Survey of the authenticity of prawn and shrimp species in commercial food products by PCR-RFLP analysis of a 16S rRNA/tRNA mitochondrial region

A novel PCR-RFLP method was evaluated as a tool to assess the incidence of incorrect labelling of prawns and shrimps in commercial food products. The whole method can be performed in less than 8 h in only one day of work. PCR amplification with primers 16Scru4/16Scru3, targeted to the amplification of a ca. 530 bp region of 16S rRNA and tRNA^Val mitochondrial genes, was coupled to restriction analysis with AluI, TaqI or HinfI. Forty-one commercial food products were considered. The molecular method considered allowed the identification of up to 17 different prawn and shrimp species in all the processed products considered. Seven (28%) of the 25 food products declaring one or more species in their labels were incorrectly labelled. Authentication was successfully assessed in commercial peeled products subjected to industrial processing, in which none of the products displayed labelling at species level. Overall, incorrect labelling was detected in 10 (24.4%) of the 41 commercial products tested, while another 16 samples (39%) exhibited incomplete labelling. The molecular method evaluated in this study proved to be a rapid and easy-to-perform two-step analytical approach to achieve species identification of commercial whole specimens of frozen prawns and shrimps and in peeled processed products where such raw materials are included as added-value ingredients.     

副溶血性弧菌定量检测的不确定度分析和评定

目的 分析和评定冷冻天然海虾中副溶血性弧菌定量检测最大可能数(most probable number, MPN)计数的不确定度。方法 依据JJF 1059.1-2012《测量不确定度评定与表示》、SN/T 4091–2015《食品微生物学测量不确定度评估指南》, 分析和评定检测过程中的不确定度主要来源, 运用统计学方法计算合成不确定度和扩展不确定度。结果 在95%的置信水平下, 当检验结果为75 MPN/g 和93 MPN/g时(k=2), 扩展不确定度为0.3526取值范围分别为(49~101) MPN/g、(60~126) MPN/g。评定结果表明, 实验过程的不确定度主要由样品匀液、加样体积、不同人员重复实验引入。结论 该方法适用于对冷冻天然海虾中副溶血性弧菌最大可能数计数结果的不确定度评定, 对检测结果准确度的提高具有指导意义。     

速冻去头虾品质改良工艺的研究

通过实验分析的方法,对冷冻去头虾产品的加工工艺流程中每一道工序的微生物变化及遭受的污染进行研究。结果表明:去头、去壳挑腺肠和分级工序中细菌的污染比较严重。采用独特的自制挑腺肠工具可以减少污染与提高工作效率。     

碱液处理-活性炭柱固相萃取结合GC-MS/MS法检测鱼干、虾皮和虾仁中8种N-亚硝胺

建立了碱液处理-活性炭柱固相萃取结合气相色谱-串联质谱联用（GC-MS/MS）技术检测鱼干、虾皮和虾仁中N-亚硝基二甲胺(NDMA)、N-亚硝基甲乙胺(NMEA)、N-亚硝基二乙胺(NDEA)、N-亚硝基二正丙胺(NDPA)、N-亚硝基哌啶(NPIP)、N-亚硝基吡咯烷(NPYR)、N-亚硝基吗啉(NMOR)和N-亚硝基二正丁胺(NDBA)等有害物质。以NDMA-d6、NDPA-d14和NPYR-d8为内标, 用Ba(OH)₂溶液于80 ℃处理样品1 h,离心上清液,经Sep-Pak^® plus AC-2活性炭小柱富集净化,DB-WAXUI（30 m×250 μm×0.25 μm）色谱柱分离,质谱多反应监测(MRM)模式检测。结果表明,在1~200 μg/L浓度范围内,8种N-亚硝胺的线性关系良好,R²＞0.998;检出限为0.03~0.25 μg/kg, 定量限为0.10~0.85 μg/kg,添加回收率为71.3%~119.0%（除NDBA在高添加水平时略低,为52.1%~69.0%）,相对标准偏差为0.65%~15.4%。将该方法用于23种实际样品检测,在所有样品中均检出NDMA,且含量相对较高,其他N-亚硝胺仅部分检出,含量相对较低。该方法操作简单、便于高通量分析、环境友好、定性定量可靠,可为水产品中N-亚硝胺类物质的检测提供参考。     

Bioactive Substances from Marine Fishes,Shrimps, and Algae and Their Functions: Present and Future

Marine fishes, shrimps, and algae have many important bioactive substances, such as peptides, unsaturated fatty acids, polysaccharides, trace elements, and natural pigments. The introduction of these substances contributes to a significant improvement in developing them in final processed products. In fact, the knowledge of these bioactive substances has experienced a rapid increase in the past 20 years and prompted the relevant technological revolution with a decisive contribution to the final application. The purpose of this review was to introduce critically and comprehensively the present knowledge of these bioactive substances and pointed out their future developmental situation.     

Detection and quantification of pathogenic Vibrio parahaemolyticus in shellfish by using multiplex PCR and loop-mediated isothermal amplification assay

Vibrio parahaemolyticus is a halophilic bacterium that commonly inhabits the marine and estuarine environments. This organism is also one of the leading causative pathogen of gastroenteritis often related to consumption of raw or undercooked seafood. In this study, molluscan shellfish (bloody clams and surf clams) and crustaceans (shrimps) were monitored in wet markets and hypermarkets. Two molecular methods were employed and compared to detect total and pathogenic V. parahaemolyticus in MPN enrichments: multiplex PCR and LAMP assay. The multiplex PCR was optimized to detect the total (toxR+), tdh+ and trh+ V. parahaemolyticus. On the other hand, the LAMP assay was employed to target the pathogenic strains only, the tdh+ and trh+, respectively. Out of 232 samples examined, 229 (98.7%) were positive for V. parahaemolyticus with counts ranging from 30 to >110, 000 MPN/g. Positive samples for tdh+ V. parahaemolyticus were obtained in 77 out of 232 (33.1%) samples ranging from 30 to >110, 000 MPN/g. Meanwhile, positive samples for trh+ were identified in 16 out of 232 (6.9%) samples examined ranging from 30 to 9600 MPN/g. Detection of samples with presence of tdh+ genes did not vary between methods, but a significant difference was observed when the LAMP assay was compared to PCR to detect trh+ V. parahaemolyticus. Therefore, on occasions where the density of the targeted genes is low, the LAMP assay serves as a better alternative. Nonetheless, this study constitutes an assessment of presence of total and potentially pathogenic V. parahaemolyticus in shellfishes for domestic consumption revealing the potential risk of contracting vibriosis if precautions and safety measures are not properly managed.     

Molecular identification of the black tiger shrimp (Penaeus monodon), the white leg shrimp (Litopenaeus vannamei) and the Indian white shrimp (Fenneropenaeus indicus) by PCR targeted to the 16S rRNA mtDNA

Polymerase chain reaction-based methodologies have been developed for the identification of three commercially-relevant penaeid shrimp species, these were: Litopenaeus vannamei, Penaeus monodon and Fenneropenaeus indicus in food products. Such three species represent more than 80% of the whole farmed shrimp production worldwide and may be fraudulently replaced by species exhibiting lower value such as Litopenaeus stylirostris, Penaeus semisulcatus and Fenneropenaeus merguiensis, respectively. For it, preliminary sequencing of a mitochondrial sequence of ca. 530 bp in the 16S rRNA/tRNA^Val mitochondrial region was performed in nearly 20 penaeid shrimp species of commercial relevance. Careful analysis of such sequences allowed the design of primers PNVF/PNVR, which allowed the combined identification of P. monodon and L. vannamei, and PNIF/PNIR, which allowed the specific identification of F. indicus. In addition, P. monodon and L. vannamei could be easily differentiated by either restriction with TspE1 or by amplification with novel primers MPNF/MPNR, specific for P. monodon. The proposed specific methods improve current general identification methods of these species based on more general RFLP analyses. In addition, these methods can be easily completed in less than 8 h.     

Untersuchungen zum mikrobiellen und viralen Kontaminationsstatus von Garnelen aus Aquakultur

Zusammenfassung: In der vorliegenden Untersuchung wurden 111 Proben roher, aus Drittl?ndern importierter Garnelen aus Aquakultur auf bakterielle Erreger und auf humanpathogene Viren untersucht. Die Garnelen stammten bis auf acht lateinamerikanische Proben aus Südostasien. Am h?ufigsten waren Proben aus Bangladesh vertreten (n=40). Das Untersuchungsspektrum umfasste die Ermittlung des aeroben mesophilen Gesamtkeimgehalts, die quantitative Untersuchung auf Escherichia coli und Staphylococcus aureus und den qualitativen Nachweis von Salmonella spp., Vibrio spp. und Listeria monocytogenes. Der Nachweis des humanpathogenen Rotavirus, des Norovirus und des Hepatitis A-Virus erfolgten jeweils mittels einer nested RT-PCR. Bei den untersuchten Proben wurden Gesamtkeimgehalte von 4,8×10² bis 1,3×10⁹ KbE/g ermittelt. Davon hatten vierzehn Proben (12,6%) einen aeroben mesophilen Gesamtkeimgehalt >10⁷ KbE/g. In einer Probe konnte Escherichia coli mit einer Keimzahl von 1,9×10³ KbE/g isoliert werden. In 16 Proben wurde Staphylococcus aureus nachgewiesen. Aus keiner der Garnelenproben lie? sich Listeria monocytogenes isolieren. Salmonella spp. war in acht der untersuchten Proben nachweisbar. Das am h?ufigsten isolierte Serovar war Salmonella Weltevreden. Vibrio spp. konnte mit 60 Isolaten in 49 Proben (44%) nachgewiesen werden. Am h?ufigsten wurde Vibrio cholerae non-O1 non-O139 isoliert (n=34). Aus 14 Proben konnte Vibrio parahaemolyticus isoliert werden und in fünf Proben konnte Vibrio vulnificusnachgewiesen werden. Erstmals konnte Norovirus in Garnelen nachgewiesen werden. 21 (18,9%) der untersuchten Proben waren positiv. Sowohl Rotavirus als auch Hepatitis A-Virus konnten hingegen nicht nachgewiesen werden.

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In this study 111 samples of raw imported aqua-cultured shrimps have been examined for bacterial pathogens and for pathogen viruses. The samples originated from South-East Asia except for eight Latin-American samples. Most samples were taken from Bangladesh (n=40). The bacteriological quality of these samples was analysed in terms of aerobic plate count, Escherichia coli, Staphylococcus aureus, Salmonella spp., Listeria monocytogenes and Vibrio species. Rotavirus, norovirus and hepatitis A-Virus were detected by using a nested RT-PCR. The aerobic plate count was found to be in a range between 4,8×10² to 1,3×10⁹ cfu/g. Fourteen samples (12,6%) showed an aerobic plate count >10⁷ cfu/g. One sample was found to be contaminated with Escherichia coli at a level of 1,9×10³ cfu/g. Staphylococcus aureus was isolated from sixteen samples. Listeria monocytogenes was not detected in any of the shrimp samples examined. Salmonella spp. was found in eight samples. Salmonella Weltevreden was the most frequently isolated serovar. Forty-nine shrimp samples (44%) were positive for Vibrio spp. and 60 Vibrio-isolates could be extracted. Vibrio cholerae non- O1 non-O139 was isolated in 34 samples as the most frequent species. 14 samples contained Vibrio parahaemolyticus, Vibrio vulnificus was present in five samples. For the first time Norovirus could be detected in shrimps. 21 (18,9%) of all examined samples were positive. However, both rotavirus and hepatitis A-virus could not be detected.

Eingegangen: 21. November 2007     

部分水生物中210Po含量及其在虾不同部位的分布

采用同位素示踪法对常见的17种水生物中²¹⁰Po含量进行了测定,还分析了虾不同部位中²¹⁰Po含量。水生物可食部分中 ²¹⁰Po含量范围为0.15～34.8 Bq/kg(鲜),不同水生物可食部分中²¹⁰Po含量差别较大。甲壳类、头足类和贝类水生物对²¹⁰Po有很强的浓集能力,尤其是牡蛎、八爪鱼、螃蟹和鱿鱼中²¹⁰Po含量很高,分别为34.8、25.0、20.3和17.2 Bq/kg(鲜),超过了我国现行国标食品中放射性物质限制浓度中的限值。虾不同部位中²¹⁰Po含量呈现规律分布,头部中²¹⁰Po含量最高,壳次之,肉最低。