有关样品培养菌落(细菌)计数的讨论

一、前盲 细菌悬液或稀释液经培养形成菌落,通过菌落计数可推算悬液中的含菌量,这是基础微生物常用实验技术方法之一。在各行业的样品检验中,常被用来检测样品中的含菌量(液体样品可直接取样培养,可溶性固体样品可制成溶液取样培养,非可溶性样品可     

蓝莓中诺如病毒检测方法的优化及其在果蔬中的应用

目的:建立蓝莓等浆果类和蔬菜类食品中GII型诺如病毒(NoV GII)的有效检测方法。方法:利用已知NoV GII病毒样本对阴性的蓝莓进行人工染毒,分别比较7种洗脱液和2种浓缩方法的回收效果,优化RNA提取方法,最后采用荧光定量PCR方法进行检测。用新建立的诺如病毒浓缩检测方法,对7种市购浆果和蔬菜样品进行检测并研究其最低检出限。结果:实时荧光RT-PCR检测结果表明,人工染毒后的蓝莓等浆果样品(表皮和基质)采用TP ALK洗脱液洗脱,经酶处理后超滤法浓缩,最后采用病毒核酸提取试剂盒(QIAamp Viral RNA Mini Kit)提取浓缩病毒的RNA,诺如病毒的回收效率达到较高的水平,其灵敏度达3.5～350 PCRU/10g。将该方法用于草莓、杨梅、葡萄、小番茄、生菜、胡萝卜、黄瓜等7种水果蔬菜中诺如病毒的检测,其中杨梅和小番茄的检测灵敏度达到3.5～3 500 PCRU/10 g,草莓、生菜和胡萝卜次之,为35～35 000 PCRU/10 g,黄瓜和葡萄的灵敏度较低。结论:所建立的果蔬中诺如病毒富集方法和核酸提取方法操作简便,灵敏度较高,适合于蓝莓等浆果和蔬菜类样品中NoV GII的检测。     

醇化烟叶表面可培养微生物的鉴定和系统发育分析

以K326烤烟品种为材料,对不同醇化时间烟叶表面的可培养微生物进行了分离,并通过分子生物学方法对这些微生物的种、属分类地位和系统发育关系进行了分析.结果表明:醇化烟叶表面可培养微生物的数量和种类随着醇化时间的延长呈明显的下降趋势;醇化烟叶表面的细茵主要包括芽孢杆茵和肠杆茵两个类群,其中,芽孢杆菌属的细菌为优势微生物类群.此外,醇化烟叶表面细菌的数量和烟叶部位等级也存在一定的相关性,即中部烟叶样品中细菌的数量高于下部烟叶样品,而上部烟叶样品中的细菌数量最少.而真茵的数量和种类与醇化时问和烟叶样品部位等级之间没有明显的相关性.     

仓储方式对复烤烟叶醇化品质及表面细菌多样性的影响

为比较烤烟烟叶在机械除湿仓储(磷化氢熏蒸杀虫、硫酰氟熏蒸杀虫)和气调仓储3种处理模式下,烟叶感官品质、常规化学成分和烟叶表面细菌多样性的差异,对样品分别进行磷化氢熏蒸杀虫机械除湿仓储、硫酰氟熏蒸杀虫机械除湿仓储和气调仓储处理。仓储24个月后取样检测烟叶常规化学成分,评价样品感官品质,提取烟叶表面细菌总DNA,通过巢式PCR扩增及变性梯度凝胶电泳法分析3种处理条件下仓储后相同来源烟叶表面的细菌多样性指标。结果显示:(1)相同来源的烟叶样品机械除湿加磷化氢熏蒸仓储感官品质综合指标优于气调和机械除湿加硫酰氟熏蒸仓储,不同来源的烟叶样品化学指标变化呈现出不同的规律,气调仓储烟叶样品的含水率保持最好、质量损失最少。(2)对于云南师宗C3F烟叶在3种处理条件下仓储24个月后,机械除湿加磷化氢熏蒸仓储条件下烟叶表面的细菌种类最多、多样性指数最大;在机械除湿加硫酰氟熏蒸仓储条件下烟叶表面的细菌多样性指数最小;在气调仓储方式下烟叶样品表面一些种类的细菌数量存在较大差异,说明烟叶表面细菌的均匀性较差。     

鲜切生菜在贮藏期间的微生物生长模型

主要研究了在 0℃、4℃和 2 5℃贮藏温度下鲜切生菜中细菌的生长趋势和感官质量的变化。研究结果表明 ,实验中所建立的Gompertz模型能有效地拟合在不同贮藏温度下鲜切生菜中细菌总数的动态变化 ,从而能预测不同贮藏温度下、不同贮藏时间内鲜切生菜中的细菌总数 ,为快速、有效地评估鲜切生菜的货架期和微生物安全性 ,提供了一个方便有效的方法。同时探讨了鲜切生菜中细菌总数与细胞组织腐烂的关系 ,即当细菌总数≤ 1 0 6 cfu/ g ,鲜切生菜的细胞组织不会腐烂。     

样品前处理对含油脂性成分化妆品微生物检测结果的影响

研究含油脂性成分化妆品样品前处理方式的不同对检测结果的影响。按《化妆品卫生规范》（2007版）中样品前处理方法（A法）和用生理盐水直接处理样品（B法）同时对50份油脂性样品做微生物检测。结果表明：按A法处理样品,检测结果细菌总数、霉菌和酵母菌计数50份样品的平均合格率分别为：88.87%和84.33%;按B法处理样品,检测结果细菌总数、霉菌和酵母菌计数50份样品的平均合格率分别为：97.37%和97.5%,致病菌均未检出。含油脂性成分化妆品按照A法处理,样品溶解性更好,微生物更易检出。     

天津市2015~2019年生鲜果蔬病原微生物污染排查分析

为排查天津市即食生鲜果蔬病原微生物污染情况,2015~2019年在生产环节和流通环节抽取生菜、番茄、黄瓜、苦菊、桃和梨等6种即食果蔬品种共计654批次样品,采用国家标准方法分析样品中菌落总数、大肠菌群、沙门氏菌、金黄色葡萄球菌、单增李斯特氏菌、蜡样芽孢杆菌和大肠杆菌O157:H7污染状况进行风险分析。结果显示,654批次样品中检出食源性致病菌135批次,检出率为20.6%,其中包括沙门氏菌1份,金黄色葡萄球菌6份,蜡样芽孢杆菌128份,其他致病菌均未检出。食源性致病菌检出率较高的样品多出现在超市和农贸市场,表明即食果蔬致病菌的污染易发生在流通环节。即食生鲜果蔬中食源性致病菌污染对消费者健康存在潜在的安全隐患,需加强采后流通环节污染防控,保障即食果蔬食品安全,防止食源性疾病暴发。     

食品微生物检验中两种均质方法的比较

目的:探究在食品微生物的检测过程中旋刀式均质方法以及拍击式均质方法的优缺点。方法:选取了食品样本后,分别使用两种均质方法,对食品样本中所含有的细菌数量和霉菌数量进行分析。结果:拍击式均质方法回收的霉菌数量相对较高,而旋刀式均质方法则更易回收细菌类。结论:在后期食品微生物检验过程中,选择均质方法可以依据需要回收细菌率较高,还是霉菌率较高进行选择,同时也可以根据样品的不同形态以及检验的不同要求进行均质方法的选择。     

氢化物原子荧光光度法测定粮食中无机砷的前处理方法比较

参照GB/T 5009.11—2003中氢化物原子荧光光度法无机砷检测样品的前处理方法,对粮食试样的前处理方法做了改进,并将3种不同前处理条件下的测定结果进行对比分析。通过加标回收试验和样品精度试验结果可知,3种前处理方法测定结果的回收率在94%~102%之间,且精密度好。试验结果表明,采用70℃超声1 h和70℃水浴1 h对粮食样品进行前处理,可极大缩短试验时间,提高分析速度。     

土壤中重金属检测样品前处理技术现状探讨

在化学检测过程中,最容易导致检测误差的就是取样前的技术处理,特别是随着技术设备的不断更新升级,对检测样品前的技术处理提出了更高的要求,在不同条件下的取样处理技术不同.以土壤中重金属检测样品前处理技术为研究对象,重点分析技术应用现状,以更好地提升检测的科学性和精准度.     

阿氏芽孢杆菌(Bacillus aryabhattai)分离鉴定及其对蔬菜中甲萘威农药降解作用的研究

目的 从寒地黑土中筛选甲萘威降解菌,并对筛选得到的菌株进行形态学及16S rDNA分子鉴定,将该菌株用于蔬菜样品中甲萘威农药残留降解规律的研究。方法 从土壤样品中用含甲萘威农药的培养基进行平板稀释法初筛、96孔板法复筛,分别在37℃(1 d)、4℃(6 d)下进行培养,筛选在OD₆₀₀ nm处吸光值高的菌株,即为可利用甲萘威生长的菌株。获得OD₆₀₀ nm为0.906的菌株D7,并对该菌株进行形态学以及分子生物学鉴定。结果 菌株D7经鉴定为阿氏芽孢杆菌(Bacillus aryabhattai);菌株D7对所选5种叶菜和5种果菜中残留的甲萘威均有一定降解作用,且对生菜中甲萘威的降解率最高为58.7%。结论 筛选出阿氏芽孢杆菌D7,在低温条件生长良好。为微生物法降解甲萘威提供优良降解菌株,解决蔬菜中甲萘威残留的问题。     

Microbial safety considerations of flooding in primary production of leafy greens: A case study

This study evaluated the effects of a flood event, floodplain and climatic parameters on microbial contamination of leafy greens grown in the floodplains. Additionally, correlations between pathogenic bacteria and levels of indicator microorganisms have been also determined. To diagnose the microbial contamination after the flood event, sampling was carried out in weeks 1, 3, 5 and 7 after the flooding in four flooded lettuce fields. To assess the impact of flooding on the microbial contamination of leafy greens, indicator microorganisms (coliforms, Escherichia coli and Enterococcus) and pathogenic microorganisms (Salmonella spp., VTEC (E. coli O157:H7 and other verocytotoxin producing E. coli, O26, O103, O111, O145) and Listeria monocytogenes) were evaluated. Irrigation water, soil and lettuce samples showed levels of coliforms and E. coli higher than 5 and 3 log cfu/g or 100 mL, respectively when sampled 1 week after flooding. However, bacterial counts drastically declined three weeks after the flooding. Climatic conditions after flooding, particularly the solar radiation (6–8 MJ/m²), affected the survival of bacteria in the field. L. monocytogenes was not detected in lettuce samples, except for 2 samples collected 3 weeks after the flooding. The presence of Salmonella was detected in irrigation water, soil and lettuce by multiplex PCR one week after the flooding, but only 2 samples of soil and 1 sample of water were confirmed by colony isolation. Verotoxigenic E. coli was detected in soil and lettuce samples by multiplex PCR. Therefore, the implication of flood water as the source of VTEC contamination of soil and lettuce was not clear. E. coli counts in irrigation water were positively correlated with those in lettuce. A significant correlation (P < 0.005) was found between the presence of pathogens and E. coli counts, highlighting a higher probability of detection of pathogens when high levels of E. coli are found. The results obtained in the present study confirm previous knowledge which defined flooding as a main risk factor for the microbial contamination of leafy greens.     

UNE-EN ISO/IEC 17025:2005-accredited method for the determination of pesticide residues in fruit and vegetable samples by LC-MS/MS

A rapid, simple and sensitive multi-residue method was developed and validated for the simultaneous quantification and confirmation of 69 pesticides in fruit and vegetables using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The samples were extracted following the quick, easy, cheap, effective, rugged and safe method known as QuEChERS. Mass spectrometric conditions were individually optimised for each analyte in order to achieve maximum sensitivity in multiple reaction monitoring (MRM) mode. Using the developed chromatographic conditions, 69 pesticides can be separated in less than 17 min. Two selected reaction monitoring (SRM) assays were used for each pesticide to obtain simultaneous quantification and identification in one run. With this method in SRM mode, more than 150 pesticides can be analysed and quantified, but their confirmation is not possible in all cases according to the European regulations on pesticide residues. Nine common representative matrices (zucchini, melon, cucumber, watermelon, tomato, garlic, eggplant, lettuce and pepper) were selected to investigate the effect of different matrices on recovery and precision. Mean recoveries ranged from 70% to 120%, with relative standard deviations (RSDs) lower than 20% for all the pesticides. The proposed method was applied to the analysis of more than 2000 vegetable samples from the extensive greenhouse cultivation in the province of Almería, Spain, during one year. The methodology combines the advantages of both QuEChERS and LC-MS/MS producing a very rapid, sensitive, accurate and reliable procedure that can be applied in routine analytical laboratories. The method was validated and accredited according to UNE-EN-ISO/IEC 17025:2005 international standard (accreditation number 278/LE1027).     

微波消解-电感耦合等离子体原子发射光谱法测定海参中铝的残留量

目的建立微波消解-电感耦合等离子体原子发射光谱测定海参中铝含量的方法,并对北京市场中海参中铝含量进行初步调查。方法抽取市场上37个海参样品,经5 m L硝酸微波消解后,采用电感耦合等离子体原子发射光谱法进行测定。结果该方法检出限为5.0 mg/kg,加标回收率为95%~105%。通过单因素方差分析,本方法的测定结果与国标方法测定所得的数据结果无显著差异。测定的37个样本中均检出铝,其中最低含量为29.2 mg/kg,最高含量为304 mg/kg,100 mg/kg以上的样本占62.2%。结论本方法具有准确度高、精密度好、操作简单、分析速度较快等特点。从市场抽样检测结果来看,海参中铝安全应该引起一定的重视,建议生产企业进一步改进生产工艺,确保海参质量安全。     

QuEChERS结合GC-MS/MS法测定果蔬汁中21种农药残留

建立一种采用QuEChERS前处理结合Quick Pro净化柱-气相色谱串联质谱（GC-MS/MS）法测定果蔬汁中的21种农药残留量的方法。样品采用乙腈提取,EN方法盐包盐析,经Quick Pro净化柱净化,DB-35MS UI色谱柱分离,采用选择离子监测（SIM）模式扫描,外标法定量。在质量浓度0.01～0.50 μg/mL范围内相关系数（R）均＞0.995,样品在0.01 mg/kg、0.02 mg/kg、0.10 mg/kg的添加水平下,21种农药的回收率在75.2%～117.8%之间,回收率实验结果相对标准偏差（RSD）在1.9%～11.5%之间。该方法操作简便,分析时间短,精密度好,适用于果蔬汁中多种农药残留的检测。     

DISINFECTION OF SHREDDED SALAD INGREDIENTS WITH SODIUM DICHLOROISOCYANURATE

Raw salads are frequently subjected to hypochlorite wash to reduce microbial numbers, retard enzymatic activity and improve shelf-life of sensory quality. Hypochlorite dips significantly reduce the initial population of natural contaminants on lettuce. However, populations on treated and untreated samples are not significantly different after four days storage at refrigeration temperature. Increasing the level of free available chlorine does not improve the microbicidal effect.
In this study sodium dichloroisocyanurate (NaDCC) was used to disinfect salad vegetables. Though the principal salad vegetable used was butterhead lettuce, results show that the efficacy of NaDCC in reducing total aerobic mesophile levels varies with vegetable type. NaDCC gives an acid pH in solution and has significantly higher activity than sodium hypochlorite.
For longer term storage, preparation of lettuce which included a 30 min soak in NaDCC improved the microbiological quality of the produce at the end of the storage period. The microbial load on disinfected produce was between 0.62 and 0.98 log cycles lower than on lettuce prepared without a disinfection step for up     

气相色谱-火焰光度检测器法检测果蔬样品中3种二硫代氨基甲酸盐类农药残留

目的建立检测福美铁、丙森锌、代森锰农药残留的气相色谱-火焰光度检测器(硫滤光片)[GC-FPD(S)]测定方法,提高该类农药的检测专一性。方法用带有强螯合作用的碱性缓冲溶液将不溶于水的二硫代氨基甲酸重金属盐转化为可溶于水的钠盐,在离子对甲基化的作用下将二硫代氨基甲酸钠盐转化为可供GC-FPD(S)测定的含硫酯类物质,优化样品前处理技术和色谱检测条件,并进行方法学论证。结果衍生后的二硫代氨基甲酸盐类农药在一定范围内,其浓度的平方与GC-FPD(S)的响应峰高呈良好的线性关系。福美铁、丙森锌、代森锰的最低检出浓度分别为0.1、0.5、0.5 mg/kg,平均回收率为75.3%～94.6%,相对标准偏差最大为14.1%。结论该方法操作简便、定量准确可靠,各项指标均符合农药残留检测的要求。     

Optimisation and validation of a specific analytical method for the determination of thiram residues in fruits and vegetables by LC–MS/MS

Thiram is a non-systemic dithiocarbamate fungicide, which is easily degraded during sample preparation since it is affected by pH, matrix components and temperature. In this work, specific methodology for thiram analysis in vegetable (eggplant and lettuce) and fruit (strawberry) samples has been developed based on liquid chromatography coupled to tandem mass spectrometry (LC–MS/MS). Minimising thiram degradation during standards storage and sample preparation was carefully studied. The effect of low temperature (about 5 °C), addition of a dehydrating agent (Na₂SO₄ anhydrous), pH regulator (NaHCO₃), and enzymatic activity reduction (EDTA) during extraction was evaluated. The optimised procedure was validated for eggplants, lettuces, and strawberries. Satisfactory recoveries, between 80% and 106%, and relative standard deviations below 10% were obtained at 0.1 and 0.01 mg/kg fortification levels (n = 5). Limits of detection below 0.0012 mg/kg were achieved. The validated method has been applied to eggplant and lettuce samples collected from different field trials as well as several strawberry and apple samples.     

Headspace solid-phase microextraction for the evaluation of pesticide residue contents in cucumber and strawberry after washing treatment

The headspace solid-phase microextraction was developed to examine the organophosphorus (diazinon, malathion, chloropyrifos, quinalphos, profenofos) and organochlorine (chlorothalonil, α-endosulfan and β-endosulfan) pesticide residues in vegetable (cucumber) and fruit (strawberry) samples. The effects of washing by different solutions were evaluated for the reduction of organophosphorus and organochlorine pesticide residues contents. Gas chromatography with electron capture detection was used to analysis the investigated pesticides. The results showed that washing by a non-toxic solution can decrease the concentration of pesticide residues in the fruit and vegetable samples. The data further indicated that acetic acid was the most effective solution in removing the residues of the investigated pesticides from the fruit and vegetable samples when compared to sodium carbonate, sodium chloride and tap water. The amount of pesticides removed by solution washing is related to their water solubility and vapour pressure properties.     

Miniaturization of the QuEChERS Method in the Fast Gas Chromatography-Tandem Mass Spectrometry Analysis of Pesticide Residues in Vegetables

Two reduced-scale quick, easy, cheap, effective, rugged, and safe (QuEChERS) procedures, combined with fast gas chromatography-triple quadrupole mass spectrometry (GC-QqQ MS), were developed and then validated for the determination of 35 pesticides in different vegetable products (tomatoes, zucchini, red peppers, and lettuce). The proposed reduced-scale methods, involving the use of only 3 g of sample, were compared with an official European Union method, namely EN15662:2008, based on the use of a 10-g sample. Method validation was performed considering the following figures of merit: recovery, linearity, precision, matrix effects, and limits of detection and quantification. Specifically, recovery was in the 67–126% range, regression coefficients were between 0.991 and 0.999, and coefficients of variation were between 1 and 13% (at the 50 μg kg^?1 level), while limits of quantification were always below European legislation residue limits. Additionally, the measurement of matrix effects confirmed the necessity of matrix-matched calibration. The developed QuEChERS GC-QqQ MS method is both simple and rapid (analysis of six samples in 2.5 h) and is sensitive enough for EU regulation purposes. To demonstrate the applicability of the proposed reduced-scale method, multi-residue analysis was performed on 20 samples.