Analysis of virulence genes and antimicrobial resistance of Vibrio parahaemolyticus isolated from shellfish in 2018

Objective To analyze the virulence genes and antimicrobial resistance of 140 Vibrio parahaemolyticus strains from shellfish collected from farms and markets. Methods Virulence genes and antimicrobial resistance genes were detected by PCR amplification method, and antimicrobial resistance was determined by disk diffusion test. Results None of the 140 strains carried tdh gene, and trh gene was detected in one strain. All strains were resistant to ampicillin, some strains were resistant to cefazolin, cefuroxime sodium, streptomycin, amikacin, sulphamethoxazole/trimethoprim, tetracycline and florfenicol. All strains were sensitive to cefepime, gentamicin, imipenem, meropenem, ofloxacin and doxycycline. Six strains showed multi-antimicrobial resistance to two or more categories of antimicrobials. The resistance genes strA and strB were detected in streptomycin resistant strains, tetA was detected in tetracycline resistant strains, sul2 was detected in sulphamethoxazole/trimethoprim resistant strains, and floR was detected in florfenicol resistant strain. Conclusion Most of Vibrio parahaemolyticus strains didn't carry virulence genes and showed antimicrobial resistance to a certain extent. Some strains had more than one antimicrobial resistance gene, which exhibited multi-antimicrobial resistance. These results indicated that enhancing the monitoring of pathogenicity and antimicrobial resistance of Vibrio parahaemolyticus in aquatic products should be continued

Distribution of Marine Lipophilic Toxins in Shellfish Products Collected from the Chinese Market

To investigate the prevalence of lipophilic marine biotoxins in shellfish from the Chinese market, we used hydrophilic interaction liquid chromatography-tandem mass spectrometry (LC-MS/MS) to measure levels of okadaic acid (OA), azaspiracid (AZA1), pectenotoxin (PTX2), gymnodimine (GYM), and spirolide (SPX1). We collected and analyzed 291 shellfish samples from main production sites along a wide latitudinal transect along the Chinese coastline from December 2008 to December 2009. Results revealed a patchy distribution of the five toxins and highlighted the specific geographical distribution and seasonal and species variation of the putative toxigenic organisms. All five lipophilic marine biotoxins were found in shellfish samples. The highest concentrations of OA, AZA1, PTX2, GYM, and SPX1 were 37.3, 5.90, 16.4, 14.4, and 8.97 μg/kg, respectively. These values were much lower than the legislation limits for lipophilic shellfish toxins. However, the value might be significantly underestimated for the limited detection toxins. Also, these toxins were found in most coastal areas of China and were present in almost all seasons of the year. Thus, these five toxins represent a potential threat to human health. Consequently, studies should be conducted and measures should be taken to ensure the safety of the harvested product

Detection of Polycyclic Aromatic Hydrocarbons in Water Samples by Annular Platform-Supported Ionic Liquid-Based Headspace Liquid-Phase Microextraction

In this paper, a new method of annular platform-supported headspace liquid-phase microextraction (LPME) was designed using ionic liquid as an extraction solvent, wherein extraction stability and efficiency were improved by adding an annular platform inside the extraction bottle. The ionic liquid 1-silicyl-3-benzylimidazolehexafluorophosphate was first synthesized and proved to be an excellent extraction solvent. Coupled with liquid chromatography, the proposed method was employed to analysis of polycyclic aromatic hydrocarbons (PAHs) in water and optimized in aspects of extraction temperature, extraction solvent volume, extraction time, pH, stirring rate, and salt effect of solution. The results indicated that this method showed good linearity (R2 > 0.995) within 0.5 µg·L−1 to 1000 µg·L−1 for PAHs. The method was more suitable for extraction of volatile PAHs, with recoveries from 65.0% to 102% and quantification limits from 0.01 to 0.05 µg·L−1. It has been successfully applied for detection of PAHs in seawater samples

Methodology and application of PCR‐RFLP for species identification in tuna sashimi

Abstract The Thunnini, or tuna, comprise many species with very different commercial values. The principal raw tuna product on the market is sashimi, for which the species used is difficult to identify through conventional morphological analysis. The present study amplified the cytochrome b gene (Cytb) of 4 major tuna species used for preparing sashimi—yellowfin tuna (Thunnus albacares), southern bluefin tuna (Thunnus maccoyii), bigeye tuna (Thunnus obesus), and Atlantic bluefin tuna (Thunnus thynnus)—and 4 species commonly mislabeled as components of tuna sashimi—albacore tuna (Thunnus alalunga), skipjack tuna (Katsuwonus pelamis), striped marlin (Tetrapturus audax), and swordfish (Xiphias gladius). Polymerase chain reaction (PCR) amplicons were digested with 5 restriction enzymes—Eco147 I, Hinf I, Mbo I, Xag I, and Hind II—to obtain characteristic restriction maps of the above‐mentioned raw tuna species and the commonly mislabeled species. An identification method using PCR restriction fragment length polymorphism (PCR‐RFLP) was established and validated using 39 commercial tuna sashimi samples, which verified that this method provides results consistent with those obtained by classical sequencing. PCR‐RFLP has several advantages over classical sequencing, such as simplicity, speed and accuracy. This technique could support species identification for raw tuna and sashimi

Solid Sampling Pyrolysis Adsorption-Desorption Thermal Conductivity Method for Rapid and Simultaneous Detection of N and S in Seafood

In this work, a rapid method for the simultaneous determination of N and S in seafood was established based on a solid sampling absorption-desorption system coupled with a thermal conductivity detector. This setup mainly includes a solid sampling system, a gas line unit for controlling high-purity oxygen and helium, a combustion and reduction furnace, a purification column system for moisture, halogen, SO2, and CO2, and a thermal conductivity detector. After two stages of purging with 20 s of He sweeping (250 mL/min), N2 residue in the sample-containing chamber can be reduced to 2, H2O, and CO2 were absorbed by the adsorption column in turn, the purification process executed the vaporization of the N-containing analyte, and then N2 was detected by the thermal conductivity cell for the quantification of N. Subsequently, the adsorbed SO2 was released after heating the SO2 adsorption column and then transported to the thermal conductivity cell for the detection and quantification of S. After the instrumental optimization, the linear range was 2.0–100 mg and the correlation coefficient (R) was more than 0.999. The limit of detection (LOD) for N was 0.66 μg and the R was less than 4.0%, while the recovery rate ranged from 95.33 to 102.8%. At the same time, the LOD for S was 2.29 μg and the R was less than 6.0%, while the recovery rate ranged from 92.26 to 105.5%. The method was validated using certified reference materials (CRMs) and the measured N and S concentrations agreed with the certified values. The method indicated good accuracy and precision for the simultaneous detection of N and S in seafood samples. The total time of analysis was less than 6 min without the sample preparation process, fulfilling the fast detection of N and S in seafood. The establishment of this method filled the blank space in the area of the simultaneous and rapid determination of N and S in aquatic product solids. Thus, it provided technical support effectively to the requirements of risk assessment and detection in cases where supervision inspection was time-dependent