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  1. Article ; Online: Venom concentrations in blisters and hemorrhagic bullae in a patient bitten by a Taiwan habu (Protobothrops mucrosquamatus)

    Chih-Chuan Lin / Po-Jung Wang / Chien-Chun Liu

    Revista da Sociedade Brasileira de Medicina Tropical, Vol 52, Iss

    2019  

    Keywords Arctic medicine. Tropical medicine ; RC955-962
    Language English
    Publishing date 2019-01-01T00:00:00Z
    Publisher Sociedade Brasileira de Medicina Tropical (SBMT)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  2. Article ; Online: Development of a Monoclonal scFv against Cytotoxin to Neutralize Cytolytic Activity Induced by Naja atra Venom on Myoblast C2C12 Cells

    Chien-Chun Liu / Cho-Ju Wu / Tsai-Ying Chou / Geng-Wang Liaw / Yung-Chin Hsiao / Lichieh-Julie Chu / Chi-Hsin Lee / Po-Jung Wang / Cheng-Hsien Hsieh / Chun-Kuei Chen / Jau-Song Yu

    Toxins, Vol 14, Iss 7, p

    2022  Volume 459

    Abstract: The Taiwanese cobra, Naja atra , is a clinically significant species of snake observed in the wild in Taiwan. Victims bitten by N. atra usually experience severe pain and local tissue necrosis. Although antivenom is available for treatment of cobra ... ...

    Abstract The Taiwanese cobra, Naja atra , is a clinically significant species of snake observed in the wild in Taiwan. Victims bitten by N. atra usually experience severe pain and local tissue necrosis. Although antivenom is available for treatment of cobra envenomation, its neutralization potency against cobra-induced necrosis is weak, with more than 60% of cobra envenoming patients developing tissue necrosis after antivenom administration. The present study found that cytotoxin (CTX) is a key component of N. atra venom responsible for cytotoxicity against myoblast cells. Anti-CTX IgY was generated in hens, and the spleens of these hens were used to construct libraries for the development of single chain variable fragments (scFv). Two anti-CTX scFv, S1 and 2S7, were selected using phage display technology and biopanning. Both polyclonal IgY and monoclonal scFv S1 reacted specifically with CTX in cobra venom. In a cell model assay, the CTX-induced cytolytic effect was inhibited only by monoclonal scFv S1, not by polyclonal IgY. Moreover, the neutralization potency of scFv S1 was about 3.8 mg/mg, approximately three times higher than that of conventional freeze-dried neurotoxic antivenom (FNAV). Collectively, these results suggest that scFv S1 can effectively neutralize CTX-induced cytotoxicity and, when combined with currently available antivenom, can improve the potency of the latter, thereby preventing tissue damage induced by cobra envenoming.
    Keywords cobra venom ; Naja atra ; cytotoxin (CTX) ; cytotoxicity ; necrosis ; single-chain variable fragment (scFv) ; Medicine ; R
    Subject code 572
    Language English
    Publishing date 2022-07-01T00:00:00Z
    Publisher MDPI AG
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  3. Article ; Online: Development of Antibody Detection ELISA Based on Immunoreactive Toxins and Toxin-Derived Peptides to Evaluate the Neutralization Potency of Equine Plasma against Naja atra in Taiwan

    Chien-Chun Liu / Yung-Chin Hsiao / Lichieh Julie Chu / Po-Jung Wang / Chien-Hsin Liu / Wen-Chin Hsieh / Jau-Song Yu

    Toxins, Vol 13, Iss 818, p

    2021  Volume 818

    Abstract: Naja atra , also known as Taiwanese cobra, is one of the most prevalent venomous snakes in Taiwan. Clinically, freeze-dried neurotoxic antivenom (FNAV) produced from horses by Taiwan Centers for Disease Control (CDC) has been the only approved treatment ... ...

    Abstract Naja atra , also known as Taiwanese cobra, is one of the most prevalent venomous snakes in Taiwan. Clinically, freeze-dried neurotoxic antivenom (FNAV) produced from horses by Taiwan Centers for Disease Control (CDC) has been the only approved treatment for N. atra envenoming for the last few decades. During antivenom production, large numbers of mice are used in the in vivo assay to determine whether the neutralization potency of hyperimmunized equines is satisfactory for large-scale harvesting. However, this in vivo assay is extremely laborious, expensive, and significantly impairs animal welfare. In the present study, we aimed to develop an in vitro ELISA-based system that could serve as an alternative assay to evaluate the neutralization potency of plasma from hyperimmunized equines. We initially obtained 51 plasma samples with known (high or low) neutralization potency assessed in vivo from 9 hyperimmunized equines and subsequently determined their antibody titers against the five major protein components of N. atra venom (neurotoxin (NTX), phospholipase A2 (PLA 2 ), cytotoxin (CTX), cysteine-rich secretory protein (CRISP), and snake venom metalloproteinase (SVMP)) via ELISA. The antibody titer against NTX was the most effective in discriminating between high and low potency plasma samples. To identify the specific epitope(s) of NTX recognized by neutralization potency-related antibodies, 17 consecutive NTX-derived pentadecapeptides were synthesized and used as antigens to probe the 51 equine plasma samples. Among the 17 peptides, immunoreactive signals for three consecutive peptides (NTX1-8, NTX1-9, and NTX1-10) were significantly higher in the high potency relative to low potency equine plasma groups ( p < 0.0001). Our ELISA system based on NTX1-10 peptide (RWRDHRGYRTERGCG) encompassing residues 28–42 of NTX displayed optimal sensitivity (96.88%) and specificity (89.47%) for differentiating between high- and low-potency plasma samples (area under the receiver operating characteristic curve (AUC) = ...
    Keywords Naja atra ; freeze-dried neurotoxic antivenom ; neutralization potency ; neurotoxin ; epitope ; peptide-based ELISA ; Medicine ; R
    Subject code 630
    Language English
    Publishing date 2021-11-01T00:00:00Z
    Publisher MDPI AG
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  4. Article ; Online: Rapid and Efficient Enrichment of Snake Venoms from Human Plasma Using a Strong Cation Exchange Tip Column to Improve Snakebite Diagnosis

    Chien-Chun Liu / Ya-Han Yang / Yung-Chin Hsiao / Po-Jung Wang / Jo-Chuan Liu / Chien-Hsin Liu / Wen-Chin Hsieh / Chih-Chuan Lin / Jau-Song Yu

    Toxins, Vol 13, Iss 2, p

    2021  Volume 140

    Abstract: Snake envenomation is a serious public health issue in many tropical and subtropical countries. Accurate diagnosis and immediate antivenom treatment are critical for effective management. However, the venom concentration in the victims’ plasma is usually ...

    Abstract Snake envenomation is a serious public health issue in many tropical and subtropical countries. Accurate diagnosis and immediate antivenom treatment are critical for effective management. However, the venom concentration in the victims’ plasma is usually low, representing one of the bottlenecks in developing clinically applicable assays for venom detection and snakebite diagnosis. In this study, we attempted to develop a simple method for rapid enrichment of venom proteins from human plasma to facilitate detection. Our experiments showed that several major protein components of both Naja atra ( N. atra ) and Bungarus multicinctus ( B. multicinctus ) venoms have higher isoelectric point (pI) values relative to high-abundance human plasma proteins and could be separated via strong cation exchange–high-performance liquid chromatography (SCX-HPLC). Based on this principle, we developed an SCX tip column-based protocol for rapid enrichment of N. atra and B. multicinctus venom proteins from human plasma. Application of liquid chromatography-tandem mass spectrometry (LC-MS/MS) led to the identification of cytotoxin and beta-bungarotoxin as the major proteins enriched by the SCX tip column in each venom sample. The entire process of venom enrichment could be completed within 10–15 min. Combination of this method with our previously developed lateral flow strip assays (rapid test) significantly enhanced the sensitivity of the rapid test, mainly via depletion of the plasma protein background, as well as increase in venom protein concentration. Notably, the SCX tip column-based enrichment method has the potential to efficiently enrich other Elapidae snake venoms containing proteins with higher pI values, thereby facilitating venom detection with other assays. This simple and rapid sample preparation method should aid in improving the clinical utility of diagnostic assays for snakebite.
    Keywords snakebite ; venom ; strong cation exchange chromatography ; lateral flow strip ; clinical diagnostic assay ; Medicine ; R
    Subject code 500
    Language English
    Publishing date 2021-02-01T00:00:00Z
    Publisher MDPI AG
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  5. Article ; Online: Development of sandwich ELISA and lateral flow strip assays for diagnosing clinically significant snakebite in Taiwan.

    Chien-Chun Liu / Jau-Song Yu / Po-Jung Wang / Yung-Chin Hsiao / Chien-Hsin Liu / Yen-Chia Chen / Pei-Fang Lai / Chih-Po Hsu / Wen-Chih Fann / Chih-Chuan Lin

    PLoS Neglected Tropical Diseases, Vol 12, Iss 12, p e

    2018  Volume 0007014

    Abstract: Taiwan is an island located in the south Pacific, a subtropical region that is home to 61 species of snakes. Of these snakes, four species-Trimeresurus stejnegeri, Protobothrops mucrosquamatus, Bungarus multicinctus and Naja atra-account for more than 90% ...

    Abstract Taiwan is an island located in the south Pacific, a subtropical region that is home to 61 species of snakes. Of these snakes, four species-Trimeresurus stejnegeri, Protobothrops mucrosquamatus, Bungarus multicinctus and Naja atra-account for more than 90% of clinical envenomation cases. Currently, there are two types of bivalent antivenom: hemorrhagic antivenom against the venom of T. stejnegeri and P. mucrosquamatus, and neurotoxic antivenom for treatment of envenomation by B. multicinctus and N. atra. However, no suitable detection kits are available to precisely guide physicians in the use of antivenoms. Here, we sought to develop diagnostic assays for improving the clinical management of snakebite in Taiwan. A two-step affinity purification procedure was used to generate neurotoxic species-specific antibodies (NSS-Abs) and hemorrhagic species-specific antibodies (HSS-Abs) from antivenoms. These two SSAbs were then used to develop a sandwich ELISA (enzyme-linked immunosorbent assay) and a lateral flow assay comprising two test lines. The resulting ELISAs and lateral flow strip assays could successfully discriminate between neurotoxic and hemorrhagic venoms. The limits of quantification (LOQ) of the ELISA for neurotoxic venoms and hemorrhagic venoms were determined to be 0.39 and 0.78 ng/ml, respectively, and the lateral flow strips were capable of detecting neurotoxic and hemorrhagic venoms at concentrations lower than 5 and 50 ng/ml, respectively, in 10-15 min. Tests of lateral flow strips in 21 clinical snakebite cases showed 100% specificity and 100% sensitivity for neurotoxic envenomation, whereas the sensitivity for detecting hemorrhagic envenomation samples was 36.4%. We herein presented a feasible strategy for developing a sensitive sandwich ELISA and lateral flow strip assay for detecting and differentiating venom proteins from hemorrhagic and neurotoxic snakes. A useful snakebite diagnostic guideline according to the lateral flow strip results and clinical symptoms was proposed to help physicians to use antivenoms appropriately. The two-test-line lateral flow strip assay could potentially be applied in an emergency room setting to help physicians diagnose and manage snakebite victims.
    Keywords Arctic medicine. Tropical medicine ; RC955-962 ; Public aspects of medicine ; RA1-1270
    Subject code 610
    Language English
    Publishing date 2018-12-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  6. Article ; Online: Analysis of the efficacy of Taiwanese freeze-dried neurotoxic antivenom against Naja kaouthia, Naja siamensis and Ophiophagus hannah through proteomics and animal model approaches.

    Chien-Chun Liu / Chen-Hsien You / Po-Jung Wang / Jau-Song Yu / Guo-Jen Huang / Chien-Hsin Liu / Wen-Chin Hsieh / Chih-Chuan Lin

    PLoS Neglected Tropical Diseases, Vol 11, Iss 12, p e

    2017  Volume 0006138

    Abstract: In Southeast Asia, envenoming resulting from cobra snakebites is an important public health issue in many regions, and antivenom therapy is the standard treatment for the snakebite. Because these cobras share a close evolutionary history, the amino acid ... ...

    Abstract In Southeast Asia, envenoming resulting from cobra snakebites is an important public health issue in many regions, and antivenom therapy is the standard treatment for the snakebite. Because these cobras share a close evolutionary history, the amino acid sequences of major venom components in different snakes are very similar. Therefore, either monovalent or polyvalent antivenoms may offer paraspecific protection against envenomation of humans by several different snakes. In Taiwan, a bivalent antivenom-freeze-dried neurotoxic antivenom (FNAV)-against Bungarus multicinctus and Naja atra is available. However, whether this antivenom is also capable of neutralizing the venom of other species of snakes is not known. Here, to expand the clinical application of Taiwanese FNAV, we used an animal model to evaluate the neutralizing ability of FNAV against the venoms of three common snakes in Southeast Asia, including two 'true' cobras Naja kaouthia (Thailand) and Naja siamensis (Thailand), and the king cobra Ophiophagus hannah (Indonesia). We further applied mass spectrometry (MS)-based proteomic techniques to characterize venom proteomes and identify FNAV-recognizable antigens in the venoms of these Asian snakes. Neutralization assays in a mouse model showed that FNAV effectively neutralized the lethality of N. kaouthia and N. siamensis venoms, but not O. hannah venom. MS-based venom protein identification results further revealed that FNAV strongly recognized three-finger toxin and phospholipase A2, the major protein components of N. kaouthia and N. siamensis venoms. The characterization of venom proteomes and identification of FNAV-recognizable venom antigens may help researchers to further develop more effective antivenom designed to block the toxicity of dominant toxic proteins, with the ultimate goal of achieving broadly therapeutic effects against these cobra snakebites.
    Keywords Arctic medicine. Tropical medicine ; RC955-962 ; Public aspects of medicine ; RA1-1270
    Subject code 616
    Language English
    Publishing date 2017-12-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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