PCR experion automated electrophoresis system to detect <i>Listeria monocytogenes</i> in foods

Delibato, Elisabetta; Gattuso, Antonietta; Minucci, Angelo; Auricchio, Bruna; Medici, Dario De; Toti, L.; Castagnola, Massimo; Capoluongo, Ettore; Gianfranceschi, Monica Virginia

doi:10.1002/jssc.200900166

Cited by 27 publications

(22 citation statements)

References 23 publications

(25 reference statements)

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“…4.0.0) based on reference sequence NM_173957. The size of the RGN amplicon (100 bp) was verified using an automated electrophoresis station (Experion Instrument, Bio-Rad), and DNA analysis was performed using the Experion DNA 1K Analysis Kit (Bio-Rad) [63].…”

Section: Methodsmentioning

confidence: 99%

Regucalcin Expression in Bovine Tissues and Its Regulation by Sex Steroid Hormones in Accessory Sex Glands

et al. 2014

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Regucalcin (RGN) is a mammalian Ca2+-binding protein that plays an important role in intracellular Ca2+ homeostasis. Recently, RGN has been identified as a target gene for sex steroid hormones in the prostate glands and testis of rats and humans, but no studies have focused on RGN expression in bovine tissues. Thus, in the present study, we examined RGN mRNA and protein expression in the different tissues and organs of veal calves and beef cattle. Moreover, we investigated whether RGN expression is controlled through sex steroid hormones in bovine target tissues, namely the bulbo-urethral and prostate glands and the testis. Sex steroid hormones are still illegally used in bovine husbandry to increase muscle mass. The screening of the regulation and function of anabolic sex steroids via modified gene expression levels in various tissues represents a new approach for the detection of illicit drug treatments. Herein, we used quantitative PCR, western blot and immunohistochemistry analyses to demonstrate RGN mRNA and protein expression in bovine tissues. In addition, estrogen administration down-regulated RGN gene expression in the accessory sex glands of veal calves and beef cattle, while androgen treatment reduced RGN gene expression only in the testis. The confirmation of the regulation of RGN gene expression through sex steroid hormones might facilitate the potential detection of hormone abuse in bovine husbandry. Particularly, the specific response in the testis suggests that this tissue is ideal for the detection of illicit androgen administration in veal calves and beef cattle.

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Section: Methodsmentioning

confidence: 99%

Regucalcin Expression in Bovine Tissues and Its Regulation by Sex Steroid Hormones in Accessory Sex Glands

et al. 2014

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“…Wild-type and deleted mRNA before and after siRNA treatment was evaluated by capillary electrophoresis (Experion automated electrophoresis station, 21 Bio-Rad Laboratories) of reverse-transcriptase PCR products (F1, R1, and R2 primers, supplemental Table 1). …”

Section: Silencingmentioning

confidence: 99%

The dominant-negative von Willebrand factor gene deletion p.P1127_C1948delinsR: molecular mechanism and modulation

Casari

Pinotti

Lancellotti

et al. 2010

Blood

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Understanding molecular mechanisms in the dominant inheritance of von Willebrand disease would improve our knowledge of pathophysiologic processes underlying its prevalence. Cellular models of severe type 2 von Willebrand disease, caused by a heterozygous deletion in the von Willebrand factor (VWF) gene, were produced to investigate the altered biosynthesis. Coexpression of the wild-type and in-frame deleted (p.P1127_C1948delinsR) VWF forms impaired protein secretion, high molecular weight multimer formation and function (VWF collagen-binding 1.9% ؎ 0.5% of wild-type), which mimicked the patient's phenotype. mRNA, protein, and cellular studies delineated the highly efficient dominant-negative mechanism, based on the key role of heterodimers as multimer terminators. The altered VWF, synthesized in large amounts with the correctly encoded "cysteine knot" domain, formed heterodimers and heterotetramers with wild-type VWF, in addition to deleted homodimers. Impaired multimerization was associated with reduced amounts of VWF in late endosomes. Correction of the dominant-negative effect was explored by siRNAs targeting the mRNA breakpoint, which selectively inhibited the in-frame deleted VWF expression. Although the small amount of the deleted protein synthesized after inhibition still exerted dominant, even though weakened, negative effects, the siRNA treatment restored secretion of large multimers with improved function (VWF collagen-binding 28.0% ؎ 3.3% of wild-type). (Blood. 2010; 116(24):5371-5376) IntroductionDominant inheritance characterizes von Willebrand disease (VWD), the most prevalent 1 disorder of hemostasis, and particularly type 1 VWD, a quantitative deficiency, and type 2 VWD, which includes a large and still increasing group of von Willebrand factor (VWF) qualitative deficiencies. 2 Differently, the rare and severe type 3 VWD is recessively inherited. Based on functional and structural defects impairing platelet adhesion or factor VIII binding, type 2 has been classified into 4 subtypes: A, B, M, and N. 2 Type 2A refers to qualitative defects characterized by a decreased proportion of large multimers 3 and by numerous mutations interfering with the multiple biosynthetic steps of the protein maturation, subgrouped in IIA, IIC, IID, and IIE. 2,4,5 We have previously described a large, de novo, and heterozygous deletion in VWF gene associated with severe type 2 VWD, 6 characterized by reduced VWF antigen levels, impaired function, and virtual absence of high molecular weight multimers (HMWMs). The altered VWF mRNA 7 (Figure 1) was supposed to encode an in-frame deleted protein, the characterization of which would contribute to define its dominant-negative features.VWF dimerization and multimerization provide a rationale for the dominant inheritance, caused by the interaction of wild-type and mutant monomers during polymerization. The strength of the dominant-negative effects produced by VWF variants [8][9][10][11][12] would be associated with the ample variation of residual VWF level and function, and w...

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“…The detection and monitoring of foodborne pathogens by food industries and food safety authorities facilitate outbreak prevention [1] . Over 25% of food productions worldwide are lost every year due to food contaminations [2] .…”

Section: Introductionmentioning

confidence: 99%

“…A detecção e monitoramento dos patógenos alimentares por parte das indústrias alimentícias e autoridades de segurança alimentar facilitam a prevenção de surtos [1] . Mais de 25% das produções alimentares em todo o mundo são perdidas todos os anos devido a contaminação alimentar [2] .…”

Section: Introductionunclassified

Comparison between Listeria monocytogenes single and multipathogen detection methods

Souza¹,

Ferreira²,

Pasquini-Netto³

et al. 2017

BBR

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143 Biomedical and Biopharmaceutical Research J o r n a l d e I n v e s t i g a ç ã o B i o m é d i c a e B i o f a r m a c ê u t i c a Comparison between Listeria monocytogenes single and multipathogen detection methods Comparação entre os métodos de detecção de Listeria monocytogenes simples e multipatógenos AbstractMultipathogen detection methods have been used to provide a broad range of microorganism detection. We aimed to identify the most searched pathogens in L. monocytogenes detection methods and evaluate if there are sensitivity and specificity differences between single and multipathogen methods. A systematic review was performed by including studies carried out in order to detect L. monocytogenes in a broad range of food sample. A total of 2770 records were retrieved, of which 191 were selected. The majority of the studies (104) presented multipathogen detection, other 87 searched for L. monocytogenes specifically. From the studies for multipathogen detection; other Listeria species (19.4%), Salmonella spp. (21.6%), E. coli O157:H7 (13.2%), Staphylococcus spp. (9.7%), E. coli (5.7%) were more frequently reported. Sensitivity and specificity calculations were derived from only 26 studies, because they compared their data with an official method. Sensitivity and specificity values were close to 100%, showing that others characteristics such as time and cost should be considered to evaluate alternative methods in further research. As a conclusion, evidence generated regarding L. monocytogenes identification methods contribute to method improvements and listeriosis control.Keywords: L. monocytogenes, methods, pathogens, sensitivity, specificity. ResumoOs métodos de detecção para múltiplos patógenos têm sido usados para fornecer uma ampla faixa de detecção de microrganismos. Objetivou-se identificar os patógenos mais buscados nos métodos de detecção para L. monocytogenes e avaliar se existem diferenças nas taxas de sensibilidade e especificidade entre os métodos que buscaram identificar somente L. monocytogenes e os para múltiplos patógenos. Uma revisão sistemática foi realizada incluindo estudos desenvolvidos para detectar L. monocytogenes em amostras alimentares.

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PCR experion automated electrophoresis system to detect Listeria monocytogenes in foods

Cited by 27 publications

References 23 publications

Regucalcin Expression in Bovine Tissues and Its Regulation by Sex Steroid Hormones in Accessory Sex Glands

Regucalcin Expression in Bovine Tissues and Its Regulation by Sex Steroid Hormones in Accessory Sex Glands

The dominant-negative von Willebrand factor gene deletion p.P1127_C1948delinsR: molecular mechanism and modulation

Comparison between Listeria monocytogenes single and multipathogen detection methods

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