Investigation on Cheyletoidea mites breeding in culture environment of Eupolyphaga sinensis and morphologic observation of Eucheyletia reticulata Cunliffe / 中国血吸虫病防治杂志

Objective To investigate the species of Cheyletoidea mites breeding in the culture environment of Eupolyphaga sinensis and to observe the morphology of Eucheyletia reticulata Cunliffe. Methods The soil samples from an E. sinensis farm in northern Anhui were collected. The mites in the soil samples were separated directly under a microscope and the glass speci?mens were made to observe the morphological feature of the mites under a light microscope,then the mites species were identi?fied and classified based on the morphological characteristics. Results In the culture soil of E. sinensis,totally 7 kinds of Chey?letoidea mites were isolated,namely Eucheyletia reticulata Cunliffe,Cheyletus eruditus Schrank,Cheyletus malaccensis Oudemans,Cheyletus troussarti Oudemans,Cheyletus aveisor Rohdendorz,Acaropsis sollers Rohdendorz and Cheletomor?pha lepidopterorum Shaw. They belonged to genera Eucheyletia,Cheletomorpha,Acaropsis and Cheyletus of Cheyletidae Leach family. The Eucheyletia reticulata Cunliffe was firstly found in the culture environment of E. sinensis,and its gnathosoma was large,the pedipalpal femurs were expanding and there were two strips of comb hair and two smooth bristles on the pedipal?pal tarsus,and the back of the body was covered with two pieces of tergum,which were decorated with reticular pattern. The body and foot setae were fan?shape. Conclusions There are various of Cheyletoidea mites found in the breeding environment of E. sinensis. These mites are important species for pest control in the culture environments of E. sinensis. Related measures should be taken to prevent the excessive growth of Cheyletoidea mites,so as to avoid the adverse effects on the quality and quan?tity of E. sinensis.

Cloning and purification of the first termicin-like peptide from the cockroach Eupolyphaga sinensis

Termicin is an antimicrobial peptide with six cysteines forming three disulfide bridges that was firstly isolated from the salivary glands and hemocytes of the termite Pseudacanthotermes spiniger. In contrast to many broad-spectrum antimicrobial peptides, termicin is most active against filamentous fungi. Although more than one hundred complementary DNAs (cDNAs) encoding termicin-like peptides have been reported to date, all these termicin-like peptides were obtained from Isoptera insects. Methods The cDNA was cloned by combination of cDNA library construction kit and DNA sequencing. The polypeptide was purified by gel filtration and reversed-phase high performance liquid chromatography (RP-HPLC). Its amino acid sequence was determined by Edman degradation and mass spectrometry. Antimicrobial activity was tested against several bacterial and fungal strains. The minimum inhibitory concentration (MIC) was determined by microdilution tests. Results A novel termicin-like peptide with primary structure ACDFQQCWVTCQRQYSINFISARCNGDSCVCTFRT was purified from extracts of the cockroach Eupolyphaga sinensis (Insecta: Blattodea). The cDNA encoding Es-termicin was cloned by cDNA library screening. This cDNA encoded a 60 amino acid precursor which included a 25 amino acid signal peptide. Amino acid sequence deduced from the cDNA matched well with the result of protein Edman degradation. Susceptibility test indicated that Es-termicin showed strong ability to kill fungi with a MIC of 25 g/mL against Candida albicans ATCC 90028. It only showed limited potency to affect the growth of Gram-positive bacteria with a MIC of 200 g/mL against Enterococcus faecalis ATCC 29212. It was inactive against gram-negative bacteria at the highest concentration tested (400 g/mL). Es-termicin showed high sequence similarity with termicins from many species of termites (Insecta: Isoptera). Conclusions This is the first report of a termicin-like peptide isolated from E. sinensis that belongs to the insect order Blattodea. Our results demonstrate the diversity of termicin-like peptides, as well as antimicrobial peptides in insects.

Cloning and purification of the first termicin-like peptide from the cockroach Eupolyphaga sinensis

Abstract Background Termicin is an antimicrobial peptide with six cysteines forming three disulfide bridges that was firstly isolated from the salivary glands and hemocytes of the termite Pseudacanthotermes spiniger. In contrast to many broad-spectrum antimicrobial peptides, termicin is most active against filamentous fungi. Although more than one hundred complementary DNAs (cDNAs) encoding termicin-like peptides have been reported to date, all these termicin-like peptides were obtained from Isoptera insects. Methods The cDNA was cloned by combination of cDNA library construction kit and DNA sequencing. The polypeptide was purified by gel filtration and reversed-phase high performance liquid chromatography (RP-HPLC). Its amino acid sequence was determined by Edman degradation and mass spectrometry. Antimicrobial activity was tested against several bacterial and fungal strains. The minimum inhibitory concentration (MIC) was determined by microdilution tests. Results A novel termicin-like peptide with primary structure ACDFQQCWVTCQRQYSINFISARCNGDSCVCTFRT was purified from extracts of the cockroach Eupolyphaga sinensis (Insecta: Blattodea). The cDNA encoding Es-termicin was cloned by cDNA library screening. This cDNA encoded a 60 amino acid precursor which included a 25 amino acid signal peptide. Amino acid sequence deduced from the cDNA matched well with the result of protein Edman degradation. Susceptibility test indicated that Es-termicin showed strong ability to kill fungi with a MIC of 25 μg/mL against Candida albicans ATCC 90028. It only showed limited potency to affect the growth of Gram-positive bacteria with a MIC of 200 μg/mL against Enterococcus faecalis ATCC 29212. It was inactive against gram-negative bacteria at the highest concentration tested (400 μg/mL). Es-termicin showed high sequence similarity with termicins from many species of termites (Insecta: Isoptera). Conclusions This is the first report of a termicin-like peptide isolated from E. sinensis that belongs to the insect order Blattodea. Our results demonstrate the diversity of termicin-like peptides, as well as antimicrobial peptides in insects.(AU)

Establishment and optimization of RAMP-PCR reaction system for DNA of Eupolyphaga sinensis / 中草药

Objective: To provide the basis for the genetic diversity research of Eupolyphaga sinensis by establishing and optimizing the random amplified microsatellite polymorphism RAMP-PCR reaction system and amplification procedure for genomic DNA. Methods: Phenol chloroform extraction of genomic DNA was performed on female E. sinensis. Based on RAMP Primer (ISSR807 + RAPD A5), the orthogonal test was adopted to optimize the RAMP-PCR amplification system on E. sinensis in five factors (Mg2+, dNTPs, primer, Taq DNA polymerase, and template DNA) at four levels, and the suitable anneal temperature of the primer was selected. Results: The optimal Primer (ISSR807+RAPD A5) for RAMP-PCR system (25 μL reaction volume) of E. sinensis was: 10 × PCR buffer (2.5 μL), Mg2+ (1.0 mmol/L), dNTPs (2.0 mmol/L), primer (0.5 μmol/L), Taq DNA polymerase (1 U), and template DNA (1.5 ng/μL); The optimized anneal temperature was 46.8°C. Conclusion: The established and optimized RAMP-PCR reaction system is stable and repeatable, which could provide the basis for the analysis of germplasm resources and genetic diversity of E. sinensis.

Mechanism of polypeptide extracts of Eupolyphaga sinensis Walker against oxidative aging in mice / 中国实验动物学报

Objective The purpose of this study was to investigate the mechanism of action of polypeptide extracts of Eupolyphaga sinensis Walker ( ESW) against oxidative aging.Methods Mice were intraperitoneally injected D-galac-tose for consecutive 20 days to establish an aging mouse model.The model mice were administered with different doses of ESW polypeptide (0, 40, 80, 160 mg/kg/d).The normal activity, movement and anti-stress ability of the mice were ob-served.The activity of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-PX) in blood and different tissues and the content of glutathione ( GSH) and malondialdehyde ( MDA) of the aging mice were assessed by xanthin oxidase activity measurement and spectrophotometry, respectively.The expression of nuclear factor erythroid 2-re-lated factor 2 (Nrf2) in Caco-2 cells was detected by immunofluorescence.Results Comparing the control and polypep-tide groups, there were significant decreases of body weight gain, organ indexes, anti-stress ability and activity capacity, the activity of SOD, CAT, GSH-PX and the content of GSH, and an increase of the content of MDA in blood and different tissues in the aging mice.With the increasing dose of polypeptide extracts of ESW, the body weight gain, organ indexes of the liver, spleen and kidney were significantly increased, the static and dynamic exercise time was prolonged in the poly-peptide group, and their abilities of hypoxia tolerance and heat tolerance were close to that of normal controls.The SOD, CAT, GSH-PX activity and GSH level in blood and different tissues were significantly increased, but MDA content de-creased.The expression of Nrf2 in Caco-2 cell nuclei was significantly increased in the polypeptide group, close to that of the positive control group.Conclusions The results of our study show that polypeptide extracts of ESW improve the anti-stress and antioxidative capacity in D-galactose-induced mouse models of oxidative aging by initiating Nrf2-ARE antioxidant signaling pathway, therefore, delay the oxidative aging in mice.

Study on Anticoagulant Activity in Vivo of Active Component F2-2 from Eupolyphaga seu Steleophaga / 世界科学技术-中医药现代化

Prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time (TT), plasma fibrino-gen (FIB), platelet aggregation rate and blood clots-fibrinolytic dynamic figure were taken as indexes in the evalua-tion of anticoagulant activity in vivo of active component F2-2 from Eupolyphaga seu Steleophaga. After 5 days of hypodermic injection of adrenaline, the rat model of acute blood stasis was established. Indexes were determined af-ter the model rats were treated with an intragastric administration of F2-2 for 9 days. The results showed that com-pared with the model group, PT/APTT was prolonged, FIB content was decreased, platelet aggregation rate and the largest of blood coagulation were declined after 9 days of intragastric administration in the model group. However, there was no difference on TT. It was concluded that the anticoagulant component F2-2 separated from E. seu Steleophaga showed favorable anticoagulant activity in vivo. However, its mechanism remained unknown and request-ed further researches.

The inhibitory effects and immunoregulatory activity of the fibrinolytic protein from Eupolyphaga sinensis Walker (EFP) in vivo on line H22 in mice / 中国药理学通报

Aim To explore the inhibitory effects and immunoregulatory activity of the extract from Eupolyphaga sinensis Walker (EFP) on the growth of liver cancer H22 in mice.Methods The chemical composition of extract from Eupolyphaga sinensis Walker(EFP)was separated with water extraction methods.The tumor bearing mouse model was constructed by injecting tumor cells subcutaneously.The EFP was given to the mouse by oral and hypodermic injection to observe its antitumor activity and immunoregulatory activity in vivo.The antibody level of anti-H22 in serum was measured with ELISA methods.The weight,tumor weight,spleen index,liver index were noted,and series of enzymes,such as,MDA,SOD,GPT and GOT activity of mouse serum and liver were measured.Results The extracts from Eupolyphaga sinensis Walker markedly inhibited the proliferation of tumor H22.The EFP constituent purified might be the main antitumor active part of Eupolyphaga sinensis Walker as shown by antitumor test in vitro.After treatment with EFP,the general condition of the test mice was much better than that of control mice,with spleen index,liver index increasing more obviously,antibody level of anti-H22,and MDA,SOD,GPT and GOT activity increasing more,and tumor growing more slowly.Conclusion EFP may be the main antitumor active part of Eupolyphaga sinensis Walker. EFP also has antitumor effect in vivo.EFP can exert antitumor effect in vivo by enhancing antibody level of anti-H22 and the series enzymes to be related to immunity activity.

Experimental study of Eupolyphaga Sinensis Walker on red blood cell immunity in mice with blood-deficiency / 中国免疫学杂志

Objective:To study the effect of Eupolyphaga Sinensis Walker (ESW) on red blood cell immune adherence (RCIA) in mice model with blood-deficiency by Cyclophosphamide , aimingatfurther research on the mechanism of blood stasis syndrome and promoting blood circulation and removing blood stasis.Methods:This model was made in mice by i.p. cyclophosphamide (0.01 g/kg).It has been observed that how the influence of ESW on the function of red cell immune adherence. The serum anticardiolipin antibodies (ACA),ACA-IgG?ACA-IgA and ACA-IgM levels were measured by using enzymelinked immunosorbent assay (ELISA). The mice body weight were weighted before model and after model and glandular weight were weighted. The serum levels of trace element zinc calcium were determined in healthy mice.Results:The results showed there is a decline in garland rate of red blood cell C3b receptors (RBC-C3bRR) in mice when animal model of Blood - deficiency by cyclophosphamide (0.01 g/kg) form, the serum ACA-IgG and ACA-IgA levels were markedly elevated,while mice body weight and spleen thymus weight markedly depressed. ESW (25 g/kg) increased RBC-C3bRR in the mice model with Blood-deficiency,restified the mice model body weight loss by cyclophosphamide, increased spleen thymus weight .ESW (25 g/kg) increased serum zinc calcium level in the normal mice.Conclusion:ESW increases activity of CR 1 of red blood cells and RCIA, inhibits serum ACA-IgG and ACA-IgA.ESW enhances serum zinc calcium level, that can be a good biological pharmacodynamics.