Creation and resolution of non-B-DNA structural impediments during replication.

During replication, folding of the DNA template into non-B-form secondary structures provides one of the most abundant impediments to the smooth progression of the replisome. The core replisome collaborates with multiple accessory factors to ensure timely and accurate duplication of the genome and epigenome. Here, we discuss the forces that drive non-B structure formation and the evidence that secondary structures are a significant and frequent source of replication stress that must be actively countered. Taking advantage of recent advances in the molecular and structural biology of the yeast and human replisomes, we examine how structures form and how they may be sensed and resolved during replication.

HLA-Cw6 allele and biologic therapy are protective factors against liver fibrosis in psoriatic arthritis patients.

OBJECTIVES: To evaluate the association between liver fibrosis and the HLACw6 allele in psoriatic arthritis (PsA) patients. METHODS: A retrospective longitudinal study involving PsA patients with determination of the HLA-Cw6 allele was performed. Liver fibrosis was estimated by using the FIB-4 (fibrosis-4) score. A multivariate logistic model was undertaken to assess the odds ratio (OR), with its 95% confidence interval, of liver fibrosis after adjustment for potential confounding factors. RESULTS: A total of 209 PsA patients were included: 25.3% HLA-Cw6 were positive, 59.8% were receiving biological disease-modifying anti-rheumatic drugs (bDMARDs), 29.6% had arterial hypertension (AHT), 24% dyslipidaemia, and 4.2% acute myocardial infarction (AMI). The HLA-Cw6 allele was more frequent in PsA patients with normal FIB-4 values (p=0.024), as opposed to AHT (p=0.002), AMI (p=0.023) and dyslipidaemia (p=0.030), which were found more frequently in subjects with altered FIB-4 values. The presence HLA-Cw6 and the use of bDMARDs were confirmed as protective factors against liver fibrosis (OR 0.210, 0.062-0.707, p=0.012 and OR 0.397, 0.166-0.949, p=0.038, respectively). Conversely, AHT emerged as a risk factor (OR 2.973, 1.125-7.858, p=0.028). CONCLUSIONS: In PsA, the HLA-Cw6 allele and bDMARDs behave as protective factors for liver fibrosis, while AHT is an independent risk factor.

Partnering with immigrant families to promote language justice and equity in education.

Despite US federal legislation mandates institutions to provide meaningful access and participation to students and families in educational settings, culturally and linguistically diverse (CLD) families and caregivers of children in special education experience cultural and linguistic barriers. A Community Advisory Team (CAT) of parents, advocates, community interpreters and translators, researchers, and teachers explored CLD families' experiences and advocacy efforts. Critical bifocality and circuits of dispossession, privilege, and resistance informed the documentation of inequities and resistance to understand the linkages of structural arrangements of power. Focus groups with families (n = 21) speakers of Spanish, Portuguese, and Cantonese were conducted. Findings indicate perceived discrimination, poor and inadequate interpretation and translation services impact children's access to special education services, hinder family's communication with schools and reduce the perceptions of schools as trustworthy institutions. Families advocate relentlessly for their children and recommend schools listen to families and hire culturally and linguistically competent interpreters and translators. Community psychologists can make significant contributions to promote language justice in education settings through participatory approaches to inquiry that value CLD families' knowledge and expertise.

Eradication of Staphylococcus aureus Post-Sternotomy Mediastinitis Following the Implementation of Universal Preoperative Nasal Decontamination With Mupirocin: An Interrupted Time-Series Analysis.

BACKGROUND: Although presurgical nasal decontamination with mupirocin (NDM) has been advocated as a measure for preventing postsurgical mediastinitis (PSM) due to Staphylococcus aureus, this strategy is not universally recommended due to lack of robust supporting evidence. We aimed to evaluate the role of preoperative NDM in the annual incidence of S. aureus PSM at our institution. METHODS: An interrupted time-series analysis, with an autoregressive error model, was applied to our single-center cohort by comparing preintervention (1990-2003) and postintervention (2005-2018) periods. Logistic regression was performed to analyze risk factors for S. aureus PSM. RESULTS: 12 236 sternotomy procedures were analyzed (6370 [52.1%] and 5866 [47.9%] in the pre- and postintervention periods, respectively). The mean annual percentage adherence to NDM estimated over the postintervention period was 90.2%. Only 4 of 127 total cases of S. aureus PSM occurred during the 14-year postintervention period (0.68/1000 sternotomies vs 19.31/1000 in the preintervention period; P < .0001). Interrupted time-series analysis demonstrated a statistically significant annual reduction in S. aureus PSM of -9.85 cases per 1000 sternotomies (-13.17 to -6.5; P < .0001) in 2005, with a decreasing trend maintained over the following 5 years and an estimated relative reduction of 84.8% (95% confidence interval [CI], 89.25-74.09%). Chronic obstructive pulmonary disease was the single independent risk factor for S. aureus PSM (odds ratio, 3.7; 95% CI, 1.72-7.93) and was equally distributed in patients undergoing sternotomy during pre- or postintervention periods. CONCLUSIONS: Our experience suggests the implementation of preoperative NDM significantly reduces the incidence of S. aureus PSM.

Internal standard application strategies in mass spectrometry imaging by desorption electrospray ionization mass spectrometry.

RATIONALE: We developed a model case study to evaluate three internal standard (IS) application strategies (methods I-III) using the psycholeptic phenobarbital (PB) and the isotopically labelled IS phenobarbital-D5 (PB-D5) from in vitro dosed tissues of the golden apple snail (Pomacea diffusa) by desorption electrospray ionization mass spectrometry imaging (DESI-MSI). METHODS: In method I, the IS was deposited as microspots on top of 10 µm thick snail tissues; in method II, a thin IS film was applied; and in method III, the IS was spiked into the DESI solvent spray. DESI-MSI analyses were performed using a Thermo LTQ mass spectrometer equipped with a custom-built DESI source and two-dimensional moving stage. PB (m/z 231) and PB-D5 (m/z 236) were monitored in selected ion monitoring mode between m/z 227 and 239. RESULTS: The analytical performance of two IS strategies (methods I and II) in DESI-MSI was evaluated based on an intra- and inter-day precision assay, an accuracy assessment, and statistical analysis. In the inter-day DESI-MSI assay, method I exhibited better precision (6.5%-7.4%) than method II (10.7%-17.6%) between 10 and 100 ng/µL. In the accuracy assessment, PB quality controls of 75 ng/µL were back-calculated as 71 ± 4 and 83 ± 9 ng/µL, resulting in relative errors of -5% and 11% for methods I and II, respectively. Method III did not work under the experimental design and was not evaluated. CONCLUSIONS: Three IS application strategies were investigated and compared for a routine quantitative DESI-MSI approach. Methods I and II were not statistically significantly different as shown by a Bland-Altman plot, suggesting that these two methods can be used interchangeably. However, method III requires further research for future quantitative DESI-MSI analyses.

Fragmentation study of clerodane diterpenes from Casearia species by tandem mass spectrometry (quadrupole time-of-flight and ion trap).

RATIONALE: Clerodane-type diterpenes from Casearia species show important pharmacological activites such as antitumor, antimicrobial and anti-inflamatory. There are several mass spectrometry (MS)-based methods for identification of diterpenes; however, there is still a lack of MS procedures capable of providing characteristic fragmentation pathways for a rapid and unambiguous elucidation of casearin-like compounds. METHODS: Casearin-like compounds were investigated by electrospray ionization tandem mass spectrometry (ESI-MS/MS). The fragmentation studies were carried out by tandem mass spectrometry in space (quadrupole time-of-flight (QTOF)) using different collision energies and also by tandem mass spectrometry in time (QIT) by selective isolation of product ions. RESULTS: Casearin-like compounds presented a predominance of sodium- and potassium-cationized precursor ions. Both QIT and QTOF techniques provided sequential neutral losses of esters related to the R1 to R5 substituents linked to the nucleus of the clerodane diterpenes. The fragmentation pathway is initiated with a cleavage of the ester moieties R2 followed by the elimination of the ester groups R3 , both losing neutral carboxylic acids. Using QIT, it was also possible to observe the cleavage of the ester groups R1 or R5 by MS4 experiments. CONCLUSIONS: Through a rational analysis of the fragmentation mechanisms of Casearia diterpenes it was possible to suggest an annotation strategy based on the sequential cleavages of the ester groups related to the R2 , R3 and R5 substituents. These results will assist studies of the dereplication and metabolomics involving casearin-like compounds present in complex extracts of Casearia species.

Rapid structural characterisation of benzylisoquinoline and aporphine alkaloids from Ocotea spixiana acaricide extract by HPTLC-DESI-MSn.

INTRODUCTION: Lauraceae alkaloids are a structurally diverse class of plant specialised secondary metabolites that play an important role in modern pharmacotherapy, being useful as well as model compounds for the development of synthetic analogues. However, alkaloids characterisation is challenging due to low concentrations, the complexity of plant extracts, and long processes for accurate structural determinations. OBJECTIVE: The use of high-performance thin layer chromatography coupled with desorption electrospray ionisation multistage mass spectrometry (HPTLC DESI-MSn ) as a fast tool to identify alkaloids present in Ocotea spixiana extract and evaluate the extract's acaricide activity. METHODS: Ocotea spixiana twigs were extracted by conventional liquid-liquid partitioning. HPTLC analysis of the ethyl acetate extract was performed to separate isobaric alkaloids prior to DESI-MSn analysis, performed from MS3 up to MS7 . The extract's acaricide activity against Rhipicephalus microplus was evaluated by in vitro (larval immersion test) and in silico tests. RESULTS: HPTLC-DESI-MSn analysis was performed to identify a total of 13 aporphine and four benzylisoquinoline-type alkaloids reported for the first time in O. spixiana. In vitro evaluation of the extract and the alkaloid boldine showed significant activity against R. microplus larvae. It was established in silico that boldine had important intermolecular interactions with R. microplus acetylcholinesterase enzyme. CONCLUSION: The present study demonstrated that HPTLC-DESI-MSn is a useful analytical tool to identify isoquinoline alkaloids in plant extracts. The acaricide activity of the O. spixiana ethyl acetate extract can be correlated to the presence of alkaloids.

Review and perspectives on the applications of mass spectrometry imaging under ambient conditions.

Ambient mass spectrometry (AMS)-based techniques are performed under ambient conditions in which the ionization and desorption occur in the open environment allowing the direct analysis of molecules with minimal or no sample preparation. A selected group of AMS techniques demonstrate imaging capabilities that can provide information about the localization of molecules on complex sample surfaces such as biological tissues. 2D, 3D, and multimodal imaging have unlocked an array of applications to systematically address complex problems in many areas of research such as drug monitoring, natural products, forensics, and cancer diagnostics. In the present review, we summarize recent advances in the field with respect to the implementation of new ambient ionization techniques and current applications in the last 5 years. In more detail, we mainly focus on imaging applications in topics related to animal whole bodies and tissues, single cells, cancer diagnostics and biomarkers, microbial cultures and co-cultures, plant and natural product metabolomics, and forensic applications. Finally, we discuss new areas of research, future perspectives, and the overall direction that the field may take in the years to come.

Rapid determination of the tumour stroma ratio in squamous cell carcinomas with desorption electrospray ionization mass spectrometry (DESI-MS): a proof-of-concept demonstration.

Squamous cell carcinomas constitute a major class of head & neck cancers, where the tumour stroma ratio (TSR) carries prognostic information. Patients affected by stroma-rich tumours exhibit a poor prognosis and a higher chance of relapse. As such, there is a need for a technology platform that allows rapid determination of the tumour stroma ratio. In this work, we provide a proof-of-principle demonstration that Desorption Electrospray Ionization Mass Spectrometry (DESI-MS) can be used to determine tumour stroma ratios. Slices from three independent mouse xenograft tumours from the human FaDu cell line were subjected to DESI-MS imaging, staining and detailed analysis using digital pathology methods. Using multivariate statistical methods we compared the MS profiles with those of isolated stromal cells. We found that m/z 773.53 [PG(18:1)(18:1) - H]-, m/z 835.53 [PI(34:1) - H]- and m/z 863.56 [PI(18:1)(18:0) - H]- are biomarker ions that can distinguish FaDu cancer from cancer associated fibroblast (CAF) cells. A comparison with DESI-MS analysis of controlled mixtures of the CAF and FaDu cells showed that the abundance of the biomarker ions above can be used to determine, with an error margin of close to 5% compared with quantitative pathology estimates, TSR values. This proof-of-principle demonstration is encouraging and must be further validated using human samples and a larger sample base. At maturity, DESI-MS thus may become a stand-alone molecular pathology tool providing an alternative rapid cancer assessment without the need for time-consuming staining and microscopy methods, potentially further conserving human resources.

Correction: Rapid determination of the tumour stroma ratio in squamous cell carcinomas with desorption electrospray ionization mass spectrometry (DESI-MS): a proof-of-concept demonstration.

Correction for 'Rapid determination of the tumour stroma ratio in squamous cell carcinomas with desorption electrospray ionization mass spectrometry (DESI-MS): a proof-of-concept demonstration' by Michael Woolman et al., Analyst, 2017, 142, 3250-3260.

Reactive DESI-MS imaging of biological tissues with dicationic ion-pairing compounds.

This work illustrates reactive desorption electrospray ionization mass spectrometry (DESI-MS) with a stable dication on biological tissues. Rat brain and zebra fish tissues were investigated with reactive DESI-MS in which the dictation forms a stable bond with biological tissue fatty acids and lipids. Tandem mass spectrometry (MS/MS) was used to characterize the dication (DC9) and to identify linked lipid-dication compounds formed. The fragment m/z 85 common to both DC9 fragmentation and DC9-lipid fragmentation was used to confirm that DC9 is indeed bonded with the lipids. Lipid signals in the range of m/z 250-350 and phosphoethanolamines (PE) m/z 700-800 observed in negative ion mode were also detected in positive ion mode with reactive DESI-MS with enhanced signal intensity. Reactive DESI-MS imaging in positive ion mode of rat brain and zebra fish tissues allowed enhanced detection of compounds commonly observed in the negative ion mode.

Imprint Desorption Electrospray Ionization Mass Spectrometry Imaging for Monitoring Secondary Metabolites Production during Antagonistic Interaction of Fungi.

Direct analysis of microbial cocultures grown on agar media by desorption electrospray ionization mass spectrometry (DESI-MS) is quite challenging. Due to the high gas pressure upon impact with the surface, the desorption mechanism does not allow direct imaging of soft or irregular surfaces. The divots in the agar, created by the high-pressure gas and spray, dramatically change the geometry of the system decreasing the intensity of the signal. In order to overcome this limitation, an imprinting step, in which the chemicals are initially transferred to flat hard surfaces, was coupled to DESI-MS and applied for the first time to fungal cocultures. Note that fungal cocultures are often disadvantageous in direct imaging mass spectrometry. Agar plates of fungi present a complex topography due to the simultaneous presence of dynamic mycelia and spores. One of the most devastating diseases of cocoa trees is caused by fungal phytopathogen Moniliophthora roreri. Strategies for pest management include the application of endophytic fungi, such as Trichoderma harzianum, that act as biocontrol agents by antagonizing M. roreri. However, the complex chemical communication underlying the basis for this phytopathogen-dependent biocontrol is still unknown. In this study, we investigated the metabolic exchange that takes place during the antagonistic interaction between M. roreri and T. harzianum. Using imprint-DESI-MS imaging we annotated the secondary metabolites released when T. harzianum and M. roreri were cultured in isolation and compared these to those produced after 3 weeks of coculture. We identified and localized four phytopathogen-dependent secondary metabolites, including T39 butenolide, harzianolide, and sorbicillinol. In order to verify the reliability of the imprint-DESI-MS imaging data and evaluate the capability of tape imprints to extract fungal metabolites while maintaining their localization, six representative plugs along the entire M. roreri/T. harzianum coculture plate were removed, weighed, extracted, and analyzed by liquid chromatography-high-resolution mass spectrometry (LC-HRMS). Our results not only provide a better understanding of M. roreri-dependent metabolic induction in T. harzianum, but may seed novel directions for the advancement of phytopathogen-dependent biocontrol, including the generation of optimized Trichoderma strains against M. roreri, new biopesticides, and biofertilizers.

DNA replication initiation shapes the mutational landscape and expression of the human genome.

The interplay between active biological processes and DNA repair is central to mutagenesis. Here, we show that the ubiquitous process of replication initiation is mutagenic, leaving a specific mutational footprint at thousands of early and efficient replication origins. The observed mutational pattern is consistent with two distinct mechanisms, reflecting the two-step process of origin activation, triggering the formation of DNA breaks at the center of origins and local error-prone DNA synthesis in their immediate vicinity. We demonstrate that these replication initiation-dependent mutational processes exert an influence on phenotypic diversity in humans that is disproportionate to the origins' genomic size: By increasing mutational loads at gene promoters and splice junctions, the presence of an origin significantly influences both gene expression and mRNA isoform usage. Last, we show that mutagenesis at origins not only drives the evolution of origin sequences but also contributes to sculpting regulatory domains of the human genome.

Chemical profiling and separation of bioactive secondary metabolites in Maca (Lepidium peruvianum) by normal and reverse phase thin layer chromatography coupled to desorption electrospray ionization-mass spectrometry.

Maca is a Peruvian tuberous root of the Brassicaceae family grown in the central Andes between altitudes of 4000 and 4500 m. The medicinal plant is a nutraceutical with important biological activities and health effects. In this study, we report a rapid high-performance thin layer chromatography (HPTLC)-(-)desorption electrospray ionization (DESI)-mass spectrometry (MS) method to profile and separate intact glucosinolates without prior biochemical modifications from the hydromethanolic extracts of two phenotypes, red and black Maca (Lepidium peruvianum) seeds. In the first stage of the plant's life cycle, aromatic glucosinolates were the main chemical constituents whereby six aromatic, three indole, and one aliphatic glucosinolate were tentatively identified. At the seedling stage, glucolepigramin/Glucosinalbin was the most predominant precursor, rather than Glucotropaeolin, which is mainly found in hypocotyls and roots. These findings lead us to suggest that glucolepigramin/glucosinalbin play a major role as active precursors in the biosynthetic pathways of other secondary metabolites in the early stages of plant development. Between red and black Maca seeds, only minor differences in the relative abundances of glucosinolates were observed rather than different plant metabolites. For the first time, we report six potential plant antibiotics, phytoanticipins: glycosylated ascorbigens and dihydroascorbigens from Maca seeds. We also investigated a targeted reverse phase C18 functionalized TLC-DESI-MS method with high sensitivity and specificity for Brassicaceae fatty acids in Maca seeds and health supplements such as black Maca root lyophilized powder and tinctures. The investigation of secondary metabolites by normal and reverse phase TLC-DESI-MS methods, described in this study, can aid in their identification as they begin to emerge in later stages of development in plant tissues such as leaves, hypocotyls, and roots.

Phosphatidylcholine restores neuronal plasticity of neural stem cells under inflammatory stress.

The balances between NSCs growth and differentiation, and between glial and neuronal differentiation play a key role in brain regeneration after any pathological conditions. It is well known that the nervous tissue shows a poor recovery after injury due to the factors present in the wounded microenvironment, particularly inflammatory factors, that prevent neuronal differentiation. Thus, it is essential to generate a favourable condition for NSCs and conduct them to differentiate towards functional neurons. Here, we show that neuroinflammation has no effect on NSCs proliferation but induces an aberrant neuronal differentiation that gives rise to dystrophic, non-functional neurons. This is perhaps the initial step of brain failure associated to many neurological disorders. Interestingly, we demonstrate that phosphatidylcholine (PtdCho)-enriched media enhances neuronal differentiation even under inflammatory stress by modifying the commitment of post-mitotic cells. The pro-neurogenic effect of PtdCho increases the population of healthy normal neurons. In addition, we provide evidences that this phospholipid ameliorates the damage of neurons and, in consequence, modulates neuronal plasticity. These results contribute to our understanding of NSCs behaviour under inflammatory conditions, opening up new venues to improve neurogenic capacity in the brain.

Identification of Sassafras albidum alkaloids by high-performance thin-layer chromatography tandem mass spectrometry and mapping by desorption electrospray ionization mass spectrometry imaging.

Sassafras albidum is an important tree species that occurs across North America. The presence of benzylisoquinoline and aporphine alkaloids has been previously described; however, the spatial distribution of these compounds within S. albidum and other plants of Lauraceae family is still unclear. Mass spectrometry imaging has become an important tool in analysis of plants metabolites, uncovering important contributions about the functional role, biosynthetic pathway, and accumulation of these compounds in the plant. This work aimed to identify further alkaloids present in S. albidum roots, twigs, and leaves by high-performance thin-layer chromatography coupled to desorption electrospray ionization multistage mass spectrometry (HPTLC DESI-MSn ) and to map the spatial distribution of these compounds by DESI-MS imaging. A total of 12 alkaloids were indentified in the roots and twigs, and six of them were detected for the first time in S. albidum. A high number of alkaloids was found in S. albidum roots; however, alkaloids were not detected in the leaves. Cross sections of roots and twigs were blotted onto TLC plates assisted by heating and solvent extraction, and these imprints were analyzed by DESI-MS imaging. The profile of alkaloid spatial distribution in DESI-MS images showed different accumulation patterns across and within different plant parts. Most alkaloids displayed higher intensities in the outer-most layer of the roots and twigs. The detailed spatial localization pattern of these alkaloids analyzed by DESI-MS imaging in different plant parts could contribute to a better understanding of the profile of distribution, accumulation, and biosynthesis of benzylisoquinoline and aporphine alkaloids.

Chemical and sensory profile of new genotypes of Brazilian Coffea canephora.

The study of Brazilian Conilon coffee genotypes with unknown chemical composition and sensory quality is extremely important since these data may contribute to the launching of new coffee cultivars in the international market with high cup quality. The present study aimed to investigate the metabolic profile of 3 genotypes of Conilon and compared them to Robusta Tropical and Arabica coffees, all collected at 3 different levels of ripeness. The extracts were analysed by ESI-LTQ-ORBITRAP, and 11 attributes were evaluated by sensory analysis. To correlate sensory, composition and maturation, chemometric analysis was used. The metabolites trigonelline, caffeine, caffeoylquinic acid and sugars revealed higher concentrations in genotypes 105 and 108. According to the sensorial analysis, genotype 108 showed the highest final score (82), which was even higher than the Arabica coffees. Among the new coffees studied, genotype 108 presented promising characteristics, sparking interest in its national and international commercialization.

Repositioning Salirasib as a new antimalarial agent.

Malaria is a serious tropical disease that kills thousands of people every year, mainly in Africa, due to Plasmodium falciparum infections. Salirasib is a promising cancer drug candidate that interferes with the post-translational modification of Ras. This S-farnesyl thiosalicylate inhibits isoprenylcysteine carboxyl methyltransferase (ICMT), a validated target for cancer drug development. There is a high homology between the human and the parasite enzyme isoforms, in addition to being a druggable target. Looking to repurpose its structure as an antimalarial drug, a collection of S-substituted derivatives of thiosalicylic acid were prepared by introducing 1,2,3-triazole as a diversity entry point or by direct alkylation of the thiol. We further investigated the in vitro toxicity of FTS analogues to Plasmodium falciparum in the asexual stages and in Vero cells. An antiplasmodial activity assay was performed using a simple, high-sensitivity methodology based on nanoluciferase (NLuc)-transfected P. falciparum parasites. The results showed that some of the analogs were active at low micromolar concentration, including Salirasib. The most potent member of the series has S-farnesyl and the 1,2,3-triazole moiety substituted with phytyl. However, the compound substituted with methyl-naphthyl shows promising physicochemical and activity values. The low cytotoxicity in eukaryotic cells of the most active analogs provided good therapeutic indices, being starting-point candidates for future antimalarial drug development.

Thermostability of oxytetracycline, tetracycline, and doxycycline at ultrahigh temperatures.

The thermostability parameters of three tetracycline antibiotics at high and ultrahigh temperatures (110-140 degrees C) as well as the influence of treatment medium pH and water activity on their thermotolerance have been investigated. The thermal degradation of the three antibiotics followed a first-order reaction kinetic within the 1.5-2 log(10) cycles investigated. A linear relationship was observed between the log of the DT values and the treatment temperature. The temperature dependence of the DT values was similar for the three molecules (z=28+/-2 degrees C). DT values of doxycycline were approximately 1.5 and 3 times higher than those of tetracycline and oxytetracycline, respectively. Changes in the treatment medium pH (7.0-4.0) and water activity (0.99-0.93) scarcely varied the antibiotics' thermal stability. Only when doxycycline was heat-treated at pH 4.0 did its thermal resistance increase by 3 times. The thermostability parameters obtained would allow the effect of different cooking and sterilization procedures to be estimated. Whereas low-temperature-long-time treatments (conventional sterilization) would destroy >98% of the initial concentration of the residues of the three antibiotics, high-temperature-short-time treatments (UHT) would leave unaltered residues in the 50-90% range.

Evaluation of a Method for Rapid Detection of Listeria monocytogenes in Dry-Cured Ham Based on Impedanciometry Combined with Chromogenic Agar.

The food industry is in need of rapid, reliable methodologies for the detection of Listeria monocytogenes in ready-to-eat products, as an alternative to the International Organization of Standardization (ISO) 11290-1 reference method. The aim of this study was to evaluate impedanciometry combined with chromogenic agar culture for the detection of L. monocytogenes in dry-cured ham. The experimental setup consisted in assaying four strains of L. monocytogenes and two strains of Listeria innocua in pure culture. The method was evaluated according to the ISO 16140:2003 standard through a comparative study with the ISO reference method with 119 samples of dry-cured ham. Significant determination coefficients ( R2 of up to 0.99) for all strains assayed in pure culture were obtained. The comparative study results had 100% accuracy, 100% specificity, and 100% sensitivity. Impedanciometry followed by chromogenic agar culture was capable of detecting 1 CFU/25 g of food. L. monocytogenes was not detected in the 65 commercial samples tested. The method evaluated herein represents a promising alternative for the food industry in its efforts to control L. monocytogenes. Overall analysis time is shorter and the method permits a straightforward analysis of a large number of samples with reliable results.