CRISPR Typing and Subtyping for Improved Laboratory Surveillance of Salmonella Infections

Laboratory surveillance systems for salmonellosis should ideally be based on the rapid serotyping and subtyping of isolates. However, current typing methods are limited in both speed and precision. Using 783 strains and isolates belonging to 130 serotypes, we show here that a new family of DNA repeats named CRISPR (clustered regularly interspaced short palindromic repeats) is highly polymorphic in Salmonella. We found that CRISPR polymorphism was strongly correlated with both serotype and multilocus sequence type. Furthermore, spacer microevolution discriminated between subtypes within prevalent serotypes, making it possible to carry out typing and subtyping in a single step. We developed a high-throughput subtyping assay for the most prevalent serotype, Typhimurium. An open web-accessible database was set up, providing a serotype/spacer dictionary and an international tool for strain tracking based on this innovative, powerful typing and subtyping tool

Détection d'activités humaines par l'utilisation du CSI

RÉSUMÉ: La technologie Wireless Fidelity (Wi-Fi) a été développée à la fin des années 1990 et peut être considérée comme arrivée pleinement à maturité. L’une des preuves de cette maturité est l’existence d’usages qui se détournent de l’objectif premier de la technologie, qui est de fournir un accès internet sans fil. Le Wi-Fi est désormais utilisé dans des applications de localisation, de domotique et internet des objets ou de communication vocale. Ces applications utilisent certaines caractéristiques techniques du Wi-Fi de manière créative. Par exemple, la puissance du signal reçu (RSSI) est un indicateur qui a pour but de déterminer la qualité du signal reçu, permettant de moduler dynamiquement la vitesse de transmission ou d’initier un processus de déconnexion quand le signal est trop faible. Certaines applications de localisation utilisent le RSSI pour déterminer la position d’un utilisateur, une manière détournée de mettre à profit des spécifications préexistantes. L’atout principal d’applications s’appuyant sur le Wi-Fi est le potentiel de les commercialiser à très grande échelle. En effet, le Wi-Fi domine largement les réseaux domestiques et publics. De même, il est prévalant dans les réseaux professionnels. Ainsi, le déploiement de nouvelles fonctionnalités peut s’adresser à un public particulièrement large. On peut citer le protocole VoIP qui permet de transmettre de la voix sur les réseaux IP, développé au cours des années 1980 et 1990 et dont l’utilisation était relativement confidentielle jusqu’à la démocratisation du Wi-Fi. Il est désormais utilisé par des applications populaires de messagerie et d’appels, telles que WhatsApp, Skype, FaceTime, Viber, Zoom, Microsoft Teams, dont le nombre d’utilisateurs est estimé à des centaines de millions. Nous allons appuyer notre étude sur la connaissance de l’état du canal (CSI). Cet indicateur s’appuie sur les nombreuses sous-porteuses du Wi-Fi. Il nous informe sur la qualité du signal transmis sur chaque sous-porteuse entre plusieurs antennes. Le CSI est impacté par les différents phénomènes physiques qui s’appliquent à un signal qui se propage en espace libre, réflexion, réfraction et diffraction. La variation de la qualité de la transmission des signaux du Wi-Fi a été déterminée empiriquement suffisamment fine pour pouvoir différencier plusieurs activités. Les modèles théoriques sont actuellement trop difficiles à mettre en place, ces phénomènes physiques dépendant de nombreux paramètres, tandis que lors d’un mouvement réalisé par un sujet, l’environnement est modifié à chaque seconde. ABSTRACT: The Wireless Fidelity (Wi-Fi) technology was developed in the late 1990s and should be considered to have reached full maturity by now. One of the proof of this is the fact that new applications are developed and stray from Wi-Fi’s original objective, which is providing a wireless internet access. Wi-Fi may now be used for localization, home automation, Internet of Things (IoT) or voice communications. These applications use creatively technical features of Wi-Fi. For instance, the Received Signal Strength Index (RSSI) is normally used in order to quantify the intensity of a signal, thus allowing to dynamically modify the velocity of transmission or start a reconnection process if the signal is too weak. Some applications now leverage the RSSI for localization, a new way to use a preexisting technology. The main advantage of Wi-Fi-based application is scalability. We can’t ignore the fact that Wi-Fi largely dominates other internet access technology, in both domestic and public networks, and is prevalent as well in professional networks. Therefore, any new application could theoretically reach a particularly wide audience. One notable example is the VoIP protocol, developed in the late 1980s and early 1990s, this protocol allowed voice transmission over IP. While it was relatively confidential before the democratization of Wi-Fi, it is now used by applications like WhatsApp, Skype, FaceTime, Zoom and Microsoft Teams, which have hundreds of millions of users. In our study, we aim to leverage Channel State Information (CSI). This index is associated to each subcarrier of Wi-Fi in each transmitted packet. It informs us on the quality of the transmitted signal, which like every signal propagating in free space is subject to physical phenomenons such as reflection, refraction and diffraction. It was experimentally proven that the variation of CSI was a stable enough indicator to pursue Human Activity Recognition (HAR). Theoretical models are yet to be developed because the complexity of considering every physical interaction for multiple frequency in fast moving and non-reproducible settings should not be underestimated. There is a vast diversity of CSI-based applications, ranging from crowd counting to localization or HAR. Some have fairly impressive results, reaching a 90% accuracy on recognizing activities among 7 different possibilities or reaching a 95% on hand gesture recognition. There is however a shortage of real life applications, most do not leave the confined environment of a laboratory. This is explained by the high level of control one has in such a setting, which is an enviable feature as you carry out an experiment

Misuse and Dependence on Non-Prescription Codeine Analgesics or Sedative H1 Antihistamines by Adults: A Cross-Sectional Investigation in France

<div><p>Background</p><p>Given the growing worldwide market of non-prescription drugs, monitoring their misuse in the context of self-medication represents a particular challenge in Public Health. The aim of this study was to investigate the prevalence of misuse, abuse, and dependence on non-prescription psychoactive drugs.</p><p>Method</p><p>During one month, in randomly solicited community pharmacies, an anonymous questionnaire was offered to adults requesting paracetamol (control group), codeine combined with paracetamol in analgesics, or sedative H1 antihistamines. Responses about misuse (drug use not in agreement with the Patient Information Leaflet) abuse (excessive drug use having detrimental consequences), and dependence (established according to questions adapted from the Diagnostic and Statistical Manual of Mental Disorders, 4th Edition criteria) on psychoactive drugs were compared to those of the paracetamol control group.</p><p>Results</p><p>295 patients (mean age 48.5 years, 68.5% of women) having used one of the studied drugs during the previous month were included. Misuse and dependence to codeine analgesics concerned 6.8% and 17.8% of the patients exposed to these drugs, respectively, (n = 118), which was significantly higher than for paracetamol. 19.5% had used codeine analgesics daily for more than six months. Headache was the most frequent reason for persistent daily use. A high prevalence of persistent daily users of sedative H1 antihistamines was also observed. Whereas these drugs are recommended only for short treatment courses of occasional insomnia, 72.2% of the participants having taken doxylamine (n = 36) were daily users, predominantly for more than six months.</p><p>Conclusions</p><p>Results on misuse and dependence on non-prescription codeine analgesics suggest that chronic pain, in particular chronic cephalalgia, requires better medical care. In addition, as for hypnotics on prescription, persistent use of doxylamine for self-medication is not justified until an acceptable benefit-risk ratio for chronic sleep disturbance is shown by clinical data.</p></div

Characteristics of the patients responding to the questionnaire and of users of the studied substances in the previous month.

*<p>p<0.05;</p>**<p>p<0.01;</p>***<p>p<0.001 (comparison to the control group).</p

Problematic uses of codeine (combined with paracetamol), H1 antihistamines, and paracetamol (control) among the patients having used these drugs during the previous month.

**<p>p<0.01;</p>***<p>p<0.001 (comparison to the control group).</p

Positive responses to DSM-IV substance dependence items.

<p>Results are expressed as percentage of the patients having used non-prescription analgesic drugs containing codeine during the previous month.</p

Cases of misuse of doxylamine (daily use for more than 5 consecutive days) purchased without prescription at the community pharmacy.

<p>Cases of misuse of doxylamine (daily use for more than 5 consecutive days) purchased without prescription at the community pharmacy.</p

Optimized Multilocus Variable-Number Tandem-Repeat Analysis Assay and Its Complementarity with Pulsed-Field Gel Electrophoresis and Multilocus Sequence Typing for Listeria monocytogenes Clone Identification and Surveillance

International audiencePopulations of the food-borne pathogen Listeria monocytogenes are genetically structured into a small number of major clonal groups, some of which have been implicated in multiple outbreaks. The goal of this study was to develop and evaluate an optimized multilocus variable number of tandem repeat (VNTR) analysis (MLVA) subtyping scheme for strain discrimination and clonal group identification. We evaluated 18 VNTR loci and combined the 11 best ones into two multiplexed PCR assays (MLVA-11). A collection of 255 isolates representing the diversity of clonal groups within phylogenetic lineages I and II, including representatives of epidemic clones, were analyzed by MLVA-11, multilocus sequence typing (MLST), and pulsed-field gel electropho-resis (PFGE). MLVA-11 had less discriminatory power than PFGE, except for some clones, and was unable to distinguish some epidemiologically unrelated isolates. Yet it distinguished all major MLST clones and therefore constitutes a rapid method to identify epidemiologically relevant clonal groups. Given its high reproducibility and high throughput, MLVA represents a very attractive first-line screening method to alleviate the PFGE workload in outbreak investigations and listeriosis surveillance. L isteriosis is a food-borne infection caused by the bacterium Listeria monocytogenes. Invasive forms of human listeriosis include septicemia, meningitis, and maternal-fetal infections (1). Listeriosis is associated with high hospitalization and fatality rates (almost 100% and 25 to 30%, respectively). Populations at risk include pregnant women, immunocompromised individuals, and the elderly. L. monocytogenes is widely present in the environment, including soil, water, vegetation, and silage, as well as in animals and animal-derived food, and can contaminate food in processing plants and retail establishments. L. monocytogenes is recognized as a public health issue and a serious challenge for the food industry, and this has led to the establishment of national surveillance systems in several countries. L. monocytogenes also stands out as a model system in the fields of microbiology, cell biology, and im-munology and for the study of host-pathogen interactions (2-5). L. monocytogenes strain characterization on the basis of sero-typing and molecular typing methods is used for surveillance, epidemiological tracking, and outbreak investigation purposes (6, 7). Genetic variants of L. monocytogenes have diversified into four major phylogenetic lineages, with lineages 1 and 2 each containing multiple clonal groups of public health importance (8-14). As these groups appear to differ in virulence and epidemic potential (6, 15), it will be interesting to better define their epidemiological, clinical, and microbiological specificities. For this purpose, tools for the easy identification of clonal groups are needed to recognize such groups and determine their presence in a large variety of sources. Several typing methods are currently available for L. monocytogenes strains. Conventional serotyping (16) and its molecular proxy PCR serogrouping (17) discriminate major categories of strains that correlate strongly (albeit not totally) with lin-eages and clones (11, 12, 14), but these methods do not have the necessary discriminatory power in the context of outbreak investigations. Pulsed-field gel electrophoresis (PFGE) is established as the gold standard for L. monocytogenes strain subtyping and is widely used for listeriosis surveillance and outbreak investigations (18). Yet, PFGE presents several practical disadvantages, as it is time-consuming and requires stringent standardization for inter-laboratory data comparison. Multilocus sequence typing (MLST) is a well-established reference method for global epidemiology and population biology (19, 20), as it renders interlaboratory genotype comparisons easy and unambiguous and as sequence data can be used to infer useful population genetic information such as amounts of genetic diversity, recombination rates, and strain phy-logeny. MLST also provides backward compatibility with genome sequencing (21). However, MLST is neither rapid nor cheap and has limited discriminatory power within L. monocytogenes (12, 22). Given the current limitations of available methods for L. monocytogenes strain typing, a potentially useful complementary approach is multilocus variable number of tandem repeat