A Comparative Study on the Muscle and Gut Microbiota of Opsariichthys bidens from Rice Field and Pond Culture Breeding Modes.

To investigate difference in the quality of the different parts (back, tail muscles, and fish skin) of Opsariichthys bidens from pond and rice field cultures, a comparative study was conducted in terms of nutritional composition, volatile flavor profiles and gut microbiota. In detail, the texture, free amino acids, fatty acids were further assessed. The results suggested that the moisture content, crude protein and crude fat content in the skin of O. bidens are higher than those in the back and tail muscles, regardless of breeding modes. The fish cultured in the rice field had a higher protein content than those from the pond culture, while the fat content of the rice field-cultured fish was significantly low compared to the fish from the pond culture, especially in the back and tail parts. A total of 43 volatile components were detected by Gas Chromatography-Mass Spectrometry (GC-MS), with a maximum of 18 types of aldehydes and the highest concentration being nonanal. Compared to pond cultures, the fish from the rice field cultures showed more abundant flavor composition and odor-active compounds. The total content of DHA (Docosahexaenoic Acid) and EPA (Eicosapentaenoic Acid) in the rice field-cultured fish was higher than that of the pond group, while no significant disparity in amino acid composition was observed (p > 0.05). Comparative and clustering analyses of gut microbiota revealed notable discrepancies in the gut microbiota of O. bidens from two aquaculture systems. However, an inherent correlation between the gut microbiome and meat quality would be further emphasized in further studies. This study can offer a theoretical reference for the development of high-quality aquatic products by selecting the appropriate aquaculture models.

Genomic presence of gadD1 glutamate decarboxylase correlates with the organization of ascB-dapE internalin cluster in Listeria monocytogenes.

The ability to survive and proliferate in acidic environments is a prerequisite for the infection of Listeria monocytogenes. The glutamate decarboxylase (GAD) system is responsible for acid resistance, and three GAD homologs have been identified in L. monocytogenes: gadD1, gadD2, and gadD3. To examine whether GAD genes are specific to lineage, serovar, or certain subpopulation, we performed a systematic investigation on the prevalence of GAD genes in 164 L. monocytogenes. In contrast to gadD2 and gadD3 conserved in all L. monocytogenes strains, gadD1 was identified in 36.6% (60/164) of L. monocytogenes strains, including all serovar 1/2c and 68.5% (37/54) of serovar 1/2a strains, as well as a small fraction of serovar 1/2b (3.4%, 1/29) and lineage III (13.8%, 4/29) strains. All serovar 4b and lineage IV strains lacked this gene. According to the ascB-dapE structure, L. monocytogenes strains were classified into four subpopulations, carrying inlC2DE, inlGC2DE, inlGHE, or no internalin cluster, respectively. All L. monocytogenes strains with inlGC2DE or inlGHE pattern harbored gadD1, whereas those bearing inlC2DE or no internalin cluster between ascB and dapE lacked gadD1. In addition, other five non-monocytogenes Listeria species lacking ascB-dapE internalin cluster were gadD1-negative. Overall, the presence of gadD1 is not fully dependent on lineages or serovars but correlates with ascB-dapE internalin profiles, suggesting gadD1 might have co-evolved with the ascB-dapE internalin cluster in the primitive L. monocytogenes before divergence of serovars.

[Rapid screening and confirmation of 86 illegally added chemicals in fishery drugs by ultra performance liquid chromatography-quadrupole-time-of-flight mass spectrometry].

With the rapid expansion of fisheries, one of the most significant limitations to the sustainable development of fisheries in China is the quality and safety of fishery products owing to the abuse of fishery drugs and the use of illegal and/or restricted chemicals in fishery drugs. A range of chemicals that are potential hazards to fishery drugs were selected for screening in this study. A comprehensive analytical method was developed, based on ultra performance liquid chromatography-quadrupole-time-of-flight mass spectrometry (UPLC-Q-TOF-MS), for the rapid screening of 86 types of illegally added chemicals in fishery drugs. The fishery drug samples were extracted with 80% (v/v) acetonitrile aqueous solution and diluted to reduce matrix effects. The 86 target compounds were separated on an ACQUITY PREMIER HSS T3 column (100 mm×2.1 mm, 1.8 µm), with methanol and 0.1% formic acid as mobile phases, via gradient elution. The extract was directly analyzed by UPLC-Q-TOF-MS using electrospray ionization in the positive mode. The external standard method was used for quantification. In this study, the extraction reagent and purification procedure were selected to develop a simple and effective pre-treatment protocol. The effects of the chromatographic column, mobile phase, and fragmentation voltage on the separation and sensitivity of the 86 substances were evaluated to determine the optimum instrument conditions. An accurate mass database and fragment ion library were created for the rapid qualitative and quantitative analysis of the 86 illegally added chemicals in fishery drugs. The retention time, isotopic abundance and spacing, and precise mass of the principal diagnostic ion for each analyte were used for identification. The information on the fragment ions obtained from the target MS/MS profiles was compared with that from a database to ensure the accuracy of the qualitative results. The chromatographic peak area of each target analyte was used for quantification. The analytical detection was based on the retention time deviation of ±0.35 min, accurate mass deviation of ±10×10-6, and major adduct forms, including [M+H]+, [M+Na]+, and [M+NH4]+. To evaluate the matrix effects of the 86 target chemicals at varied dilution ratios, two types of antibiotics and four types of Chinese herbal medicines were selected as typical samples. Considering the instrument tolerance as well as sensitivity and accuracy of the procedure, the recommended dilution ratios for antibiotics and Chinese herbal medicines were 50 times and 10 times, respectively. Two different types of calibration curves were prepared; one was the solvent calibration curve for antibiotics and the other was the matrix calibration curve for Chinese herbal medicines. For a given concentration, the calibration curves of the 86 target chemicals were linear with correlation coefficients of at least 0.99. The recoveries ranged from 76.8% to 112.1% with relative standard deviations (RSDs) (n=3) of less than 11.7%. The limit of quantification (LOQ) ranges of the compounds in Chinese herbal medicines and antibiotics were 1-15 mg/kg and 5-75 mg/kg, respectively. To evaluate the screening detection limits (SDLs) of each compound, a mixed standard solution was added to a fishery drug sample at varied concentrations. The SDL ranges of the compounds in Chinese herbal medicines and antibiotics were 1-15 and 5-50 mg/kg, respectively. This approach resulted in SDLs that satisfy the actual screening requirements. Because of its rapid nature, simplicity, accuracy, and sensitivity, the method may be used in the high-throughput screening and identification of illegally added chemicals in many types of fishery drugs. This method was applied to a monitoring project for the quality and safety of fishery inputs in Zhejiang Province. Sixty fishery drug samples were evaluated, among which eight Chinese herbal medicine samples were found to contain unspecified ingredients and one antibiotic sample was found to be free of any active substances. Thus, an effective technical method to monitor the quality and safety of fishery drugs was developed.

Genetic organization of ascB-dapE internalin cluster serves as a potential marker for Listeria monocytogenes sublineages IIA, IIB, and IIC.

Listeria monocytogenes is an important foodborne pathogen that comprises four genetic lineages: I, II, III, and IV. Of these, lineage II is frequently recovered from foods and environments and responsible for the increasing incidence of human listeriosis. In this study, the phylogenetic structure of lineage II was determined through sequencing analysis of the ascB-dapE internalin cluster. Fifteen sequence types proposed by multilocus sequence typing based on nine housekeeping genes were grouped into three distinct sublineages, IIA, IIB, and IIC. Organization of the ascBdapE internalin cluster could serve as a molecular marker for these sublineages, with inlGHE, inlGC2DE, and inlC2DE for IIA, IIB, and IIC, respectively. These sublineages displayed specific genetic and phenotypic characteristics. IIA and IIC showed a higher frequency of recombination (rho/theta). However, recombination events had greater effect (r/m) on IIB, leading to its high nucleotide diversity. Moreover, IIA and IIB harbored a wider range of internalin and stress-response genes, and possessed higher nisin tolerance, whereas IIC contained the largest portion of low-virulent strains owing to premature stop codons in inlA. The results of this study indicate that IIA, IIB, and IIC might occupy different ecological niches, and IIB might have a better adaptation to a broad range of environmental niches.