Antibacterial activity from Siamese crocodile (Crocodylus siamensis) serum

Antibacterial agents were purified from Siamese crocodile serum by anion exchange, gel filtration and reversed phase HPLC. Six antibacterial agents designed as Hp14, Hp15, Hp17, Hp31, Hp36 and Hp51 were purified and proved to carry activity against Salmonella typhi, Escherichia coli, Staphylococcus aureus, Staphylococcus epidermidis, Klebsiella pneumoniae, Pseudomonas aeruginosa and Vibrio chorelae. The mass analysis of MALDI-TOF for antibacterial agent of Hp14, Hp15 and Hp51 revealed that they are small molecule with a molecular mass less than 1 kDa. The scanning electron microscopy demonstrated that these agents targeted the bacterial membrane and they act like as antimicrobialpeptides. The antibacterial agent in the serum may represent the first line of an immune system in a freshwater crocodile

Immunization of Basa fish (Pangasius bocourti) against Ichthyophthirius multifiliis with live and sonicated trophonts

The high density of Basa fish (Pangasius bocourti) culture leads to outbreaks of Ichthyophthirius multifiliis (Ich), also knows as white spot disease. In this research, immunization of Basa fish against Ich with live and sonicated trophonts by intraperitoneal (IP) injection was investigated. Anti-Ich antibody titer was determined using ELISA and Western immunoblotting 21 days post immunization. The results revealed that pre-immunized fish, non-immunized fish and fish immunized with bovine serum albumin (BSA) at a concentration 65 µg g -1 fish did not show specific antibody against Ich. 21 days post immunization, fish immunized with live trophonts exhibited higher anti-Ich antibody titer than fish immunized with sonicated trophonts at the same antigen concentration. Fish immunized with 65 µg trophonts protein/g fish live trophonts showed the highest titer 1:1,000 (p<0.05). The results from Western immunoblotting showed two parasite protein bands of 66 kDa and <14 kDa, which reacted with antibodies from serum of immune fish. No fish in the non-immunized group survived. At the same concentration of antigen (65 µg g^-1), fish immunized with live trophonts exhibited the highest survival rate, 63.33±5.77% (p<0.05). Therefore, these results are the Basa fish immunizing procedure will be the way to conduct immunization against Ich to prevent disease outbreaks in aquaculture

CpG-island methylation study of liver fluke-related cholangiocarcinoma

Background: Genetic changes have been widely reported in association with cholangiocarcinoma (CCA), while epigenetic changes are poorly characterised. We aimed to further evaluate CpG-island hypermethylation in CCA at candidate loci, which may have potential as diagnostic or prognostic biomarkers. Methods: We analysed methylation of 26 CpG-islands in 102 liver fluke related-CCA and 29 adjacent normal samples using methylation-specific PCR (MSP). Methylation of interest loci was confirmed using pyrosequencing and/or combined bisulfite restriction analysis, and protein expression by immunohistochemistry. Results: A number of CpG-islands (OPCML, SFRP1, HIC1, PTEN and DcR1) showed frequency of hypermethylation in >28% of CCA, but not adjacent normal tissues. The results showed that 91% of CCA were methylated in at least one CpG-island. The OPCML was the most frequently methylated locus (72.5%) and was more frequently methylated in less differentiated CCA. Patients with methylated DcR1 had significantly longer overall survival (Median; 41.7 vs 21.7 weeks, P=0.027). Low-protein expression was found in >70% of CCA with methylation of OPCML or DcR1. Conclusion: Aberrant hypermethylation of certain loci is a common event in liver fluke-related CCA and may potentially contribute to cholangiocarcinogenesis. The OPCML and DcR1 might serve as methylation biomarkers in CCA that can be readily examined by MSP

Characterization of the allergen Sol gem 2 from the fire ant venom, Solenopsis geminata

Sol i 2 is a potent allergen in Solenopsis invicta venom, and most humans exhibit reactivity to it. The Sol gem 2 allergen found in the venom of the Thai tropical fire ant Solenopsis geminata was analysed in the present study. The protein was present in higher amounts than other proteins, as determined by SDS-PAGE, and presumably has allergenic properties similar to those of Sol i 2. Sol gem 2 molecular weight is 28 and 15 kDa, respectively, under non-reducing and reducing conditions, indicating that its native form is a dimer. LC-MS/MS analysis confirmed its similarity to Sol i 2. The mono/dimeric form of Sol gem 2 was determined to be relevant by proteomic approach and immunoblotting. An anti-Sol gem 2 antibody was produced in mice, with a titer greater than 1:800 according to the Western blotting analysis. The Sol gem 2-neutralising activity of this antibody was determined in crickets. The paralytic dose 50 (PD50) of crude S. geminata venom was elevated from 0.18 mg/g of body weight to more than 0.90 mg/g of body weight after preincubation with antibody at a ratio of 1:1. These results suggest that Sol gem 2 plays an important role in mediating the effects of the piperidine derivatives in the venom

Gallic acid conjugated with gold nanoparticles: antibacterial activity and mechanism of action on foodborne pathogens

Narintorn Rattanata,1 Sompong Klaynongsruang,1 Chanvit Leelayuwat,2 Temduang Limpaiboon,2 Aroonlug Lulitanond,2 Patcharee Boonsiri,3 Sirinart Chio-Srichan,4 Siriwat Soontaranon,4 Supagorn Rugmai,4 Jureerut Daduang2 1Department of Biochemistry, Faculty of Science, 2Centre for Research and Development of Medical Diagnostic Laboratories, Faculty of Associated Medical Sciences, 3Department of Biochemistry, Faculty of Medicine, Khon Kaen University, Khon Kaen, 4Synchrotron Light Research Institute (Public Organization), Nakhon Ratchasima, Thailand Abstract: Foodborne pathogens, including Plesiomonas shigelloides and Shigella flexneri B, are the major cause of diarrheal endemics worldwide. Antibiotic drug resistance is increasing. Therefore, bioactive compounds with antibacterial activity, such as gallic acid (GA), are needed. Gold nanoparticles (AuNPs) are used as drug delivery agents. This study aimed to conjugate and characterize AuNP&ndash;GA and to evaluate the antibacterial activity. AuNP was conjugated with GA, and the core&ndash;shell structures were characterized by small-angle X-ray scattering and transmission electron microscopy. Antibacterial activity of AuNP&ndash;GA against P. shigelloides and S. flexneri B was evaluated by well diffusion method. AuNP&ndash;GA bactericidal mechanism was elucidated by Fourier transform infrared microspectroscopic analysis. The results of small-angle X-ray scattering showed that AuNP&ndash;GA conjugation was successful. Antibacterial activity of GA against both bacteria was improved by conjugation with AuNP because the minimum inhibitory concentration value of AuNP&ndash;GA was significantly decreased (P&lt;0.0001) compared to that of GA. Fourier transform infrared analysis revealed that AuNP&ndash;GA resulted in alterations of lipids, proteins, and nucleic acids at the bacterial cell membrane. Our findings show that AuNP&ndash;GA has potential for further application in biomedical sciences.Keywords: gold nanoparticles, gallic acid, antibacterial activity, foodborne bacteria, small-angle X-ray scattering (SAXS)&nbsp

Expanding the allergen repertoire of salmon and catfish

Background: Diagnostic tests for fish allergy are hampered by the large number of under‐investigated fish species. Four salmon allergens are well‐characterized and registered with the WHO/IUIS while no catfish allergens have been described so far. In 2008, freshwater‐cultured catfish production surpassed that of salmon, the globally most‐cultured marine species. We aimed to identify, quantify, and compare all IgE‐binding proteins in salmon and catfish. Methods: Seventy‐seven pediatric patients with clinically confirmed fish allergy underwent skin prick tests to salmon and catfish. The allergen repertoire of raw and heated protein extracts was evaluated by immunoblotting using five allergen‐specific antibodies and patients' serum followed by mass spectrometric analyses. Results: Raw and heated extracts from catfish displayed a higher frequency of IgE‐binding compared to those from salmon (77% vs 70% and 64% vs 53%, respectively). The major fish allergen parvalbumin demonstrated the highest IgE‐binding capacity (10%‐49%), followed by triosephosphate isomerase (TPI; 19%‐34%) in raw and tropomyosin (6%‐32%) in heated extracts. Six previously unidentified fish allergens, including TPI, were registered with the WHO/IUIS. Creatine kinase from salmon and catfish was detected by IgE from 14% and 10% of patients, respectively. Catfish L‐lactate dehydrogenase, glyceraldehyde‐3‐phosphate dehydrogenase, pyruvate kinase, and glucose‐6‐phosphate isomerase showed IgE‐binding for 6%‐13% of patients. In salmon, these proteins could not be separated successfully. Conclusions: We detail the allergen repertoire of two highly farmed fish species. IgE‐binding to fish tropomyosins and TPIs was demonstrated for the first time in a large patient cohort. Tropomyosins, in addition to parvalbumins, should be considered for urgently needed improved fish allergy diagnostics

Expanding the allergen repertoire of salmon and catfish

Background: Diagnostic tests for fish allergy are hampered by the large number of under‐investigated fish species. Four salmon allergens are well‐characterized and registered with the WHO/IUIS while no catfish allergens have been described so far. In 2008, freshwater‐cultured catfish production surpassed that of salmon, the globally most‐cultured marine species. We aimed to identify, quantify, and compare all IgE‐binding proteins in salmon and catfish. Methods: Seventy‐seven pediatric patients with clinically confirmed fish allergy underwent skin prick tests to salmon and catfish. The allergen repertoire of raw and heated protein extracts was evaluated by immunoblotting using five allergen‐specific antibodies and patients' serum followed by mass spectrometric analyses. Results: Raw and heated extracts from catfish displayed a higher frequency of IgE‐binding compared to those from salmon (77% vs 70% and 64% vs 53%, respectively). The major fish allergen parvalbumin demonstrated the highest IgE‐binding capacity (10%‐49%), followed by triosephosphate isomerase (TPI; 19%‐34%) in raw and tropomyosin (6%‐32%) in heated extracts. Six previously unidentified fish allergens, including TPI, were registered with the WHO/IUIS. Creatine kinase from salmon and catfish was detected by IgE from 14% and 10% of patients, respectively. Catfish L‐lactate dehydrogenase, glyceraldehyde‐3‐phosphate dehydrogenase, pyruvate kinase, and glucose‐6‐phosphate isomerase showed IgE‐binding for 6%‐13% of patients. In salmon, these proteins could not be separated successfully. Conclusions: We detail the allergen repertoire of two highly farmed fish species. IgE‐binding to fish tropomyosins and TPIs was demonstrated for the first time in a large patient cohort. Tropomyosins, in addition to parvalbumins, should be considered for urgently needed improved fish allergy diagnostics