咸阳市市售食品中单核细胞增生李斯特菌基因组特征及耐药和致病基因分析

目的 采用全基因组测序技术对从咸阳市市售食品中分离的64株单核细胞增生李斯特菌（单增李斯特菌）的基因组特征,耐药和致病性基因进行分析。方法 收集咸阳市市售食品中分离的64株单增李斯特菌,利用微量肉汤法进行药敏测定,同时进行全基因组测序,原始序列经拼接后利用生物信息学软件进行基因组注释、系统发育树构建及基因组特征和遗传原件分析。结果 64株分离株对于氨苄西林、青霉素、美罗培南、复方新诺明、万古霉素5种抗生素结果均为敏感。其中2株分离株对2种抗生素产生抗性,分别为四环素和红霉素。全基因组测序分析表明,64株分离株分属3个谱系,分为15个克隆群（CC）,以谱系Ⅰ和谱系Ⅱ为主;2株耐药株基因型与表型一致,耐药基因上下游遗传环境分析表明,这些基因的可能来源为猪丹毒杆菌、肠球菌和外源质粒。所有分离株均携带致病基因岛LIPI-1和LIPI-2,部分谱系Ⅰ菌株携带LIPI-3或LIPI-4,具有潜在致病风险。携带质粒和抗性相关基因的主要为谱系Ⅱ菌株。inlA基因提前终止突变均发生在谱系Ⅱ,可能降低菌株毒力。结论 咸阳市市售食品中单增李斯特菌基因组结构相对稳定,菌株存在获得性耐药,且因携带更多毒力基因而产生潜在的高致病性菌株。谱系Ⅰ和谱系Ⅱ菌株在毒力基因、抗性相关基因和质粒携带方面均具有差异,显示不同CC型菌株的毒力和环境适应性存在差异,为咸阳市单增李斯特菌的监测和防控提供了参考数据。     

苏州市食源性单核细胞增生李斯特菌的全基因组测序分析

目的 应用高通量测序技术分析苏州市食源性单核细胞增生李斯特菌(Listeria monocytogenes, Lm)的毒力基因携带情况、分子分型、遗传进化谱系及遗传进化关系等分子特征。方法 对2016—2020年分离自食品的42株Lm进行全基因组测序,运用CLC Genomics Workbench 21.0.4软件进行组装及毒力基因和耐药基因分析;通过与BIGSdb-Lm数据库比对获得谱系、克隆群(clone complexes,CC)、血清群、多位点序列分型(multilocus sequence typing,MLST)、核心基因组多位点序列分型(core genome multilocus sequence typing,cgMLST);利用柏熠微生物分析平台v4.0构建最大似然树。结果 42株Lm共包含两个谱系,谱系Ⅰ和谱系Ⅱ,以谱系Ⅱ为主(83.3%)。血清群分为Ⅱ_a、Ⅱ_b和Ⅱ_c,以Ⅱ_a为主(57.1%)。42株Lm分为11个CC型, 11个序列型(sequence type, ST型)...     

食品中单核细胞增生李斯特氏菌检测分析

目的:分析食品中单核细胞增生李斯特氏菌的检测结果,以及时发现食品安全隐患,为食品安全监管提供参考依据.方法:严格按照《全国食源性致病菌监测工作手册》中的单核细胞增生李斯特氏菌检验标准,对某市2021年1—12月定期采样的10类常用食品进行单核细胞增生李斯特氏菌检测,分析其阳性检出率.结果:897份10类常用食品中,检出...     

水产品中单核细胞增生李斯特氏菌检验研究

本文是对水产品中单核细胞增生李斯特氏菌常规检验的研究。结合国家标准、行业标准和FDA《细菌学检验手册》第八版中所述方法进行检验。自朝鲜进境的水产品23类、255批中检出李斯特氏菌7株。结果表明,由于食品加工过程对微生物损害极大,使大多数微生物处于濒死状态,因而加速受损细胞恢复是单核细胞增生李斯特氏菌检验前增菌的关键。在检验过程中最初分离时发现了可疑菌落并就此进行确认和得出结论,而不是延长增菌时间继续分离和鉴定,容易产生假阴性结果。     

食品中单核细胞增生李斯特氏菌检测结果的分析

目的:分析食品中单核细胞增生李斯特氏菌的检测结果,以及时发现食品安全隐患,为食品安全监管提供参考依据。方法:严格按照《2012年食源性致病菌监测工作手册》中的单核细胞增生李斯特氏菌检验标准,对济宁市2019年1—12月定期采样的8类常用食品(生鲜猪肉、生鲜牛肉、生鲜羊肉、生鲜鸡肉、冻虾、冻带鱼、凉拌菜与冰激凌)进行单核细胞增生李斯特氏菌检测,分析其阳性检出率。结果:567份8类常用食品中,检出单核细胞增生李斯特氏菌27份,阳性检出率为4.76%,其中包括生鲜猪肉6份(5.94%)、生鲜羊肉5份(6.02%)、生鲜鸡肉5份(5.10%)、冻虾4份(7.69%)、冻带鱼2份(3.28%)以及凉拌菜5份(11.63%),生鲜牛肉与冰激凌均未检出单核细胞增生李斯特氏菌。结论:食品中存在单核细胞增生李斯特氏菌污染的风险,其中以凉拌菜的污染风险最高,应加强该方面的防控,以确保食品安全。     

食品中单核细胞增生李斯特氏菌检测能力验证

目的通过能力验证提升食品中单核细胞增生李斯特氏菌检测能力和实验室质量管理水平。方法依据GB 4789.30-2010《食品安全国家标准食品微生物学检验单核细胞增生李斯特氏菌检验》进行检测,利用BAX system Q7全自动病原微生物检测系统对能力验证中的3个样品进行快速筛查,采用VITEK2COMPACT全自动细菌鉴定系统、基质辅助激光解析电离飞行时间质谱(matrix-assisted laser desorption ionization time-of-flight mass spectrometry,MALDI-TOF-MS)和溶血素O基因(hlyA)序列比对分析鉴定可疑菌株。结果编号CODE1和CODE3的样品检出单核细胞增生李斯特氏菌,编号CODE2的样品未检出。结论本实验室参加了中国食品药品检定研究院组织的食品中单核细胞增生李斯特氏菌检测能力验证测试,取得了满意的实验结果。     

加工环境中单核细胞增生性李斯特菌耐药特征分析

目的分析在流通过程中加工环境来源的食源性李斯特菌耐药性及耐药基因携带情况,探讨加工环境对该菌的影响。方法收集2016~2017年分离自加工环境中的71株单核细胞增生性李斯特菌,利用微量肉汤稀释法对8种抗菌药物进行药物敏感性分析,并采用PCR鉴定11种耐药基因(tet A、tet M、tet S、erm A、erm B、erm C、mec A、aac(6’)-Ib、van A、van B、cfr)。结果 71株Lm对庆大霉素(11.27%)、氨苄西林(4.22%)、头孢他啶(98.59%)、环丙沙星(97.18%)、四环素(21.12%)、红霉素(15.49%)、林可霉素(92.96%)均具有不同程度的耐药性,对万古霉素敏感。其中以耐受3~6种抗菌药为主,耐药情况较为严重。耐药基因检测表明四环素类tet A,tet M为主要耐药基因,检出率高达50%以上,其次为红霉素类erm A、erm B、erm C,检出率约11%,氨基糖苷类aac(6’)-Ib的检出率为5.63%。结论加工环境单核细胞增生性李斯特菌的耐药性呈上升趋势,其耐药表型与耐药基因型并不完全一致,从而表明加工环境中存在影响单核细胞增生性李斯特菌耐药性因素,同时为食源性李斯特菌病的防治及预警提供参考依据。     

基于全基因组测序的单核细胞增生李斯特菌食品分离株分子特征分析

目的 了解肇庆市单核细胞增生李斯特菌(以下简称单增李斯特菌)食品分离株的基因组特征、毒力岛的携带情况及遗传多样性。方法 对肇庆市13株单增李斯特菌食品分离株进行全基因组测序,将组装好的contings/Scaffolds上传至在线分析平台Center for Genomic Epidemiology、Rast和VFanalyzer进行基因组注释与毒力因子基因鉴定,并应用基于全基因组测序的单核苷酸多态性分型(wg-SNPs)方法与美国生物技术信息中心(NCBI)上获取的25株国内外单增李斯特菌的基因组进行遗传进化分析。结果 13株单增李斯特菌食品分离株的基因组大小为2.82～3.04 Mb,CG含量为37.9%～38.1%,可分为6个ST型(ST1、ST3、ST8、ST59、ST87、ST101),分属于6个克隆复合群(CC1、CC3、CC8、CC59、CC87、CC101)。其中,ST3型菌株均携带LIPI-3毒力岛基因,ST87型菌株均携带完整LIPI-4毒力岛基因。wg-SNPs遗传进化分析显示13株单增李斯特菌食品分离株可分为2个进化分支,其中ST3型菌株位于进化树的根部,与其他ST型菌株进化上存在差异。结论 肇庆市单增李斯特菌食品分离株以高毒力的ST87型和ST3型为流行型,并发现一株同时携带LIPI-1～LIPI-4毒力岛的ST87型菌株,应加强对此类菌株的监测,警惕高毒力菌株在肇庆市引起感染性暴发的风险。     

实时荧光PCR检测熟肉制品中单核细胞增生李斯特氏菌的结果分析与探讨

目的:对实时荧光PCR检测单核细胞增生李斯特氏菌假阳性结果的原因分析探讨。方法:对假阳性的样本进行传统培养法鉴定分析,将鉴定的菌落进行实时荧光PCR检测,同时对样本基质的影响和培养基本底的影响进行实时荧光PCR检测。结果:样本中的可疑菌落、样本的基质、培养基本底对实时荧光PCR检测结果均无假阳性影响。结论:实时荧光PCR检测熟肉制品中单核细胞增生李斯特氏菌的假阳性主要原因可能是样本中存在死亡目的菌基因造成的干扰。     

产单核细胞增生性李斯特氏菌致病机制的研究进展

产单核细胞增生性李斯特氏菌是一类人畜共患的食源性致病菌.本文阐述了产单核细胞增生性李斯特氏菌的生物学特性、毒力因子及流行病学,旨在为产单核细胞增生性李斯特氏菌的致病机制提供理论依据.     

Control of Listeria monocytogenes in ready-to-eat meats containing sodium levulinate, sodium lactate, or a combination of sodium lactate and sodium diacetate

ABSTRACT:  This study investigated the use of sodium levulinate to prevent outgrowth of Listeria monocytogenes in refrigerated ready-to-eat (RTE) meat products. Turkey breast roll and bologna were formulated to contain 1%, 2%, or 3% (w/w) sodium levulinate, 2% sodium lactate, a 2% combination of sodium lactate and sodium diacetate (1.875% sodium lactate and 0.125% sodium diacetate), or no antimicrobial (control). Samples of the RTE products were sliced, inoculated with 10² to 10³ CFU/cm² of a 5-strain cocktail of L. monocytogenes , vacuum packaged, and stored at refrigeration temperature for 0 to 12 wk. Counts reached 10⁸ CFU/cm² on control turkey roll product after 8 wk, and over 10⁷ CFU/cm² on control bologna after 12 wk. Addition of 2% or more sodium levulinate to turkey roll and 1% or more sodium levulinate to bologna completely prevented growth of L. monocytogenes during 12 wk of refrigerated storage. A consumer taste panel with pathogen-free samples found no differences in the overall liking among the preparations of turkey roll or among preparations of bologna. These results show that sodium levulinate is at least as effective at inhibiting outgrowth of L. monocytogenes in RTE meat products as the current industry standards of lactate or lactate and diacetate, and levulinate addition does not alter the overall liking of the RTE meat products.     

Listeria monocytogenes: Strain Heterogeneity,Methods, and Challenges of Subtyping

Listeria monocytogenes is a food‐borne bacterial pathogen that is associated with 20% to 30% case fatality rate. L. monocytogenes is a genetically heterogeneous species, with a small fraction of strains (serotypes 1/2a, 1/2b, 4b) implicated in human listeriosis. Monitoring and source tracking of L. monocytogenes involve the use of subtyping methods, with the performance of genetic‐based methods found to be superior to phenotypic‐based ones. Various methods have been used to subtype L. monocytogenes isolates, with the pulsed‐field gel electrophoresis (PFGE) being the gold standard. Although PFGE has had a massive impact on food safety through the establishment of the PulseNet, there is no doubt that whole genome sequence (WGS) typing is accurate, has a discriminatory power superior to any known method, and allows genome‐wide differences between strains to be quantified through the comparison of nucleotide sequences. This review focuses on the different techniques that have been used to type L. monocytogenes strains, their performance challenges, and the tremendous impact WGS typing could have on the food safety landscape.     

Growth or Survival of Listeria monocytogenes in Ready-to-Eat Meat Products and Combination Deli Salads During Refrigerated Storage

Growth or survival of Listeria monocytogenes in cold‐smoked salmon; sliced, cooked ham; sliced, roasted turkey; shrimp salad; and coleslaw obtained at retail supermarkets stored at 5 °C, 7 °C, or 10 °C (41 °F, 45 °F, or 50 °F, respectively) for up to 14 d was evaluated. Cold‐smoked salmon, ham, and turkey were obtained in case‐ready, vacuum packages. All food products were stored aerobically to reflect additional handling within the retail supermarket. Cold‐smoked salmon, ham, and turkey supported the growth of L. monocytogenes at all 3 storage temperatures. Fitted growth curves of initial populations (about 3 log₁₀ colony‐forming units [CFU]/g) in cold‐smoked salmon, ham, and turkey stored at 5 °C achieved maximal growth rates of 0.29, 0.45, and 0.42 log₁₀ CFU/g growth per day, respectively. Storage at 10 °C increased the estimated maximal growth rate of the pathogen by 0.56 to 1.08 log₁₀ CFU/ g growth per day compared with storage at 5 °C. A decline in populations of L. monocytogenes was observed in shrimp salad and coleslaw, and the rate of decline was influenced by storage temperature. Retention of viability was higher in shrimp salad than in coleslaw, where populations fell 1.2, 1.8, and 2.5 log₁₀ CFU/g at 5 °C, 7 °C, and 10 °C, respectively, after 14 d of storage. Inability of shrimp salad and coleslaw to support the growth of L. monocytogenes may be attributed to the acidic pH (4.8 and 4.5, respectively) of the formulations used in this study. Results show that the behavior of L. monocytogenes in potentially hazardous ready‐to‐eat foods is dependent upon the composition of individual food products as well as storage temperature.     

Occurrence and Antimicrobial Susceptibility of Listeria monocytogenes Isolated from Brined White Cheese in Jordan

Listeria monocytogenes is a serious foodborne pathogen that has been isolated from different dairy food products. Several foodborne outbreaks of listeriosis have been associated with consumption of cheese. The aims of this study were to determine the occurrence of L. monocytogenes and Listeria spp. in brined white cheese (BWC) sold in Jordan, and to determine the susceptibility of isolated L. monocytogenes to antimicrobials. Three hundred and fifty samples of 5 different types of BWC (akkawi, boiled, halloumi, pasteurized, and shellal) were collected from a local market in Jordan. The ISO (11290-1) procedure was followed for isolation and identification of Listeria spp. from cheese samples and a polymerase chain reaction (PCR) technique was used for confirmation of L. monocytogenes isolates. The VITEK2 automated system was used for testing antimicrobial susceptibility of L. monocytogenes isolates. The overall prevalence of Listeria spp. in cheese sample was 27.1%. L. monocytogenes was isolated from 39 (11.1%) samples. Other isolated species were L. grayi (6.9%), L. innocua (2%), L. ivanovii (4%), L. seeligeri (2%), and L. welshimeri (0.3%). The pH values and salt concentrations of L. monocytogenes positive cheese samples ranged from 5.10 to 6.32 and 5.64 to 13.16, respectively. L. monocytogenes isolates were sensitive or intermediate susceptible to imipenem, gentamicin, linezolid, teicoplanin, vancomycin, fusidic acid, trimethoprim/sulfamethoxazole, benzylpenicillin, ciprofloxacin, erythromycin, tetracycline, and rifampicin, but resistant to fosfomycin, oxacillin, and clindamycin.     

Antimicrobial activity of Ginkgo biloba leaf extract on Listeria monocytogenes

ABSTRACT: The antimicrobial activities of Ginkgo biloba leaf extract (GBE) and the combined effects of GBE and sodium EDTA (sodium Ethylenediaminetetraacetic acid) against Listeria monocytogenes were determined at 4 °C, 25 °C, and 37 °C. Listeria monocytogenes grown at 37 °C for 24 h was inoculated (6 to 7 log CFU/mL) into BHI broth containing either GBE or GBE and EDTA (1.6 mg/mL) with various GBE concentrations of 0.1, 0.25, 0.5, 1, 2.5, 5.0, 7.5, 10.0, 15.0, or 20.0% vol/vol and stored at 4 °C, 25 °C, and 37 °C. The inhibitory effect of the GBE was more pronounced at low temperature of 4 °C. GBE was effective in inhibiting microbial growth. Addition of EDTA enhanced antimicrobial activity of GBE.     

Combining Antimicrobial and Steam Treatments in a Vacuum-packaging System to Control Listeria monocytogenes on Ready-to-eat Franks

ABSTRACT: This study was to evaluate the effectiveness of steam or steam in combination with an antimicrobial agent to control Listeria monocytogenes on ready-to-eat (RTE) franks. The franks were surface-inoculated to contain 6 or 3 log₁₀(colony-forming units [CFU])/cm² of L. monocytogenes and treated with steam or steam in combination with an antimicrobial agent, immediately followed by vacuum-sealing the top films of frank packages (6 franks per package in a single layer). Three log (CFU) /cm² of reductions were achieved at the both inoculation levels for L. monocytogenes on franks. At an inoculation level of 3 logs, no outgrowth of L. monocytogenes was obtained on the treated franks after storing at 4.4°C or 16°C for a combined 47 d. This study provided an alternative approach for controlling L. monocytogenes in packaged franks.     

Controlling Listeria monocytogenes in Cold Smoked Salmon with the Antimicrobial Peptide Salmine

Listeria monocytogenes (LM) is a major safety concern for smoked salmon producers, as it can survive both the brining and smoking process in cold smoked salmon production. Salmine is a cationic antimicrobial peptide derived from the milt of salmon that has been shown to inhibit the growth of LM in vitro. Commercialization of this peptide would add value to a waste product produced when raising salmon. The purpose of this study was to determine the anti‐listeria activity of salmine in smoked salmon by measuring the viable counts of LM over time. Cold smoked salmon was treated with a salmine solution or coated with agar or k‐carrageenan films incorporating salmine to maintain a high surface concentration of the antimicrobial. Samples were then inoculated with approximately 1.0 × 10³ cells of LM. The viable counts were then enumerated throughout 4 wk at 4 °C storage. It was found that 5 mg/g salmine delayed the growth of LM on smoked salmon. These samples had significantly (P < 0.05) lower LM counts than on the untreated samples on days 13 and 22. Edible films did not significantly (P > 0.05) improve the antimicrobial efficacy of salmine. The peptide combined with biopolymers also had lower antimicrobial activity in vitro when compared to salmine alone. These results suggest there is potential for salmine to be used as a natural hurdle to inhibit growth of LM due to post process contamination; however, future investigations for extending this effect throughout the shelf life of smoked salmon products are warranted.     

Impact of nitrite on detection of Listeria monocytogenes in selected ready-to-eat (RTE) meat and seafood products

ABSTRACT:  The impact of sodium nitrite (NaNO₂) on detection and recovery of Listeria monocytogenes from select ready-to-eat (RTE) foods including smoked salmon, smoked ham, beef frankfurters, and beef bologna was assessed. Nitrite-containing (NC; 100 to 200 ppm NaNO₂) or nitrite-free (NF) foods were inoculated with a 5-strain cocktail of L. monocytogenes by immersion into Butterfield's buffer solution containing 5.4 to 7.4 × 10³ L. monocytogenes per milliliter. Inoculated products were vacuum-packaged and stored at 5 °C. A weekly comparative analysis was performed for presence of L. monocytogenes using 5 detection methods on products held at 5 °C for up to 8 wk. L. monocytogenes initially present at <100 CFU/g during the first 2 wk of storage increased throughout the study, attaining final populations of approximately 1 × 10⁴ to 1 × 10⁵ CFU/g. Lactic acid bacteria predominated throughout the study in all products. Exposure to NaNO₂ (100 to 200 ppm) resulted in 83% to 99% injury to the L. monocytogenes strains tested. The genetic-based BAX^® System (DuPont™ Qualicon, Wilmington, Del., U.S.A.) and modified USDA/FSIS methods detected 98% to 100% of Listeria -positive food samples and were consistently superior to and significantly different ( P < 0.05) from conventional cultural methods in recovering Listeria from NC samples. Data show that nitrite-induced injury adversely affects detection and recovery of L. monocytogenes from NC food, confirming earlier findings that nitrite-induced injury masks L. monocytogenes detection in NC RTE food products. Nitrite-injured Listeria can subsequently repair upon nitrite depletion and grow to high levels over extended refrigerated storage.     

单核细胞增生李斯特氏菌食品分离株协同 溶血现象研究

目的对从食品中分离的88株单核细胞增生李斯特氏菌(Listeria monocytogenes)进行协同溶血(christie,atkins and munch-petersen,c AMP)测试。方法根据食品安全国家标准食品微生物学检验单核细胞增生李斯特氏菌检验GB 4789.30-2010的方法进行c AMP测试。结果所测试的88株单核细胞增生李斯特氏菌分离株与GB 4789.30中描述的结果一致,即在靠近金黄色葡萄球菌(Staphylococcus aureus)的接种端溶血增强,78株靠近马红球菌一端呈阴性反应。同时,发现10株分离株c AMP测试结果与传统的测试结果不同,在靠近马红球菌(Rhodococcus equi)的接种端溶血增强。结论对88株单核细胞增生李斯特氏菌分离株c AMP测试结果表明,有大约11%的菌株在靠近马红球菌的接种端溶血增强。