Assessment of the virulence of Listeria monocytogenes: agreement between a plaque-forming assay with HT-29 cells and infection of immunocompetent mice.

Some Listeria monocytogenes strains not related to clinical cases have been found to exhibit a low virulence level in mice as well as in an in vitro test using Caco-2 cells. The purpose of this study was to validate a new in vitro test of virulence based on a plaque-forming assay (PFA) using a HT-29 cell monolayer with 118 Listeria strains. The use of HT-29 cells in 96-well tissue culture plates allowed the testing of 30 strains per day and providing results in 24 h. In addition. statistical analyses demonstrated the reproducibility and repeatability of the PFA. No quantitative relationship was observed between the virulence of the strains and the hemolytic titer or the cytotoxic effects on HT-29 cells. In contrast, good agreement was observed between virulence assessed after subcutaneous (SC) infection and virulence obtained by PFA. Three groups of L. monocytogenes strains (avirulent, hypovirulent and fully virulent) were established by comparison of the clinical origin of the strains, the number of immunocompetent contaminated mice and the numbers of Listeria strains recovered in the spleen after SC infection. With one exception, i.e. a clinical case of L. seeligeri (sensitivity 0.98), the PFA successfully detected the virulent strains only (specificity 1). Decision-tree algorithms performed by SAS and S-Plus demonstrated that this tissue culture assay discriminated between the avirulent and hypovirulent strains and the virulent strains. This test could therefore be an alternative to in vivo tests, allowing grading of virulence.     

A QUANTITATIVE MICROTITER PLATE HEMOLYSIS ASSAY FOR LISTERIA MONOCYTOGENES

All virulent strains of L. monocytogenes produce the extracellular SH-activated hemolysin, listeriolysin O, while nonhemolytic strains of L. monocytogenes are avirulent suggesting that the expression of this hemolysin is necessary for virulence of L. monocytogenes. Hence, testing for hemolysis becomes clinically relevant for an isolate identified as L. monocytogenes. However, the quantification and interpretation of this characteristic on blood agar poses several problems. Hence we have proposed a simple quantitative microtiter plate hemolysis assay. The assay is made of SRBC (3% SRBC in PBS) in microtiter plates to which CASO-cultured cell-free supernatants of L. monocytogenes or other test cultures are added. After mixing, the plates are incubated at 37C for 15–30 min and the hemolysis is visually read as CHU and MHU, as well as by a 620 nm scan for absorbancy. Percent hemolysis is calculated. We feel that this assay should prove to be of significance in a microbiology laboratory in which routine hemolysis assays are conducted.     

Hypovirulent Listeria monocytogenes strains are less frequently recovered than virulent strains on PALCAM and Rapid' L. mono media

Several selective media have been developed to detect Listeria monocytogenes contaminated foodstuffs. Polymyxin-acriflavine-LiCl-ceftazidime-aesculin-mannitol (PALCAM) and Oxford media, required for the EN ISO method 11 290-1, are used for the detection of Listeria spp. in 2 days based on the expression of esculinase activity. Selective agar media such as Rapid' L. mono and Agar Listeria according to Ottaviani and Agosti (ALOA), based on the activity of phosphatidylinositol phospholipase C (PI-PLC) that allows the specific detection of L. monocytogenes in 2 days, are also used. However, no medium can assess the level of virulence of L. monocytogenes strains. Using a plaque-forming assay followed by subcutaneous footpad inoculation in mice, 15 virulent, 8 hypovirulent and 17 avirulent strains were discriminated among L. monocytogenes strains mainly originating from food (36/40). Their growth was tested on the four selective media. After 2 days, the number of colony forming units (cfu) of all the virulent strains was significantly superior to the number obtained with avirulent strains on all the four media tested, and superior to the number obtained with hypovirulent strains on PALCAM and Oxford media. These results showed a relationship between the level of virulence of L. monocytogenes strains and their growth on the selective agar media tested. Moreover, 1 out of 8 hypovirulent and 5 out of 17 avirulent strains did not grow on Rapid' L. mono medium, and 1 hypovirulent and 8 avirulent strains grew but did not express PI-PLC activity during the 7 days of incubation. The lack of detection of PI-PLC activity on Rapid' L. mono was not related to a gene mutation since these strains expressed enzymatic activity on ALOA medium, which detected up to 92% of the hypo- and avirulent strains. In contrast, some of these strains without growth or enzymatic activity expression would not be detected with PALCAM and Rapid' L. mono in foodstuffs on the second day.     

Pathogenicity and production of virulence factors by Listeria monocytogenes isolates from channel catfish

Pathogenicity of four channel catfish Listeria monocytogenes isolates (CCF1, CCF4, HCC7, and HCC23) was examined in a comparative manner with virulent type strains L. monocytogenes ATCC 19115 and EGD and avirulent type strain ATCC 15313 in BDF and A/J mice. Isolates HCC7 and CCF1 (both serovar 1) caused similar percent mortalities and 50% lethal concentration values when compared with virulent type strains and were therefore considered pathogenic. The presence of the virulence factors listeriolysin (LLO), phosphotidylcholine-phospholipase (PC-PLC), and phosphotidylinositol-phospholipase (PI-PLC) was determined using specific activity tests. The virulent catfish isolates were positive for production of LLO, PC-PLC, and PI-PLC. However, catfish isolate HCC23 was not virulent in mice despite being hemolytic, suggesting that not every hemolytic L. monocytogenes strain is virulent. With the exception of HCC7, all virulent strains displayed enhanced LLO production in a special stress medium, whereas almost undetectable LLO activity was present when catfish isolates and virulent type strain L. monocytogenes were grown in a rich medium such as brain heart infusion. Avirulent strains were found to lack or have decreased expression of LLO, PC-PLC, or PI-PLC.     

单核细胞增生李斯特菌菌株分型与其致病潜力基因相关性研究进展

单核细胞增生李斯特菌是一种常见的食源性致病菌,该菌分型繁多。不同分型的单核细胞增生李斯特菌致病潜力不同,这与菌株所携带的抗性基因和毒力基因不同有关。本文综述了不同分型单核细胞增生李斯特菌与抗性基因和毒力基因的关系,结果发现与该菌抗性相关的基因,如热抗性、耐冷性、酸抗性、耐高渗、耐干燥、耐金属和杀菌剂及参与应激蛋白表达调控的基因较多,它们与分型之间的关系暂无明确相关性结论,但也发现应激生存岛（stress survival island,SSI）-1仅存在于谱系I和谱系II部分菌株中,SSI-2目前仅被发现在ST121菌株中携带。从功能毒力基因和李斯特菌毒力基因岛（Listeria pathogenicity islands,LIPI）两方面分述了毒力基因,LIPI-3和LIPI-4主要存在于谱系I菌株中。此外,ST5、ST8、ST9和ST121菌株的inlA基因中出现提前终止密码子,使得菌株的毒力降低。研究不同分型单核细胞增生李斯特菌致病潜力基因的差异有助于单核细胞增生李斯特菌的预警预测、防控,并为不同分型菌株致病机制的深入研究提供参考。     

Virulence characterization and genotypic analyses of Listeria monocytogenes isolates from food and processing environments in eastern China

In this study, twenty L. monocytogenes food-related isolates collected from eastern China Zhejiang province were compared by in vivo LD50 assays as well as in vitro cytopathic plaque forming assay. Nineteen L. monocytogenes isolates (19/20) were as virulent as reference strain 10403S, while the isolate M4 had low pathogenicity. The unique isolate M4 fell into lineage III based on the partial nucleotide variations of actA, while the other isolates belonged to the more common lineages I and II. L. monocytogenes isolates were grouped in 17 to 19 subtypes using pulsed-field gel electrophoresis (PFGE) with SmaI digestion, and multilocus sequence typing (MLST) based on three virulence genes (actA, inlA and inlB) and four housekeeping genes (betL, dat, recA and sigB). The virulence genes based MLST had better discriminatory power than that targeting the housekeeping genes (0.990 vs 0.895), similar to PFGE (0.976). An isolate from the processing desk was found having the same pulsotype as the two isolates from final shrimp products in the same plant, indicating that process contamination could be the source of Listeria contamination.     

应用PCR技术快速测定食品中单核细胞增生李斯特菌毒力

目的：采用PCR 技术准确、快速测定单核细胞增生李斯特菌(单增李斯特氏菌)分离株的毒力基因,鉴别强毒株和弱毒株或无毒株,以有效地限制李斯特氏菌病的传播。方法：以单增李斯特氏菌相关毒力基因(hly、plcB、inlA、inlB、inlC、inlJ、prfA)设计引物,检测重庆市2007 - 2009 年分离的40 株单增李斯特氏菌分离株中的毒力基因携带率,并进行小鼠毒力实验。结果：40 株分离株中有15 株7 种毒力基因检测结果均为阳性,6 株hly 基因阴性,4 株plcB 基因阴性,4 株prfA 基因性,5 株inlA、inlB 基因阴性,11 株inlC 基因阴性,9 株inlJ基因阴性,1 株inlA、inlB、inlC、inlJ 基因均为阴性。4 株分离株的毒力与标准菌株ATCC191161E 相当,小鼠LD50 在1.2 × 108～6.0 × 108CFU/mL 之间,为强毒株。筛选出弱毒株09-132,LD50 为1.7 × 1011CFU/mL。结论：重庆市存在发生李斯特菌食物中毒的潜在危险,内化素基因(inlA、inlB、inlC、inlJ)的缺失可能是导致菌株毒力降低的原因,而prfA 和plcB 基因与菌株毒力的相关性较小。     

Nonsense-mutated inlA and prfA not widely distributed in Listeria monocytogenes isolates from ready-to-eat seafood products in Japan

InlA is a surface protein participating in the entry of Listeria monocytogenes into mammalian non-phagocytic cells. PrfA is a positive regulatory factor that regulates the expression of a set of virulence genes. Recent studies revealed that some L. monocytogenes strains have a truncated form of these proteins because of nonsense mutations in their sequences, and these truncations contribute to the significant reduction in virulence of this pathogen. In this study, sequence analyses of inlA and prfA among L. monocytogenes isolated from ready-to-eat seafood revealed that only one out of 59 isolates had a nonsense-mutated inlA and all had non-mutated prfA. This indicated that these strains could be fully virulent based on the sizes of these proteins.     

台州市食源性单核细胞增生李斯特菌分子特征研究

目的了解台州市食品中分离的单核细胞增生李斯特菌的血清型、毒力基因以及基因分型情况,建立食源性单核细胞增生李斯特菌的分子特征本底信息,为食源性疾病的防治提供技术支持。方法对近几年从食品中分离的37株单核细胞增生李斯特菌进行多重PCR血清分型、9种毒力基因(prf A、inl A、inl B、iap、fla A、hly A、plc B、mpl和act A)PCR检测、PFGE基因分型,用Bio Numerics 6.6软件对分型数据进行聚类分析。结果 37株食源性单核细胞增生李斯特菌的血清型以1/2a或3a型别为主;所有菌株均检出4种以上毒力基因,有15株菌携带所有9种毒力基因;37株菌经Apa I酶切PFGE分型后,共得到22种带型,每种带型包含1~5株不等,相似度区间为67%~100%。结论台州市食源性单核细胞增生李斯特菌存在致病流行风险,建立的指纹图谱数据库可为食源性疾病的防治提供技术支持。     

A simple method for the differentiation of Listeria monocytogenes based on induction of lecithinase activity by charcoal

The PlcB phospholipase C, or lecithinase, is an important listerial virulence factor of potential use as a pathogenicity marker for Listeria spp. food isolates. However, wild-type strains of Listeria monocytogenes express virulence factors very poorly in vitro and their lecithinase activity is normally difficult to detect on agar plates. We recently reported that the production of listerial virulence factors is strongly induced if L. monocytogenes is grown in the presence of activated charcoal. We report here a simple method for the rapid differentiation of L. monocytogenes from other Listeria spp. based on a comparison of lecithinase reactions in egg yolk agar plates with and without charcoal supplementation. All L. monocytogenes wild-type isolates tested showed a clear induction of lecithinase activity in charcoal-supplemented medium (CEYM), while nonpathogenic Listeria spp. remained negative in CEYM. The animal pathogen L. ivanovii was easily differentiated from L. monocytogenes because it showed a strong lecithinase reaction independently of the presence or absence of charcoal in the medium.     

Comparison of Listeria monocytogenes virulence in a mouse model

Listeriosis results from exposure to the foodborne pathogen Listeria monocytogenes. Although many different strains of L. monocytogenes are isolated from food, no definitive tests currently predict which isolates are most virulent. The objectives of this study were to address two major data gaps for risk assessors, variability among L. monocytogenes strains in pathogenicity and virulence. Strains used in our monkey clinical trial or additional food isolates were evaluated for their virulence and infectivity in mice. All strains were equally pathogenic to immunocompromised mice, causing deaths to 50% of the population 3 days after exposure to doses ranging from 2 to 3 log CFU. Doses resulting in 50% deaths on the fifth day after administration were 1 to 2 log lower than those on the third day, indicating that the full course of pathogenicity exceeds the 3-day endpoint in immunocompromised mice. Three strains were chosen for further testing for their virulence and infectivity in liver and spleen in normal (immunocompetent) mice. Virulence was not significantly different (P > 0.05) among the three strains, all resulting in deaths to 50% of mice at 5 to 7 log CFU by 5 days after administration. All strains were equally infective in liver or spleen, with higher numbers of L. monocytogenes directly correlated with higher doses of administration. In addition, there was no preference of organs by any strains. The lack of strain differences may reflect the limitation of the mouse model and suggests the importance of using various models to evaluate the pathogenicity and virulence of L. monocytogenes strains.     

Characterization of Listeria monocytogenes isolates in import food products of China from 8 provinces between 2005 and 2007

A total of 48 Listeria monocytogenes isolates of different import food products from 8 provinces between 2005 and 2008 were characterized. The serotype and virulence were confirmed for each strain and molecular subtyping were analyzed by multilocus sequence typing (MLST). Twenty five strains were assigned to serotype 1/2a, and 11 isolates to serotype 1/2b, serotype 4b were found in 7 isolate, and the remaining 5 strains were grouped into serotypes 1/2c, 4a, and 4e. Molecular subtyping schemes found thirty two sequence types (STs) among these isolates and the majority of L. monocytogenes strains belonged to lineage II (56%), followed by lineage I (38%), lineage III (6%). Two molecular subtype clusters, cluster A included all isolates of lineage II, while cluster B contained the isolates of lineages I and lineages III. Two L. monocytogenes strains were not grouped in either of the two clusters. Fifty three isolates were as virulent as L. monocytogenes reference strain EGD in mouse virulence assay, while the isolates 22213 and 22265 had low pathogenicity. These results provide the first molecular insight into the L. monocytogenes strains isolated from import food products of 8 provinces in China and indicate the potential risk to cause human disease if intake by contaminated foods. MLST could be used as a routine subtyping method of L. monocytogenes isolates. In China, inspection and quarantine strategies of imported foods should be strengthened. PRACTICAL APPLICATION: There is a potential risk of listeriosis in China and routine subtyping of L. monocytogenes isolates is important. It is necessary for food hygiene management to strengthen the supervision of imported foods.     

云南省牛乳中单核细胞增生李斯特氏菌的分布特征、耐药性及毒力研究

目的分析云南省牛乳中单核细胞增生李斯特氏菌(Listeria monocytogenes)的分布特征、耐药性及毒力现状。方法采集云南省主要奶产区牛乳样本,按照GB 4789.30—2016方法分离单核细胞增生李斯特氏菌疑似菌株,采用飞行时间质谱技术结合16SrDNA测序对疑似菌株进行鉴定;采用微量肉汤稀释法对分离菌株进行抗生素药敏实验;通过聚合酶链式反应(polymerasechainreaction,PCR)技术检测单核细胞增生李斯特氏菌中7种毒力基因的存在状况。结果158份样品共有4份检出单核细胞增生李斯特氏菌,检出率为2.53%;共分离获得8株菌。其中,生鲜乳中检出率为3.36%,巴氏杀菌乳中未检出。在生鲜乳中,荷斯坦牛乳、水牛乳、牦牛乳检出率分别为2.13%、0.00%和20.00%;标准化牛场、奶牛合作社和个体户的检出率分别为2.50%、1.51%和15.38%;手工挤奶和机械挤奶检出率为11.76%和1.96%。药敏实验结果显示8株菌均对青霉素耐药,对四环素、复方新诺明、红霉素耐药率分别为87.50%、75.00%和50.00%;多重耐药率为75.00%;但所有菌株对氨苄西林均敏感。8株菌对7种毒力基因的携带程度从12.50%~75.00%不等。结论云南省牛乳尤其生鲜乳中存在一定程度的单核细胞增生李斯特氏菌污染,且该菌具有普遍耐药性并携带一定程度的毒力基因,对消费者的安全具有潜在风险,应加强卫生监督管理。     

Intragastric inoculation with a cocktail of Listeria monocytogenes strains does not potentiate the severity of infection in A/J mice compared to inoculation with the individual strains comprising the cocktail

Although multistrain cocktails of Listeria monocytogenes are used in food inoculation experiments, no studies, to our knowledge, have been reported that use these cocktails in an intragastric mouse model. In this study, we used a five-strain L. monocytogenes cocktail consisting of strains Scott A, MFS108, 101M, V7, and 310 and a four-strain L. monocytogenes cocktail containing strains Scott A, EGD, H7738, and F2365. Here, we report that intragastric inoculation of anesthetized mice with approximately 106 CFU of a cocktail of L. monocytogenes strains does not result (P > 0.05) in a more severe infection (on the basis of the CFU of Listeria spp. recovered from the spleen, liver, and blood) than inoculation of mice with similar numbers of the individual strains comprising the cocktail. Nor did we observe any consistent relationship between susceptibility of L. monocytogenes strains to inactivation in synthetic gastric fluid in vitro and virulence in mice.     

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。     

Detection of multiple virulence-associated genes in Listeria monocytogenes isolated from bovine mastitis cases

Clinical samples (n=725) were collected from bovines (n=243) which were positive for mastitis using the California mastitis test (CMT) and somatic cell count (SCC). The clinical samples comprising blood (n=239), milk (n=243), and faecal swabs (n=243) were examined for the presence of pathogenic Listeria spp. Isolation of the pathogen was done using selective enrichment in University of Vermont Medium and plating onto Dominguez-Rodriguez isolation agar. Confirmation of the isolates was based on biochemical tests and Christie, Atkins, Munch-Petersen (CAMP) test followed by pathogenicity testing. Pathogenicity of the isolates was tested by phosphatidylinositol-specific phospholipase C (PI-PLC) assay as well as in vivo tests namely, chick embryo and mice inoculation tests. The isolates were subjected to PCR assay for five virulence-associated genes, plcA, prfA, hlyA, actA and iap. Listeria spp. were isolated from 12 (1.66%) samples. Of these 4 (0.55%) and 1 (0.14%) were confirmed as Listeria monocytogenes and Listeria ivanovii, respectively. L. monocytogenes and L. ivanovii were recovered from milk samples (2) and faecal (3) of mastitic cattle (3) and buffaloes (2). L. monocytogenes recovered from the milk of mastitic cattle and L. ivanovii from the faecal swab of buffalo turned out to be pathogenic. However, the remaining three hemolytic isolates exhibiting positive CAMP test turned out to be negative in PI-PLC assay, chick embryo and mice inoculation. L. monocytogenes and L. ivanovii isolates characterized as pathogenic by PI-PLC assay and in vivo pathogenicity tests were found to possess all the five virulence-associated genes and three genes, plcA, prfA and actA respectively. The remaining three hemolytic but non-pathogenic L. monocytogenes isolates were negative for plcA by PCR. It seems that the plcA gene and its expression (in the PI-PLC assay) have an important role as virulence determinants in pathogenic Listeria spp. In conclusion, the PI-PLC assay and virulence genes targeted PCR (plcA, prfA and hlyA genes for L. monocytogenes and plcA, prfA and actA genes for L. ivanovii) hold a good promise as rapid and reliable in vitro alternatives to in vivo pathogenicity tests.     

Processing plant persistent strains of Listeria monocytogenes appear to have a lower virulence potential than clinical strains in selected virulence models

Listeria monocytogenes is an important foodborne bacterial pathogen that can colonize food processing equipment. One group of genetically similar L. monocytogenes strains (RAPD type 9) was recently shown to reside in several independent fish processing plants. Persistent strains are likely to contaminate food products, and it is important to determine their virulence potential to evaluate risk to consumers. We compared the behaviour of food processing persistent and clinical L. monocytogenes strains in four virulence models: Adhesion, invasion and intracellular growth was studied in an epithelial cell line, Caco-2; time to death in a nematode model, Caenorhabditis elegans and in a fruit fly model, Drosophila melanogaster and fecal shedding in a guinea pig model. All strains adhered to and grew in Caco-2 cells in similar levels. When exposed to 10(6) CFU/ml, two strains representing the persistent RAPD type 9 invaded Caco-2 cells in lower numbers (10(2)-10(3) CFU/ml) as compared to the four other strains (10(4)-10(6) CFU/ml), including food and human clinical strains. In the D. melanogaster model, the two RAPD type 9 strains were among the slowest to kill. Similarly, the time to reach 50% killed C. elegans worms was longer (110 h) for the RAPD type 9 strains than for the other four strains (80 h). The Scott A strain and one RAPD type 9 strain were suspended in whipping cream before being fed to guinea pigs and the persistent RAPD type 9 strain was isolated from feces in a lower level (approximately 10(2) CFU/g) than the Scott A strain (approximately 10(5) CFU/g) (P<0.05). The addition of NaCl has been shown to cause autoaggregation and increases adhesion of L. monocytogenes to plastic. However, growth in the presence of NaCl did not alter the behaviour of the tested L. monocytogenes strains in the virulence models. Overall, the two strains representing a very common fish processing plant persistent group (RAPD type 9) appear to have a lower virulence potential in all four virulence models than Scott A and a strain isolated from a clinical case of listeriosis.     

Comparison of the cell surface properties and growth characteristics of Listeria monocytogenes and Listeria innocua

Growth kinetics and physicochemical surface properties were compared for three Listeria strains with differing degrees of virulence: L. monocytogenes LO28; its isogenic, nonhemolytic mutant L. monocytogenes Bof415; and a nonvirulent species, L. innocua (strain Lin9). The influences of growth stage (mid-exponential phase, early stationary phase, and mid-stationary phase) and culture temperature (20 and 37 degrees C) were assessed by determining the electrical properties and the hydrophobic-hydrophilic and Lewis acid-base characteristics of the three strains. L. innocua, although taxonomically very similar to L. monocytogenes, exhibited physicochemical surface properties that differed significantly from those of L. monocytogenes LO28 and L. monocytogenes Bof415. Indeed, under our experimental conditions, L. innocua cells presented a more marked electronegative character (particularly when cultured at 20 degrees C), as well as greater variability in their Lewis acid-base characteristics as a function of temperature and growth stage. Furthermore, the growth kinetics of the three strains revealed the onset of a decay phase after 16 h of culture at 37 degrees C for the L monocytogenes Bof415 mutant. All of these results demonstrate that under our experimental conditions, the growth and/or physicochemical characteristics of the slightly pathogenic or nonpathogenic Listeria strains (Bof415 and Lin9) differed from those of the virulent strain (L. monocytogenes LO28). Consequently, the use of Listeria strains recognized as nonvirulent appeared to provide a model that was not fully suitable for simulating the bioadhesive behavior of the pathogenic strains involved in foodborne diseases.     

ISOLATION,PURIFICATION AND PARTIAL CHARACTERIZATION OF A NEW EXTRACELLULAR CYTOTOXIN FROM A VIRULENT CLINICAL STRAIN OF LISTERIA MONOCYTOGENES SEROTYPE 4b,AND AN AVIRULENT,NONHEMOLYTIC VARIANT ATCC 15313 SEROTYPE 1/2a1

A number of pathologies oflisteric infections in man remain unexplained. The objective of this study was to investigate whether strains of Listeria monocytogenes synthesize additional toxin(s), which may be implicated in these unexplained pathologies. Results showed that both virulent (4b) and avirulent (ATCC 15313) strains of L. monocytogenes synthesize a previously unrecognized extracellular cytotoxin of Mr 34,000. Cytotoxin synthesis is possibly dependent on the type of salt and the concentration of calcium ions. The cytotoxin is highly thermostable (D_121C > 10 min) but is susceptible to digestion by trypsin/chymotrypsin. The cytotoxin is thermostable as the absence of viable L. monocytogenes cells in a food does not preclude the possibility of a toxigenic food product. The role of the cytotoxin in a listeric infection is at present unknown.