In vivo antibacterial activity of medicinal plant Sophora flavescens against Streptococcus agalactiae infection.

Streptococcosis disease caused by Streptococcus agalactiae (Group B Streptococcus, GBS) results in a huge economic loss of tilapia culture. It is urgent to find new antimicrobial agents against streptococcosis. In this study, 20 medicinal plants were evaluated in vitro and in vivo to obtain medicinal plants and potential bioactive compounds against GBS infection. The results showed that the ethanol extracts of 20 medicinal plants had low or no antibacterial properties in vitro, with a minimal inhibitory concentration ≥256 mg/L. Interestingly, in vivo tests showed that 7 medicinal plants could significantly inhibit GBS infection in tilapia, and Sophora flavescens (SF) had the strongest anti-GBS activity in tilapia, reaching 92.68%. SF could significantly reduce the bacterial loads of GBS in different tissues (liver, spleen and brain) of tilapia after treated with different tested concentrations (12.5, 25.0, 50.0 and 100.0 mg/kg) for 24 h. Moreover, 50 mg/kg SF could significantly improve the survival rate of GBS-infected tilapia by inhibiting GBS replication. Furthermore, the expression of antioxidant gene cat, immune-related gene c-type lysozyme and anti-inflammatory cytokine il-10 in liver tissue of GBS-infected tilapia significantly increased after treated with SF for 24 h. Meanwhile, SF significantly reduced the expression of immune-related gene myd88 and pro-inflammatory cytokines il-8 and il-1ß in liver tissue of GBS-infected tilapia. The negative and positive models of UPLC-QE-MS, respectively, identified 27 and 57 components of SF. The major components of SF extract in the negative model were α, α-trehalose, DL-malic acid, D- (-)-fructose and xanthohumol, while in the positive model were oxymatrine, formononetin, (-)-maackiain and xanthohumol. Interestingly, oxymatrine and xanthohumol could significantly inhibit GBS infection in tilapia. Taken together, these results suggest that SF can inhibit GBS infection in tilapia, and it has potential for the development of anti-GBS agents.

Natural component geniposide enhances survival rate of crayfish Procambarus clarkii infected with white spot syndrome virus.

White Spot Disease (WSD), caused by white spot syndrome virus (WSSV), is an acute and highly lethal viral disease of shrimp. Currently, there are no commercially available drugs to control WSD. It is urgent and necessary to find anti-WSSV drugs. Natural compounds are an important source of antiviral drug discovery. In this study, the anti-WSSV activity of natural compound geniposide (GP) was investigated in crayfish Procambarus clarkii. Results showed that GP had a concentration-dependent inhibitory effect on WSSV replication in crayfish at 24 h, and highest inhibition was more than 98%. In addition, GP significantly inhibited the expression of WSSV immediate-early gene ie1, early gene DNApol, late gene VP28. The mortality of WSSV-infected crayfish in control groups was 100%, while it reduced by 70.0% when treated with 50 mg/kg GP. Co-incubation, pre-treatment and post-treatment experiments showed that GP could prevent and treat WSSV infection in crayfish by significantly inhibiting WSSV multiplication. Mechanistically, the syntheses of WSSV structural proteins VP19, VP24, VP26 and VP28 were significantly inhibited by GP in S2 cells. Furthermore, GP could also suppress WSSV replication by blocking the expression of antiviral immunity-related factor STAT to reduce ie1 transcription. Moreover, GP possessed anti-inflammatory and anti-oxidative activity in crayfish. Overall, GP has the potential to be developed as a preventive or therapeutic agent against WSSV infection.

Natural component plumbagin as a potential antibacterial agent against Streptococcus agalactiae infection.

Streptococcus agalactiae, also known as Group B Streptococcus (GBS), can infect humans, terrestrial animals and fish. The emergence of bacterial resistance of S. agalactiae to antibiotics leads to an urgent need of exploration of new antimicrobial agents. In the study, the antibacterial activity of natural component plumbagin (PLB) against S. agalactiae was investigated in vitro and in vivo. The results showed that the minimal inhibitory concentration (MIC) of PLB against S. agalactiae was 8 mg/L. The growth curve assay revealed that PLB could inhibit the growth of S. agalactiae. In addition, the time-killing curve showed that S. agalactiae was killed almost completely by 2-fold MIC of PLB within 12 h. Transmission electron microscopy results showed obvious severe morphological destruction and abnormal cells of S. agalactiae after treated with PLB. The pathogenicity of S. agalactiae to zebrafish was significantly decreased after preincubation with PLB for 2 h in vitro, further indicating the bactericidal activity of PLB. Interestingly, PLB could kill S. agalactiae without inducing resistance development. Furthermore, pretreatment and post-treatment assays suggested that PLB also exhibited the antibacterial activity against S. agalactiae infection in vivo by effectively reducing the bacterial load and improving the survival rate of S. agalactiae-infected zebrafish. In summary, PLB had potent antibacterial activity against S. agalactiae in vitro and in vivo, and it could be an excellent antimicrobial candidate to prevent and control S. agalactiae infection.

Evaluation on the antiviral activity of genipin against white spot syndrome virus in crayfish.

White spot syndrome virus (WSSV) is a serious epidemic pathogen of crustaceans and cause severe economic losses to aquaculture. However, no commercial drugs presently available to control WSSV infection. Genipin (GN) is a bioactive compound extracted from the fruit of Gardenia jasminoides and exhibits potential antiviral activity. In the study, the antiviral activity of GN against WSSV was investigated in crayfish Procambarus clarkii and in shrimp Litopenaeus vannamei. In vitro antiviral test showed that GN could inhibit WSSV replication in crayfish and in shrimp, and the highest inhibition on WSSV was over 99% when treatment with 50 mg/kg of GN for 24 h. In vivo antiviral test proved that GN could be used to treat and prevent WSSV infection. GN could also effectively protect crayfish from WSSV infection by reducing the mortality rate of WSSV-infected crayfish. Moreover, GN attenuated the WSSV-induced oxidative stress and inflammatory by upregulation the expression of antioxidant-related genes and downregulation the expression of inflammatory-related genes, respectively. Mechanically, GN inhibited WSSV replication at least via decreasing STAT (signal transducer and activator of transcription) gene expression to block WSSV immediate-early gene ie1 transcription. Additionally, the inhibition of BI-1 (Bax inhibitor-1) gene expression also played an important role in the suppression of WSSV infection. In conclusion, GN represented a potential therapeutic and preventive agent to block WSSV infection.

Immunosuppression-induced alterations in fish gut microbiota may increase the susceptibility to pathogens.

Intestinal bacteria play an important role in the health and provide a variety of beneficial effects to host. Immunosuppressant can reduce the immunity of host and increase the susceptibility to pathogens. But it is not clear whether the increased susceptibility caused by immunosuppressant is related to changes of gut microbiota. In this study, we used crucian carp administrated with dexamethasone to explore the effects of immunosuppressants on gut microbial communities and further evaluate the potential association between changes in gut microbiota and susceptibility to pathogens. The results of MANOVA based on the top 10 PCoA axis scores from unweighed/weighted UniFrac distances showed that administration of dexamethasone (P = 0.021) and the administration time (P = 0.027) had a significant impact on the gut microbial composition, regardless of pathogens infection status (P = 0.35). After administration with dexamethasone, the fish had higher abundance of Cetobacterium and lower abundance of Bacillus and Lactococcus, and the abundance of genus Bacillus, Pseudomonas and Lactococcus decreased along with prolong administration time of dexamethasone. The results may help us understand the correlation between the host susceptibility to pathogenic bacteria and gut microbial community shift, and extend our knowledge regarding the role of gut microbiota in keeping the balance between pathogenic and symbiotic bacteria.

Screening of plant extracts for anthelmintic activity against Dactylogyrus intermedius (Monogenea) in goldfish (Carassius auratus).

With the aim of finding natural anthelmintic agents against Dactylogyrus intermedius (Monogenea) in goldfish (Carassius auratus), 26 plants were screened for antiparasitic properties using in vivo anthelmintic efficacy assay. The results showed that Caesalpinia sappan, Lysima chiachristinae, Cuscuta chinensis, Artemisia argyi, and Eupatorium fortunei were found to have 100% anthelmintic efficacy at 125, 150, 225, 300, and 500 mg L(-1) after 48 h of exposure. Crude extract of the five plants were further partitioned with petroleum ether, chloroform, ethyl acetate, methanol, and water to obtain anthelmintically active fractions with various polarity. Among these fractions tested, the ethyl acetate extract of L. chiachristinae was found to be the most effective with a 50% effective concentration (EC50) value of 5.1 mg/L after 48 h of exposure. This was followed by ethyl acetate extract of C. chinensis (48 h-EC50 = 8.5 mg L(-1)), chloroform extracts of C. sappan (48 h-EC50 = 15.6 mg L(-1)), methanol extract of C. chinensis (48 h-EC50 = 15.9 mg L(-1)), and chloroform and petroleum ether extract of L. chiachristinae (EC50 values of 17.2 and 21.1 mg/L, respectively), suggesting that these plants, as well as the active fractions, provide potential sources of botanic drugs for the control of D. intermedius in aquaculture.

Application of Virus Targeting Nanocarrier Drug Delivery System in Virus-Induced Central Nervous System Disease Treatment.

Virus-induced central nervous system (CNS) diseases represent a significant burden to animal health worldwide. The difficulty in treating these diseases is mainly attributable to the elaborate barrier system, which limits the transport of drugs to the infected sites. Therefore, it is necessary to develop smart delivery technologies for treatment of these diseases. In the study, viral nervous necrosis disease was studied as a model to evaluate the feasibility of multiwalled carbon nanotubes (MWCNTs) conjugated with virus-specific nanobody and antiviral drug for targeted therapy of virus-induced CNS diseases. The virus (named as PGNNV) was isolated, identified and purified from diseased grouper. A naïve phage-displayed alpaca nanobody library was constructed, and the purified PGNNV was used for biopanning of PGNNV-specific nanobody from the library. The targeted delivery system based on MWCNTs conjugated with polyethylenimine, ribavirin, and PGNNV-specific nanobody was constructed and designated as MWCNTs-PEI-R-Nb. Targeting ability and treatment effects of the MWCNTs-PEI-R-Nb were checked both in vitro and in vivo. MWCNTs-PEI-R-Nb showed an increasing distribution in PGNNV-infected cells, and an obvious accumulation in the brain of PGNNV-infected zebrafish larvae. MWCNTs-PEI-R-Nb also showed a strong anti-PGNNV ability both in vitro and in vivo. The mortality of larvae treated with MWCNTs-PEI-R-Nb (equivalent to 100 mg/L ribavirin) was 27% during 10 days post infection, whereas it was 100% for the control group. The results so far indicate that MWCNTs conjugated with antiviral drugs and viral-specific antibody are effective means for virus-induced CNS disease targeted therapy.

Enhanced protective immunity against spring viremia of carp virus infection can be induced by recombinant subunit vaccine conjugated to single-walled carbon nanotubes.

As a successful prevention strategy for controlling the highly contagious and pathogenic disease of spring viremia of carp (SVC), DNA vaccines reported in recent years could trigger protective responses against SVC with the means of injection. However, there remains many concerns and uncertainties related to DNA vaccination as well as injection is labor intensive, costly and not suitable to vaccinate large numbers of fish. Therefore, more efficient and safe prophylactic measures should be urgently investigated. In this research, single-walled carbon nanotubes (SWCNTs) as the candidate SWCNTs-pET32a-G subunit vaccine carrier were administrated via bath (1, 5, 10, 20, 40 mg L-1) or injection (1, 4, 8, 12, 20 µg) in common carp juvenile, and the different immune treatments to induce immunoprotective effect were analyzed. The results showed that SWCNTs-pET32a-G could enter fish body after immersion for 10 h, furthermore, compared to control groups, antibody levels, the non-specific immune parameters (complement activity, superoxide dismutase activity and alkaline phosphatase activity), and immune-related genes (especially the TNF-α and IFNg2b) in vaccinated groups were significantly enhanced in fish immunized with SWCNTs-subunit vaccine. In addition, as a promising carrier, SWCNTs can increase the immune protective effect of naked subunit vaccine by ca. 16% in bath immunization group and by ca. 23% in injection group. This study suggests that SWCNTs-vaccine may represent a potentially efficient immersion vaccine against viral pathogens of fish in the future.

The oxidative stress response of myclobutanil and cyproconazole on Tetrahymena thermophila.

Using Tetrahymena thermophila as experimental models, the oxidative stress of triazole fungicides myclobutanil (MYC) and cyproconazole (CYP) was investigated. Results showed that 24-h EC50 values for MYC and CYP were 16.67 (13.37-19.65) and 20.44 (18.85-21.96) mg/L, respectively; 48-h EC50 values for MYC and CYP were 14.31 (13.13-15.42) and 18.76 (17.09-20.31) mg/L, respectively. Reactive oxygen species was significantly induced and cytotoxicity was caused by MYC and CYP by increasing propidium iodide (PI) fluorescence. Damage of regular wrinkles and appearing of small holes on the cell surface were observed by SEM. Furthermore, MYC and CYP also caused notable changes in enzyme activities and mRNA levels. Overall, the present study points out that MYC and CYP lead to oxidative stress on T. thermophila. The information presented in this study will provide insights into the mechanism of triazoles-induced oxidative stress on T. thermophila.

[The surgical timing and effects for vitreous hemorrhage caused by ocular blunt trauma].

OBJECTIVE: To investigate the surgical timing and effects for vitreous hemorrhage caused by blunt ocular trauma. METHOD: 116 eyes of vitreous hemorrhage caused by ocular blunt trauma were divided into two groups, non-vitrectomy group (46 eyes) and vitrectomy group (70 eyes). The treating results and follow-up results were compared. RESULTS: 50 of 70 eyes (71.4%) in vitrectomy group and 10 of 46 eyes (21.7%) in non-vitrectomy group achieved a visual acuity of 0.1 or better over a follow-up period of 1 month, which indicated a significant difference between two groups. In the non-vitrectomy group, visual acuity of 10 eyes were better than 0.1, 7 eyes (70.0%) of them improved within 2 weeks of injury. In surgery group, 27.1% underwent only vitrectomy and 70.9% needed combining with other manipulations. In vitrectomy group the retina detachment was found in 3 eyes (4.3%) post-operatively and were cured by second operation. In non-vitrectomy group, the retinal detachment was found in 14 eyes (30.4%) during the follow-up period, and 8 eyes (72.7%) of 11 eyes received operation were cured. CONCLUSION: Early vitreous surgery can improve curative effect for vitreous hemorrhage caused by blunt ocular trauma when no improvement was observed after applied drugs 2 - 3 weeks.

[On the thinking of medical psychology in Pre-Qin Period].

There are abundant medical psychological thoughts in the ancient Chinese medical and cultural heritage. Based on the time of appearance of the above materials, it indicates that China is one of the earliest countries where medical psychology originates. There are systematic recognition and descriptions on the psychological factors as the etiology of diseases, and achievements on psychotherapy as well in the Pre-Qin Period. All these thoughts and achievements are basically identical with research of modern medical psychology. The theories are essential experiences of our ancestors about two thousand years ago. Today, these theories still play a significant role in guiding the clinical practices and scientific researches.