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Loop-Mediated Isothermal Amplification (LAMP) for the Detection of Listeria monocytogenes and Major Pathogenic Serotypes  [PDF]
Ana Paula Rocha da Costa, Mariana de Lira Nunes, Carina Lucena Mendes-Marques, Alzira Maria Paiva de Almeida, Nilma Cintra Leal
American Journal of Analytical Chemistry (AJAC) , 2014, DOI: 10.4236/ajac.2014.516112
Abstract: Rapid identification and characterization of Listeria monocytogenes are required for the food industry, epidemiological studies, and disease prevention and control. However, typing procedures are labor-intensive and time-consuming, and they require technical expertise, a panel of sera and reference culture strains or sophisticated and expensive equipment. To improve upon traditional diagnostic methods for L. monocytogenes we developed and evaluated an efficient procedure for the specific identification of L. monocytogenes and the major pathogenic serotypes of the species based on loop-mediated isothermal amplification (LAMP). Four individual reactions were designed using primers targeting any L. monocytogenes serotypes (LAMP-AS) and the 1/2a (LAMP-1/2a), 1/2b (LAMP-1/2b), and 4b (LAMP-4b) serotypes. The procedure distinguished L. monocytogenes from closely genetically related species and the targeted serotypes. Cross-reactivity with a few rare serotypes isolated from food or clinical samples did not impair the usefulness of the procedure. Thus, our approach constitutes a fast, easy and low-cost alternative for L. monocytogenes diagnosis and serotyping and may be useful for surveillance and epidemiological investigation programs.
Different sensitivity of various serotypes of Listeria monocytogenes to lactic acid bacteria bacteriocins
Dimitrijevi? Mirjana,Teodorovi? V.,Balti? Milan ?.,Karabasil Ne?eljko
Acta Veterinaria , 2004, DOI: 10.2298/avb0403201d
Abstract: In this study the sensitivity of various serotypes of Listeria monocytogenes towards five lactic bacteria bacteriocins was investigated, at two incubation temperatures (37oC for 24 h and 4oC for 12 days); six serotypes were indentified in 50 clinical/human isolates of L. monocytogenes and eight among 48 isolates from foodstuffs. This microorganism was found in many foodstuffs in numerous studies during the last decade. Among the methods for typing Listeria, the greatest attention has been dedicated to serological typing which defines the basic characteristics of Listeria antigens. Namely, it is well known that bactericideal or bacteriostatic effects of bacteriocins are not only expressed towards closely related bacterial species, but also towards less closely related Gram positive bacteria, such as L. monocytogenes. Bacteriocins can be eventually added to food, with the aim of decreasing the risk of listeriosis to the minimum. It was discovered that the bacteriocins, originating from Lactococcus UW and Lactobacillus sake 148 did not express inhibitory effects on any Listeria serotypes, while those bacteriocins originating from Lactobacillus sake 265, Pediococcus 347 and Lactobacillus sake 706 had a listericidal effect towards almost every assessed serotype. The highest bactericid effect was expressed by bacteriocin towards serotypes 4c and 4, at 37oC after 24h incubation and towards serotypes 1/2b and 4b after 12 days incubation at 4oC. Thus, the incubation temperature and time influenced the inhibitory effects of bacteriocins.
Listeria monocytogenes serotypes in human infections (Italy, 2000-2010)
Pontello,Mirella; Guaita,Anna; Sala,Giuliana; Cipolla,Micaela; Gattuso,Antonietta; Sonnessa,Michele; Gianfranceschi,Monica Virginia;
Annali dell'Istituto Superiore di Sanità , 2012, DOI: 10.4415/ANN_12_02_07
Abstract: background: in developed countries invasive listeriosis is an infection of great concern to public health to due its clinical severity and high fatality rate, despite its low incidence. in europe, statistically significant increasing trends in listeriosis notification rates from 2005 to 2009 were noted in austria, denmark, hungary, italy, spain and sweden. materials and methods: the standardized techniques based on phenotype to typing listeria monocytogenes is the serotyping. in europe, as elsewhere in the world, about 95% of l. monocytogenes strains isolated from clinical and food samples belongs to serovars 1/2a, 1/2b, 1/2c and 4b. results: the target of this work is to draw attention to this important and atypical foodborne disease, reporting epidemiological data and serotypes distribution of 251 human l. monocytogenes isolates reported during 2000-2010 to veterinary public health and food safety department of istituto superiore di sanità, focusing on epidemiological trend of invasive listeriosis in lombardia, a north italian region. the serotypes most frequently identified are 1/2a, 4b, 1/2b (in total 92%), but the detection of uncommon serotypes is not missing (1/2c, 3a, 3b, 4d). conclusions: in italy the surveillance laboratory network, as well as the foodborne disease network (enter-net), has revealed in the last 11 years an increase trend of listeriosis cases reported likewise with results of notificable national infectious disease surveillance system. this is probably due to a real increase of listeriosis, even if there is a greater sensitivity of the network in some regions.
Antimicrobial activity of some of the south-Indian spices against serotypes of Escherichia coli, Salmonella, Listeria monocytogenes and Aeromonas hydrophila
Indu, M.N.;Hatha, A.A.M.;Abirosh, C.;Harsha, U.;Vivekanandan, G.;
Brazilian Journal of Microbiology , 2006, DOI: 10.1590/S1517-83822006000200011
Abstract: antibacterial activity of extracts of allium sativum (garlic), myristica fragrans (nutmeg), zingiber officinale (ginger), allium cepa (onion) and piper nigrum (pepper) has been evaluated against 20 different serogroups of escherichia coli, 8 serotypes of salmonella, listeria monocytogenes and aeromonas hydrophila. garlic extract showed excellent antibacterial activity against all the test organisms, except l. monocytogenes. nutmeg showed good anti-listerial activity, although activity against e. coli and salmonella were serotype dependent. both garlic and nutmeg extracts were effective against a. hydrophila. extracts of ginger showed inhibitory activity against two serogroups of e. coli: as o8 (enterotoxigenic e. coli) and o88 only. extracts of onion and pepper did not show any antibacterial activity against the test organisms.
Antimicrobial activity of some of the south-Indian spices against serotypes of Escherichia coli, Salmonella, Listeria monocytogenes and Aeromonas hydrophila  [cached]
Indu M.N.,Hatha A.A.M.,Abirosh C.,Harsha U.
Brazilian Journal of Microbiology , 2006,
Abstract: Antibacterial activity of extracts of Allium sativum (garlic), Myristica fragrans (nutmeg), Zingiber officinale (ginger), Allium cepa (onion) and Piper nigrum (pepper) has been evaluated against 20 different serogroups of Escherichia coli, 8 serotypes of Salmonella, Listeria monocytogenes and Aeromonas hydrophila. Garlic extract showed excellent antibacterial activity against all the test organisms, except L. monocytogenes. Nutmeg showed good anti-listerial activity, although activity against E. coli and Salmonella were serotype dependent. Both garlic and nutmeg extracts were effective against A. hydrophila. Extracts of ginger showed inhibitory activity against two serogroups of E. coli: as O8 (enterotoxigenic E. coli) and O88 only. Extracts of onion and pepper did not show any antibacterial activity against the test organisms.
Estudio mediante PCR múltiple de serotipos de Listeria monocytogenes aislados en Argentina Study by multiplex PCR of Listeria monocytogenes serotypes isolated in Argentine  [cached]
R. Callejo,M. Prieto,C. Martínez,L. Aguerre
Revista argentina de microbiolog?-a , 2008,
Abstract: Se comparó una PCR múltiple recientemente validada para la caracterización de serotipos de Listeria monocytogenes con el método tradicional de serotipificación. Se estudiaron 342 aislamientos de origen humano, alimentario, veterinario y ambiental obtenidos durante el período 1992-2005. La concordancia entre ambos métodos para los serotipos 1/2a, 1/2b y 1/2c fue del 100%, y para el serotipo 4b fue del 98%. La serotipificación constituye una herramienta importante como primer nivel de diferenciación de cepas de L. monocytogenes para llevar a cabo la vigilancia epidemiológica y, sobre todo, el estudio de brotes. La PCR múltiple es una técnica alternativa rápida, de bajo costo y fácilmente adaptable en laboratorios de bacteriología clínica y bromatología. A multiplex PCR assay, recently validated to characterize the serotypes of Listeria monocytogenes was evaluated in comparison to conventional serotyping. Three hundred forty two L. monocytogenes strains isolated from human, food, animal and environmental sources during the 1992-2005 period were assayed. The concordance between the two methods for serotypes 1/2a, 1/2b and 1/2c was 100%, whereas for serotype 4b it was 98%. Serotyping is a useful tool for first line strain differentiation during epidemiological surveillance and outbreaks. The multiplex PCR assay offers a fast and low-cost alternative, which is easily adaptable to clinical bacteriology and bromatology laboratories.
Distribution of Listeria monocytogenes serotypes isolated from foods, Colombia, 2000-2009 Distribución de serotipos de Listeria monocytogenes aislados de alimentos, Colombia, 2000-2009  [cached]
Ana Isabel Mu?oz
Biomédica , 2012,
Abstract: Introduction. Listeria monocytogenes is an intracellular, opportunistic pathogen which can cause severe infections such as meningitis, encephalitis and bacteremia, and also abortions in human beings. Foods are the vehicle for infection of the host.Serotypification has discriminated thirteen serotypes: 1/2a,1/2b, 1/2c, 3a, 3b, 3c, 4a, 4ab, 4b, 4c, 4d, 4e, 7. 4b is the cause of the majority of cases of listeriosis in the world.Objective. To determine the frequency of serotypes of L. monocytogenes isolated from foods during 2000-2009 in Colombia. Materials and methods. The study is descriptive and retrospective. 1599 isolates were studied, all of which were confirmed as Listeria monocytogenes and other strains of Listeria, using biochemical tests recommended by the Food and Drug Administration (USA) and API Listeria, and serotyped using the Seeliger and H hne method. Results. Of the 1,599 isolates, 1,424 were confirmed as L. monocytogenes. Serotypes identified were: 1/2a, 135 (9,5%); 1/2b, 154 (10,8%); 1/2c, 68 (4,8%); 3a, 4 (0,3%); 3b, 29 (2,0%); 3c, 2 (0,1%); 4a, 44 (3,1%); 4b, 820 (57,6%); 4c, 6 (0,4%); 4d- 4e, 140 (9,8%); 4e, 17 (1,2%); 7, 2 (0,1%); not susceptible of serotypification, three cases, (0,2%). Isolates came mainly from the Capital District of Bogotá, 1035 (73%); Antioquia 199 (14%), Nari o, 109 (8%); Valle del Cauca 50 (3,5%) and from other departments 33 (2,3%).Conclusion. At the analyzed isolates, 1,424 (89%) belonged to L. monocytogenes, showing a good quality in isolations and identification, most of this isolates belonged to serotype 4b, 820 (57,6%), highly virulent.It is recommended the obligatory surveillance of this microorganism. Introducción. La Listeria monocytogenes es un patógeno, facultativo intracelular, oportunista, causante de graves infecciones en humanos como meningitis, encefalitis y bacteremias; también es causa de abortos. Los alimentos actúan como medio de transporte para infectar al huésped.La serotipificación ha discriminado trece serotipos: 1/2a,1/2b, 1/2c, 3a, 3b, 3c, 4a, 4ab, 4b, 4c, 4d, 4e, 7. El 4b es causante de la mayoría de listeriosis en el mundo.Objetivo. Determinar la frecuencia de los serotipos de L. monocytogenes aislados de alimentos, durante los a os 2000-2009, en Colombia.Materiales y métodos. Estudio descriptivo retrospectivo. Se analizaron 1.599 aislamientos, los cuales fueron confirmados como Listeria monocytogenes y otras especies de Listeria, con pruebas bioquímicas recomendadas por la Food Drug Administration (USA) y API Listeria, y serotipificadas con la metodología de Seeliger and H hne.Re
A study on the inactivated bivalent vaccine prepared from serotypes 1/2a and 4b Listeria monocytogenes for the control of listeriosis in sheep
Baci? Dragan,Obrenovi? Sonja,Dimitrijevi? B.,Joni? B.
Acta Veterinaria , 2012, DOI: 10.2298/avb1206531b
Abstract: In this study, the protective effects of two bivalent inactivated vaccines were evaluated. Vaccines were prepared from Listeria monocytogenes, serotypes 1/2a and 4b, as the most frequent in our and surrounding epidemiological areas. Vaccine A consists of whole L. monocytogenes bacteria cells, inactivated with 0.4% formaldehyde and aluminium hydroxide as a carrier. Vaccine B contains 0.1% saponin in addition to ingredients of vaccine A. Evaluations of these vaccines were performed in 60 sheep, divided into four groups (n=10) with a corresponding negative control group (n=5). After 14 days, boosterisation of all animals was performed. In order to evaluate the immune response, blood samples were obtained every 14 days during the next 6 months. Antibody titres were determined by microaglutitation (MAT) and complement fixation tests (CFT). Comparative analyses of antibody titres, induced by vaccines A and B, show that the latter (with saponine) significantly increased the level of antibody titres (p<0.01). The levels of immune response were also significantly impacted by the total number of bacteria and vaccine dosage (p<0.01). The bivalent vaccine containing 0.1% saponin (vaccine B) in 5.0 mLx 106 cfu/mL (colony-forming units per milliliter) dosage shows a protective effect after challenge with L. monocytogenes. The protective levels of this antibody were 1/80 and 1/16, determined by MAT and CFT, respectively. Antibody titres were significantly higher after boosterisation (p<0.01) and protective levels could be detected in the sera of vaccinated animals during the next 6 months. Therefore, it is strongly recommended to perform boosterisation two weeks after the initial vaccination. [Projekat Ministarstva nauke Republike Srbije, br. 31085 i br. 31088]
Investigations of protective effects of bivalent inactivated vaccine prepared from serotypes 1/2a and 4b Listeria monocytogenes on mice  [PDF]
Baci? Dragan,Obrenovi? Sonja,Kirovski Marko,Dimitrijevi? Blagoje
Veterinarski Glasnik , 2012, DOI: 10.2298/vetgl1202049b
Abstract: The objective of these investigations was to check on laboratory white mice the protective effect of an inactivated bivalent vaccine prepared from serotypes 1/2a and 4b L. monocytogenes. Following verification of the sterility and toxicity of the prepared vaccine, the mice were divided into 6 groups with 10 animals in each group. The first and second group of mice were administered the vaccine without saponin (vaccine A) and the third and fourth group the vaccine with saponin (vaccine B). Mice of the fifth and the sixth group were not vaccinated and served as a negative control. Two weeks following vaccination, the experimental groups were revaccinated, with the exception of the two control groups. Two weeks following revaccination, all groups were artificially infected with serotypes 1/2a and 4b L. monocytogenes. During the course of the investigations (60 days) a total of 4 mice died in the vaccinated groups. Mice of the control groups started dying after day 7, and the last mouse in these groups died 14 days after the infection. Examinations of preparations of parenchymatous organs of the dead mice stained according to Gram proved the presence of L. monocytogenes. Homogenates of parenchymatous organs were sown on tryptose agar for reisolation and a pure culture of L. monocytogenes was obtained. Through the use of specific antiserums, serotypes 1/2a and 4b were confirmed. Considering the total number of vaccinated mice in the experiment and the percent deaths (10%), it can be said that the investigated vaccine with saponin had a satisfactory protective effect. [Projekat Ministarstva nauke Republike Srbije, br. TR-20142 i TR-31088]
PATOGENESIS DE Listeria monocytogenes, MICROORGANISMO ZOONOTICO EMERGENTE.
Torres,Kirvis; Sierra,Sara; Poutou,Raúl; Carrascal,Ana; Mercado,Marcela;
Revista MVZ Córdoba , 2005,
Abstract: listeria monocytogenes in addition to being a paradigm for the immunological investigation has become in an appropriate model system to the analysis of the molecular mechanisms of the intracellular parasitism of other bacteria strains. inmunologyst were interested in this microorganism when was recognized as a risky organism for public health and the food industry security. from mid of the 80's scientists have researched the molecular biology of virulence markers of this microorganism, the cellular biology of the interactions of these markers with the receptors of the host cell, the cytoskeleton, the transduction signals routes and the mechanisms of immunity mediated by the host cells. the intention of this review is to describe some taxonomic and phylogenetics characteristics of listeria monocytogenes, the human and animal incidence of several serotypes, the physiopathology of the infection, animals models and cellular culture employed for virulence studies, the risk populations, clinical manifestations of human and animal listeriosis, the treatment, the genetic organization and evolution de the virulence markers, the mechanisms used to interact with the host cell, the mechanism to escape from the cellular death processes and to pass through an infected cell to another one. the compiled information is from great importance for the health personnel, consumers and risk population; reason for which listeria monocytogenes it is a pathogen that represents a threat for world-wide the public health.
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