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Urinary tract infections (UTIs) are a worldwide health problem. Fast and accurate detection of bacterial infection is essential to provide appropriate antibiotherapy to patients and to avoid the emergence of drug-resistant pathogens. While the gold standard requires 24h to 48h of bacteria culture prior MALDI-TOF species identification, we propose a culture-free workflow, enabling a bacterial identification and quantification in less than 4 hours using 1mL of urine. After a rapid and automatable sample preparation, a signature of 82 bacterial peptides, defined by machine learning, was monitored in LC-MS, to distinguish the 15 species causing 84% of the UTIs. The combination of the sensitivity of the SRM mode on a triple quadrupole TSQ Altis instrument and the robustness of capillary flow enabled us to analyze up to 75 samples per day, with 99.2% accuracy on bacterial inoculations of healthy urines. We have also shown our method can be used to quantify the spread of the infection, from 8x10 4 to 3x10 7 CFU/mL. Finally, the workflow was validated on 45 inoculated urines and on 84 UTI-positive urine from patients, with respectively 93.3% and 87.1% of agreement with the culture-MALDI procedure at a level above 1x10 5 CFU/mL corresponding to an infection requiring antibiotherapy., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Infectious diseases (IDs) are a leading cause of death. The diversity and adaptability of microbes represent a continuing risk to health. Combining vision with passion, our transdisciplinary medical research team has been focussing its work on the better management of infectious diseases for saving human lives over the past five decades through medical discoveries and innovations that helped change the practice of medicine. The team used a multiple-faceted and integrated approach to control infectious diseases through fundamental discoveries and by developing innovative prevention tools and rapid molecular diagnostic tests to fulfill the various unmet needs of patients and health professionals in the field of ID. In this article, as objectives, we put in context two main research areas of ID management: innovative infection prevention that is woman-controlled, and the rapid molecular diagnosis of infection and resistance. We also explain how our transdisciplinary approach encompassing specialists from diverse fields ranging from biology to engineering was instrumental in achieving success. Furthermore, we discuss our vision of the future for translational research to better tackle IDs.

Objective: To identify tools that predict the risk of complications for patients presenting to an outpatient clinic or an emergency department (ED) with influenza-like illness., Methods: We searched Medline, Embase, Cochrane Library and CINAHL from inception to July 2023. We included articles reporting on the derivation or validation of a score or algorithm used to stratify the risk of hospitalization or mortality among patients with influenza-like illness in the ED or outpatient clinic., Results: Twelve articles reporting on eight scores and six predictive models were identified. For predicting the need for hospitalization, the area under the curve (AUC) of the PMEWS and the CURB-65 ranged respectively from 0.76 to 0.94, and 0.65 to 0.88. The Community Assessment Tool had an AUC of 0.62. For predicting inpatient mortality, AUC was 0.66 for PMEWS and 0.79 for CURB-65, 0.79 for the SIRS criteria and 0.86 for the qSOFA score. Two scores were developed without external validation during the Covid-19 pandemic. The CovHos score and the Canadian Covid discharge score had an AUC ranged from 0.70 to 0.91. The predictive models performed adequately (AUC from 0.76 to 0.92) but will require external validation for clinical use. Tool diversity and study population heterogeneity precluded meta-analysis., Conclusion: Although the CURB, PMEWS and qSOFA scores appear to predict accurately the risk of complications of influenza-like illness, none were reliable enough to justify their widespread ED use. Refinement of an existing tool or development of a new tool to optimize the management of these patients is needed., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:Simon Berthelot reports article publishing charges was provided by Quebec Ministry of the Economy and Innovation. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors. Published by Elsevier Ltd.)

Objectives: To characterize vancomycin-resistance vanD gene clusters and potential vanD -carrying bacteria in the intestinal microbiota of healthy volunteers exposed or not to β-lactam antibiotics., Methods: Stool samples were collected before and after 7 days of cefprozil β-lactam antibiotic exposure of 18 participants and six control participants who were not exposed to the antibiotic at the same time points. Metagenomic sequencing and culture-enriched metagenomic sequencing (with and without β-lactam selection) were used to characterize vanD gene clusters and determine potential vanD -carrying bacteria. Alteration by antimicrobials was also examined., Results: Culture enrichment allowed detection of vanD genes in a large number of participants (11/24; 46%) compared to direct metagenomics (2/24; 8%). vanD genes were detected in stool cultures only following β-lactam exposure, either after β-lactam treatment of participants or after culture of stools with β-lactam selection. Six types of vanD gene clusters were identified. Two types of vanD cluster highly similar to those of enterococci were found in two participants. Other vanD genes or vanD clusters were nearly identical to those identified in commensal anaerobic bacteria of the families Lachnospiraceae and Oscillospiraceae and/or bordered by genomic sequences similar or related to these anaerobes, suggesting that they are the origin or carriers of vanD ., Conclusions: This study showed that culture-enriched metagenomics allowed detection of vanD genes not detected by direct metagenomics and revealed collateral enrichment of bacteria containing vancomycin-resistance vanD genes following exposure to β-lactams, with a higher prevalence of the most likely gut commensal anaerobes carrying vanD . These commensal anaerobes could be the reservoir of vanD genes carried by enterococci., (© The Author(s) 2023. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy.)

Background: The ongoing coronavirus diseases 2019 (COVID-19) pandemic with its numerous variants of concern has shown the need to have a robust and complete global infectious diseases genomic surveillance network worldwide. Various clinical and research institutions have stepped up to perform SARS-CoV-2 sequencing thus enhancing the understanding of this virus' global evolution. However, given that genomic sequencing capacities and capabilities are not available in every region or country, significant gaps exist, which lead to geographic blind spots. One such region is the Caribbean. This paper measures the Caribbean region's SARS-CoV-2 genomic sequencing capacity and highlights the need to improve further regional genomics surveillance capacities and capabilities, which are essential for efficient health interventions for infectious diseases., Methods: A map showing SARS-CoV-2 sequences available for each Caribbean Island was constructed using SARS-CoV-2 genomic, epidemiological and populational data obtained from GISAID, the World Health Organization, the United Nations, and the World Bank. The number of reported SARS-CoV-2 cases and the proportion of cases sequenced in each Caribbean Island was then analysed by the Gross Domestic Product per capita and political status., Findings: As of August 6, 2022, the number of SARS-CoV-2 sequences from the Caribbean are underrepresented with only 40,190 (1.07%) of the over 3.76 million documented cases sequenced, which is further exacerbated by a disparity based not only on the country's income but also on its political status (sovereign country versus dependent or integrated) and accessibility to sequencing technologies. There are a limited number of sequencing centres based in the Caribbean islands with the majority located on the American and European continents. Using mobile sequencing technologies while concomitantly investing in data analysis training could lead to greater and more sustainable coverage., Interpretation: Considering the Caribbean region's dispersed heterogeneous populations, varying political regimes, and resource-constrained healthcare systems, further development of local next-generation sequencing capacity and capabilities in the Caribbean region is needed to achieve global public health goals., Funding: No funding source was required for this study., Competing Interests: The authors have no conflicts of interest to disclose., (© 2022 The Authors.)

Objective: To identify tools that predict the risk of complications in patients presenting to outpatient clinics or emergency departments (ED) with acute infectious diarrhea., Methods: Medline, Embase, Cochrane Library, Web of Science and CINAHL were searched from inception to July 2021. Articles reporting on the derivation or validation of a score to stratify the risk of intravenous rehydration or hospitalization among patients with acute infectious diarrhea in the ED or outpatient clinic were retained for analysis., Results: Five articles reporting on two different tools were identified. Developed to assess the risk of hospitalization of children, the EsVida scale has not been externally validated. Developed originally to assess the level of dehydration in children, the Clinical Dehydration Scale (CDS) was evaluated as a risk stratification tool. For predicting intravenous rehydration, a CDS score ≥ 1 showed a sensitivity between 0.73 and 0.88 and specificity between 0.38 and 0.69, whereas a CDS score ≥ 5 showed a sensitivity between 0.06 and 0.32 and specificity between 0.94 and 0.99. For predicting hospitalization, a CDS score ≥ 1 showed a sensitivity between 0.74 and 1.00 and specificity between 0.34 and 0.38, whereas a CDS score ≥ 5 showed a sensitivity between 0.26 and 0.62 and specificity between 0.66 and 0.96. High heterogeneity among studies and unclear risk of bias precluded meta-analysis., Conclusion: As a risk-stratification tool, the CDS has been validated only for children. Further research is needed to develop and validate a tool suitable for adults in the ED., Competing Interests: Declaration of Competing Interest None., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Comparative metagenomics studies have highlighted differences in microbiome community structure among human populations over diverse lifestyles and environments. With their unique environmental and historical backgrounds, Nunavik Inuit have a distinctive gut microbiome with undocumented health-related implications. Using shotgun metagenomics, we explored the taxonomic and functional structure of the gut microbiome from 275 Nunavik Inuit ranging from 16 to 30-year-old. Whole-metagenome analyses revealed that Nunavik Inuit youths have a more diverse microbiome than their non-industrialized and industrialized counterparts. A comparison of k-mer content illustrated the uniqueness of the Nunavik gut microbiome. Short-chain fatty acids producing species, and carbohydrates degradation pathways dominated Inuit metagenomes. We identified a taxonomic and functional signature unique to the Nunavik gut microbiome contrasting with other populations using a random forest classifier. Here, we show that the Nunavik Inuit gut microbiome exhibits high diversity and a distinct community structure., (© 2022. The Author(s).)

Antimicrobial resistance (AMR) is continuing to grow across the world. Though often thought of as a mostly public health issue, AMR is also a major agricultural and environmental problem. As such, many researchers refer to it as the preeminent One Health issue. Aerial transport of antimicrobial-resistant bacteria via bioaerosols is still poorly understood. Recent work has highlighted the presence of antibiotic resistance genes in bioaerosols. Emissions of AMR bacteria and genes have been detected from various sources, including wastewater treatment plants, hospitals, and agricultural practices; however, their impacts on the broader environment are poorly understood. Contextualizing the roles of bioaerosols in the dissemination of AMR necessitates a multidisciplinary approach. Environmental factors, industrial and medical practices, as well as ecological principles influence the aerial dissemination of resistant bacteria. This article introduces an ongoing project assessing the presence and fate of AMR in bioaerosols across Canada. Its various sub-studies include the assessment of the emissions of antibiotic resistance genes from many agricultural practices, their long-distance transport, new integrative methods of assessment, and the creation of dissemination models over short and long distances. Results from sub-studies are beginning to be published. Consequently, this paper explains the background behind the development of the various sub-studies and highlight their shared aspects.

Anti-1-amino-3- 18 fluorine-fluorocyclobutane-1-carboxylic acid ( 18 F-fluciclovine) positron emission tomography (PET) shows preferential glioma uptake but there is little data on how uptake correlates with post-contrast T1-weighted (Gd-T1) and dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) activity during adjuvant treatment. This pilot study aimed to compare 18 F-fluciclovine PET, DCE-MRI and Gd-T1 in patients undergoing chemoradiotherapy for glioblastoma (GBM), and in a parallel pre-clinical GBM model, to investigate correlation between 18 F-fluciclovine uptake, MRI findings, and tumour biology. 18 F-fluciclovine-PET-computed tomography (PET-CT) and MRI including DCE-MRI were acquired before, during and after adjuvant chemoradiotherapy (60 Gy in 30 fractions with temozolomide) in GBM patients. MRI volumes were manually contoured; PET volumes were defined using semi-automatic thresholding. The similarity of the PET and DCE-MRI volumes outside the Gd-T1 volume boundary was measured using the Dice similarity coefficient (DSC). CT-2A tumour-bearing mice underwent MRI and 18 F-fluciclovine PET-CT. Post-mortem mice brains underwent immunohistochemistry staining for ASCT2 (amino acid transporter), nestin (stemness) and Ki-67 (proliferation) to assess for biologically active tumour. 6 patients were recruited (GBM 1-6) and grouped according to overall survival (OS)-short survival (GBM-SS, median OS 249 days) and long survival (GBM-LS, median 903 days). For GBM-SS, PET tumour volumes were greater than DCE-MRI, in turn greater than Gd-T1. For GBM-LS, Gd-T1 and DCE-MRI were greater than PET. Tumour-specific 18 F-fluciclovine uptake on pre-clinical PET-CT corresponded to immunostaining for Ki-67, nestin and ASCT2. Results suggest volumes of 18 F-fluciclovine-PET activity beyond that depicted by DCE-MRI and Gd-T1 are associated with poorer prognosis in patients undergoing chemoradiotherapy for GBM. The pre-clinical model confirmed 18 F-fluciclovine uptake reflected biologically active tumour.

Background: Previous data on glycogen synthase kinase 3 (GSK-3) inhibition in cancer models support a cytotoxic effect with selectivity for tumor cells compared to normal tissue but the effect of these inhibitors in glioma has not been widely studied. Here, we investigate their potential as cytotoxics in glioma., Methods: We assessed the effect of pharmacologic GSK-3 inhibition on established (U87, U251) and patient-derived (GBM1, GBM4) glioblastoma (GBM) cell lines using cytotoxicity assays as well as undertaking a detailed investigation of the effect on cell cycle, mitosis, and centrosome biology. We also assessed drug uptake and efficacy of GSK-3 inhibition alone and in combination with radiation in xenograft models., Results: Using the selective GSK-3 inhibitor AZD2858, we demonstrated single agent cytotoxicity in two patient-derived glioma cell lines (GBM1, GBM4) and two established cell lines (U251 and U87) with IC 50 in the low micromolar range promoting centrosome disruption, failed mitosis, and S-phase arrest. Glioma xenografts exposed to AZD2858 also showed growth delay compared to untreated controls. Combined treatment with radiation increased the cytotoxic effect of clinical radiation doses in vitro and in orthotopic glioma xenografts., Conclusions: These data suggest that GSK-3 inhibition promotes cell death in glioma through disrupting centrosome function and promoting mitotic failure and that AZD2858 is an effective adjuvant to radiation at clinical doses.

Carbapenemase-producing Enterobacterales, including KPC-2 producers, have become a major clinical problem. During an outbreak in Quebec City, Canada, KPC-2-producing Klebsiella michiganensis and Citrobacter farmeri were isolated from a patient six weeks apart. We determined their complete genome sequences. Both isolates carried nearly identical IncN2 plasmids with bla KPC-2 on a Tn 4401b element. Both strains also carried IncP1 plasmids, but that of C. farmeri did not carry a Beta-lactamase gene, whereas that of K. michiganensis carried a second copy of bla KPC-2 on Tn 4401b . These results suggest recent plasmid transfer between the two species and a recent transposition event.

DNA hybridization phenomena occurring on solid supports are not understood as clearly as aqueous phase hybridizations and mathematical models cannot predict some empirically obtained results. Ongoing research has identified important parameters but remains incomplete to accurately account for all interactions. It has previously been shown that the length of the overhanging (dangling) end of the target DNA strand following hybridization to the capture probe is correlated to interactions with the complementary strand in solution which can result in unbinding of the target and its release from the surface. We have developed an instrument for real-time monitoring of DNA hybridization on spherical particles functionalized with oligonucleotide capture probes and arranged in the form of a tightly packed monolayer bead bed inside a microfluidic cartridge. The instrument is equipped with a pneumatic module to mediate displacement of fluid on the cartridge. We compared this system to both conventional (passive) and centrifugally-driven (active) microfluidic microarray hybridization on glass slides to establish performance levels for the detection of single nucleotide polymorphisms. The system was also used to study the effect of the dangling end's length in real-time when the immobilized target DNA is exposed to the complementary strand in solution. Our findings indicate that increasing the length of the dangling end leads to desorption of target amplicons from bead-bound capture probes at a rate approaching that of the initial hybridization process. Finally, bead bed hybridization was performed with Streptococcus agalactiae cfb gene amplicons obtained from randomized clinical samples, which allowed for identification of group B streptococci within 5-15 min. The methodology presented here is useful for investigating competitive hybridization mechanisms on solid supports and to rapidly validate the suitability of microarray capture probes.

MicroRNAs play an important role in the regulation of mRNA translation and have therapeutic potential in cancer and other diseases. To profile the landscape of microRNAs with significant cytotoxicity in the context of glioblastoma (GBM), we performed a high-throughput screen in adult and pediatric GBM cells using a synthetic oligonucleotide library representing all known human microRNAs. Bioinformatics analysis was used to refine this list and the top seven microRNAs were validated in a larger panel of GBM cells using state-of-the-art in vitro assays. The cytotoxic effect of our most relevant candidate was assessed in a preclinical model. Our screen identified ~100 significantly cytotoxic microRNAs with 70% concordance between cell lines. MicroRNA-1300 (miR-1300) was the most potent and robust candidate. We observed a striking binucleated phenotype in miR-1300 transfected cells due to cytokinesis failure followed by apoptosis. This was also observed in two stem-like patient-derived cultures. We identified the physiological role of miR-1300 as a regulator of endomitosis in megakaryocyte differentiation where blockade of cytokinesis is an essential step. In GBM cells, where miR-1300 is normally not expressed, the oncogene Epithelial Cell Transforming 2 (ECT2) was validated as a direct key target. ECT2 siRNA phenocopied the effects of miR-1300, and ECT2 overexpression led to rescue of miR-1300 induced binucleation. We showed that ectopic expression of miR-1300 led to decreased tumor growth in an orthotopic GBM model. Our screen provides a resource for the neuro-oncology community and identified miR-1300 as a novel regulator of endomitosis with translatable potential for therapeutic application.

Rapid and accurate diagnosis of bovine respiratory disease (BRD) presents a substantial challenge to the North American cattle industry. Here we utilize recombinase polymerase amplification (RPA), a fast and sensitive isothermal DNA-based technology for the detection of four BRD pathogens ( Mannheimia haemolytica, Pasteurella multocida, Histophilus somni, Mycoplasma bovis ), genes coding antimicrobial resistance (AMR) and integrative conjugative elements (ICE) which can harbor AMR genes. Eleven RPA assays were designed and validated including: a) one conventional species-specific multiplex assay targeting the 4 BRD pathogens, b) two species-specific real-time multiplex RPA assays targeting M. haemolytica / M. bovis and P. multocida / H. somni , respectively with a novel competitive internal amplification control, c) seven conventional assays targeting AMR genes ( tetH, tetR, msrE, mphE, sul2, floR, erm42 ), and d) one real-time assay targeting ICE. Each real-time RPA assay was tested on 100 deep nasopharyngeal swabs (DNPS) collected from feedlot cattle previously assessed for targets using either culture methods and/or polymerase chain reaction (PCR) verification (TC-PCR). The developed RPA assays enabled sensitive and accurate identification of BRD agents and AMR/ICE genes directly from DNPS, in a shorter period than TC-PCR, showing considerable promise as a tool for point-of-care identification of BRD pathogens and antimicrobial resistance genes., (Copyright © 2020 Conrad, Daher, Stanford, Amoako, Boissinot, Bergeron, Alexander, Cook, Ralston, Zaheer, Niu and McAllister.)

Fast identification of microbial species in clinical samples is essential to provide an appropriate antibiotherapy to the patient and reduce the prescription of broad-spectrum antimicrobials leading to antibioresistances. MALDI-TOF-MS technology has become a tool of choice for microbial identification but has several drawbacks: it requires a long step of bacterial culture before analysis (≥24 h), has a low specificity and is not quantitative. We developed a new strategy for identifying bacterial species in urine using specific LC-MS/MS peptidic signatures. In the first training step, libraries of peptides are obtained on pure bacterial colonies in DDA mode, their detection in urine is then verified in DIA mode, followed by the use of machine learning classifiers (NaiveBayes, BayesNet and Hoeffding tree) to define a peptidic signature to distinguish each bacterial species from the others. Then, in the second step, this signature is monitored in unknown urine samples using targeted proteomics. This method, allowing bacterial identification in less than 4 h, has been applied to fifteen species representing 84% of all Urinary Tract Infections. More than 31,000 peptides in 190 samples were quantified by DIA and classified by machine learning to determine an 82 peptides signature and build a prediction model. This signature was validated for its use in routine using Parallel Reaction Monitoring on two different instruments. Linearity and reproducibility of the method were demonstrated as well as its accuracy on donor specimens. Within 4h and without bacterial culture, our method was able to predict the predominant bacteria infecting a sample in 97% of cases and 100% above the standard threshold. This work demonstrates the efficiency of our method for the rapid and specific identification of the bacterial species causing UTI and could be extended in the future to other biological specimens and to bacteria having specific virulence or resistance factors., (© 2019 Roux-Dalvai et al.)

Science is the engine of prosperity, helping society respond to the important issues it faces. We are presently witnessing a worldwide decline in high school students' interest in science, regardless of gender. To overcome this problem, developing science promotion programs and activities that inspire young people to become the scientists of tomorrow is critical. We strongly believe in the leadership of research centers in creating such programs, which could have a significant impact on the next generation of scientists. Here we present the 'Researcher for a Day' program, which offers senior high school students immersion days in a scientific workplace dedicated to microbiology, as an example for other institutions that would like to implement such a program. Researcher for a Day has already helped more than 4,000 young students who are considering their career choices discover the world of science. Similar approaches could be implemented virtually anywhere to extend these efforts to promote science among young people., (©2019 Author(s). Published by the American Society for Microbiology.)

A rod-shaped, motile anaerobic bacterium, designated CCRI-22567 T , was isolated from a vaginal sample of a woman diagnosed with bacterial vaginosis and subjected to a polyphasic taxonomic study. The novel strain was capable of growth at 30-42 °C (optimum, 42 °C), at pH 5.5-8.5 (optimum, pH 7.0-7.5) and in the presence of 0-1.5 % (w/v) NaCl (optimally at 0.5 % NaCl). The phylogenetic trees based on 16S rRNA gene sequences showed that strain CCRI-22567 T forms a distinct evolutionary lineage independent of other taxa in the family Peptostreptococcaceae . Strain CCRI-22567 T exhibited 90.1 % 16S rRNA gene sequence similarity to Peptoanaerobacter stomatis ACC19a T and 89.7 % to Eubacterium yurii subsp. schtitka ATCC 43716. The three closest organisms with an available whole genome were compared to strain CCRI-22567 T for genomic relatedness assessment. The genomic average nucleotide identities (OrthoANIu) obtained with Peptoanaerobacter stomatis ACC19a T , Eubacterium yurii subsp. margaretiae ATCC 43715 and Filifactor alocis ATCC 35896 T were 71.8, 70.3 and 69.6 %, respectively. Strain CCRI-22567 T contained C 18 : 1  ω9 c and C 18 : 1  ω9 c DMA as the major fatty acids. The DNA G+C content of strain CCRI-22567 T based on its genome sequence was 33.8 mol%. On the basis of the phylogenetic, chemotaxonomic and other phenotypic properties, strain CCRI-22567 T is considered to represent a new genus and species within the family Peptostreptococcaceae , for which the name Criibacterium bergeronii gen. nov., sp. nov., is proposed. The type strain of Criibacterium bergeronii is CCRI-22567 T (=LMG 31278 T =DSM 107614 T =CCUG 72594 T ).

Background: Low-abundance microorganisms of the gut microbiome are often referred to as a reservoir for antibiotic resistance genes. Unfortunately, these less-abundant bacteria can be overlooked by deep shotgun sequencing. In addition, it is a challenge to associate the presence of resistance genes with their risk of acquisition by pathogens. In this study, we used liquid culture enrichment of stools to assemble the genome of lower-abundance bacteria from fecal samples. We then investigated the gene content recovered from these culture-enriched and culture-independent metagenomes in relation with their taxonomic origin, specifically antibiotic resistance genes. We finally used a pangenome approach to associate resistance genes with the core or accessory genome of Enterobacteriaceae and inferred their propensity to horizontal gene transfer., Results: Using culture-enrichment approaches with stools allowed assembly of 187 bacterial species with an assembly size greater than 1 million nucleotides. Of these, 67 were found only in culture-enriched conditions, and 22 only in culture-independent microbiomes. These assembled metagenomes allowed the evaluation of the gene content of specific subcommunities of the gut microbiome. We observed that differentially distributed metabolic enzymes were associated with specific culture conditions and, for the most part, with specific taxa. Gene content differences between microbiomes, for example, antibiotic resistance, were for the most part not associated with metabolic enzymes, but with other functions. We used a pangenome approach to determine if the resistance genes found in Enterobacteriaceae, specifically E. cloacae or E. coli, were part of the core genome or of the accessory genome of this species. In our healthy volunteer cohort, we found that E. cloacae contigs harbored resistance genes that were part of the core genome of the species, while E. coli had a large accessory resistome proximal to mobile elements., Conclusion: Liquid culture of stools contributed to an improved functional and comparative genomics study of less-abundant gut bacteria, specifically those associated with antibiotic resistance. Defining whether a gene is part of the core genome of a species helped in interpreting the genomes recovered from culture-independent or culture-enriched microbiomes.

Initially, Escherichia albertii has been described as a non-lactose fermenting bacterium and methods used to isolate it were first based on this phenotypic property. However, a recent study showed a variable lactose fermentation phenotype for E. albertii suggesting that this microorganism could have been underestimated by previous studies using isolation methods based on lactose fermentation. In this study, we present a method for the isolation and identification of both lactose fermenting and non-fermenting-E. albertii cells in stool samples, said method combining culture and isolation on mEA agar, an indole test, as well as an E. albertii-specific PCR assay for formal species identification. The ability of the procedure to detect E. albertii strains was verified using 19 E. albertii strains and 132 non-E. albertii strains representing 88 species of different origins majoritary belonging to the Enterobacteriaceae family. All indole-positive white colonies grown on mEA agar were subjected to E. albertii-specific PCR amplification; all E. albertii strains tested were detected with this assay and none of the non-E. albertii strains tested was detected. To demonstrate the ability of the procedure to directly detect E. albertii in stool samples, E. albertii-inoculated stools were tested and for all inoculated samples, E. albertii colonies were easily detected and identified. The present study provides a method enable to recover both lactose-fermenting and -non-fermenting E. albertii strains from clinical samples. This method could help to provide a better portrait of the prevalence and pathogenicity of E. albertii in clinical samples., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Lachnotalea glycerini CCRI-19302 belongs to the genus Lachnotalea The strain was isolated from a water sample harvested in Québec City, Canada. The genome assembly comprised 4,694,231 bp, with 34.6% GC content. This is the first documentation to report the genome sequence of a sporulating and motile strain of L. glycerini ., (Copyright © 2017 Maheux et al.)

The Romboutsia maritimum sp. nov. CCRI-22766 T strain was isolated from coastal estuarine mud in New Zealand. The genome assembly comprised 2,854,352 bp, with 27.1% G+C content. This is the first documentation that reports the genome sequence of R. maritimum ., (Copyright © 2017 Maheux et al.)

Romboutsia weinsteinii sp. nov. CCRI-19649 T belongs to the genus Romboutsia The strain was isolated from a water sample harvested in Québec City, Québec, Canada. The genome assembly comprised 4,134,593 bp with a 29.3% GC content. This is the first documentation that reports the genome sequence of R. weinsteinii ., (Copyright © 2017 Maheux et al.)

Successful treatment of a Candida infection relies on 1) an accurate identification of the pathogenic fungus and 2) on its susceptibility to antifungal drugs. In the present study we investigated the level of correlation between phylogenetical evolution and susceptibility of pathogenic Candida spp. to antifungal drugs. For this, we compared a phylogenetic tree, assembled with the concatenated sequences (2475-bp) of the ATP2, TEF1, and TUF1 genes from 20 representative Candida species, with published minimal inhibitory concentrations (MIC) of the four principal antifungal drug classes commonly used in the treatment of candidiasis: polyenes, triazoles, nucleoside analogues, and echinocandins. The phylogenetic tree revealed three distinct phylogenetic clusters among Candida species. Species within a given phylogenetic cluster have generally similar susceptibility profiles to antifungal drugs and species within Clusters II and III were less sensitive to antifungal drugs than Cluster I species. These results showed that phylogenetical relationship between clusters and susceptibility to several antifungal drugs could be used to guide therapy when only species identification is available prior to information pertaining to its resistance profile. An extended study comprising a large panel of clinical samples should be conducted to confirm the efficiency of this approach in the treatment of candidiasis., (Copyright © 2016. Published by Elsevier B.V.)

Antibiotics profoundly affect the gut microbiome and modulate microbial communities. We recently observed that antimicrobial drugs also impact the abundance and distribution of antibiotic resistance genes. In this addendum, we reanalyze our ∼1 trillion nucleotide shotgun metagenomic dataset to quantify comprehensive genomic differences at the sequence level before and after antibiotic treatment. We show that 7 day exposure to cefprozil leads to a statistically significant loss of metagenome sequences. Recovery of gut microbiomes 3 months after antibiotherapy was characterized by the emergence of new genome sequences not observed prior to antibiotic exposure. Participants with low initial gut microbiome diversity had an increased amount of sequences related to antibiotic resistance. Therefore, we suggest that while the taxonomical composition of microbiomes is partially affected by the antibiotic, the genomic content and population structure of bacterial communities is noticeably impacted.

Criibacterium bergeronii gen. nov., sp. nov., CCRI-22567 is the type strain of the new genus Criibacterium The strain was isolated from a woman with bacterial vaginosis. The genome assembly comprised 2,384,460 bp, with 34.4% G+C content. This is the first genome announcement of a strain belonging to the genus Criibacterium., (Copyright © 2016 Maheux et al.)

Background: First introduced in 2006, recombinase polymerase amplification (RPA) has stirred great interest, as evidenced by 75 publications as of October 2015, with 56 of them just in the last 2 years. The widespread adoption of this isothermal molecular tool in many diagnostic fields represents an affordable (approximately 4.3 USD per test), simple (few and easy hands-on steps), fast (results within 5-20 min), and sensitive (single target copy number detected) method for the identification of pathogens and the detection of single nucleotide polymorphisms in human cancers and genetically modified organisms., Content: This review summarizes the current knowledge on RPA. The molecular diagnostics of various RNA/DNA pathogens is discussed while highlighting recent applications in clinical settings with focus on point-of-care (POC) bioassays and on automated fluidic platforms. The strengths and limitations of this isothermal method are also addressed., Summary: RPA is becoming a molecular tool of choice for the rapid, specific, and cost-effective identification of pathogens. Owing to minimal sample-preparation requirements, low operation temperature (25-42 °C), and commercial availability of freeze-dried reagents, this method has been applied outside laboratory settings, in remote areas, and interestingly, onboard automated sample-to-answer microfluidic devices. RPA is undoubtedly a promising isothermal molecular technique for clinical microbiology laboratories and emergence response in clinical settings., (© 2016 American Association for Clinical Chemistry.)

Microbiome studies have demonstrated the high inter-individual diversity of the gut microbiota. However, how the initial composition of the microbiome affects the impact of antibiotics on microbial communities is relatively unexplored. To specifically address this question, we administered a second-generation cephalosporin, cefprozil, to healthy volunteers. Stool samples gathered before antibiotic exposure, at the end of the treatment and 3 months later were analysed using shotgun metagenomic sequencing. On average, 15 billion nucleotides were sequenced for each sample. We show that standard antibiotic treatment can alter the gut microbiome in a specific, reproducible and predictable manner. The most consistent effect of the antibiotic was the increase of Lachnoclostridium bolteae in 16 out of the 18 cefprozil-exposed participants. Strikingly, we identified a subgroup of participants who were enriched in the opportunistic pathogen Enterobacter cloacae after exposure to the antibiotic, an effect linked to lower initial microbiome diversity and to a Bacteroides enterotype. Although the resistance gene content of participants' microbiomes was altered by the antibiotic, the impact of cefprozil remained specific to individual participants. Resistance genes that were not detectable prior to treatment were observed after a 7-day course of antibiotic administration. Specifically, point mutations in beta-lactamase blaCfxA-6 were enriched after antibiotic treatment in several participants. This suggests that monitoring the initial composition of the microbiome before treatment could assist in the prevention of some of the adverse effects associated with antibiotics or other treatments.

Background: A point-of-care rapid test (POCRT) may help early and targeted use of antiviral drugs for the management of influenza A infection., Objective: (i) To determine whether antiviral treatment based on a POCRT for influenza A is cost-effective and, (ii) to determine the thresholds of key test parameters (sensitivity, specificity and cost) at which a POCRT based-strategy appears to be cost effective., Methods: An hybrid « susceptible, infected, recovered (SIR) » compartmental transmission and Markov decision analytic model was used to simulate the cost-effectiveness of antiviral treatment based on a POCRT for influenza A in the social perspective. Data input parameters used were retrieved from peer-review published studies and government databases. The outcome considered was the incremental cost per life-year saved for one seasonal influenza season., Results: In the base-case analysis, the antiviral treatment based on POCRT saves 2 lives/100,000 person-years and costs $7600 less than the empirical antiviral treatment based on clinical judgment alone, which demonstrates that the POCRT-based strategy is dominant. In one and two way-sensitivity analyses, results were sensitive to the POCRT accuracy and cost, to the vaccination coverage as well as to the prevalence of influenza A. In probabilistic sensitivity analyses, the POCRT strategy is cost-effective in 66% of cases, for a commonly accepted threshold of $50,000 per life-year saved., Conclusion: The influenza antiviral treatment based on POCRT could be cost-effective in specific conditions of performance, price and disease prevalence., (© 2015 The Authors. Influenza and Other Respiratory Viruses Published by John Wiley & Sons Ltd.)

Recombinase polymerase amplification (RPA) technology relies on three major proteins, recombinase proteins, single-strand binding proteins, and polymerases, to specifically amplify nucleic acid sequences in an isothermal format. The performance of RPA with respect to sequence mismatches of closely-related non-target molecules is not well documented and the influence of the number and distribution of mismatches in DNA sequences on RPA amplification reaction is not well understood. We investigated the specificity of RPA by testing closely-related species bearing naturally occurring mismatches for the tuf gene sequence of Pseudomonas aeruginosa and/or Mycobacterium tuberculosis and for the cfb gene sequence of Streptococcus agalactiae. In addition, the impact of the number and distribution of mismatches on RPA efficiency was assessed by synthetically generating 14 types of mismatched forward primers for detecting five bacterial species of high diagnostic relevance such as Clostridium difficile, Staphylococcus aureus, S. agalactiae, P. aeruginosa, and M. tuberculosis as well as Bacillus atropheus subsp. globigii for which we use the spores as internal control in diagnostic assays. A total of 87 mismatched primers were tested in this study. We observed that target specific RPA primers with mismatches (n > 1) at their 3'extrimity hampered RPA reaction. In addition, 3 mismatches covering both extremities and the center of the primer sequence negatively affected RPA yield. We demonstrated that the specificity of RPA was multifactorial. Therefore its application in clinical settings must be selected and validated a priori. We recommend that the selection of a target gene must consider the presence of closely-related non-target genes. It is advisable to choose target regions with a high number of mismatches (≥36%, relative to the size of amplicon) with respect to closely-related species and the best case scenario would be by choosing a unique target gene., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Background: Glioblastoma is the most aggressive primary brain tumor, and is associated with a very poor prognosis. In this study we investigated the potential of microRNA expression profiles to predict survival in this challenging disease., Methods: MicroRNA and mRNA expression data from glioblastoma (n = 475) and grade II and III glioma (n = 178) were accessed from The Cancer Genome Atlas. LASSO regression models were used to identify a prognostic microRNA signature. Functionally relevant targets of microRNAs were determined using microRNA target prediction, experimental validation and correlation of microRNA and mRNA expression data., Results: A 9-microRNA prognostic signature was identified which stratified patients into risk groups strongly associated with survival (p = 2.26e-09), significant in all glioblastoma subtypes except the non-G-CIMP proneural group. The statistical significance of the microRNA signature was higher than MGMT methylation in temozolomide treated tumors. The 9-microRNA risk score was validated in an independent dataset (p = 4.50e-02) and also stratified patients into high- and low-risk groups in lower grade glioma (p = 5.20e-03). The majority of the 9 microRNAs have been previously linked to glioblastoma biology or treatment response. Integration of the expression patterns of predicted microRNA targets revealed a number of relevant microRNA/target pairs, which were validated in cell lines., Conclusions: We have identified a novel, biologically relevant microRNA signature that stratifies high- and low-risk patients in glioblastoma. MicroRNA/mRNA interactions identified within the signature point to novel regulatory networks. This is the first study to formulate a survival risk score for glioblastoma which consists of microRNAs associated with glioblastoma biology and/or treatment response, indicating a functionally relevant signature., (Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

The combination of molecular diagnostic technologies is increasingly used to overcome limitations on sensitivity, specificity or multiplexing capabilities, and provide efficient lab-on-chip devices. Two such techniques, PCR amplification and microarray hybridization are used serially to take advantage of the high sensitivity and specificity of the former combined with high multiplexing capacities of the latter. These methods are usually performed in different buffers and reaction chambers. However, these elaborate methods have high complexity and cost related to reagent requirements, liquid storage and the number of reaction chambers to integrate into automated devices. Furthermore, microarray hybridizations have a sequence dependent efficiency not always predictable. In this work, we have developed the concept of a structured oligonucleotide probe which is activated by cleavage from polymerase exonuclease activity. This technology is called SCISSOHR for Structured Cleavage Induced Single-Stranded Oligonucleotide Hybridization Reaction. The SCISSOHR probes enable indexing the target sequence to a tag sequence. The SCISSOHR technology also allows the combination of nucleic acid amplification and microarray hybridization in a single vessel in presence of the PCR buffer only. The SCISSOHR technology uses an amplification probe that is irreversibly modified in presence of the target, releasing a single-stranded DNA tag for microarray hybridization. Each tag is composed of a 3-nucleotide sequence-dependent segment and a unique "target sequence-independent" 14-nucleotide segment allowing for optimal hybridization with minimal cross-hybridization. We evaluated the performance of five (5) PCR buffers to support microarray hybridization, compared to a conventional hybridization buffer. Finally, as a proof of concept, we developed a multiplexed assay for the amplification, detection, and identification of three (3) DNA targets. This new technology will facilitate the design of lab-on-chip microfluidic devices, while also reducing consumable costs. At term, it will allow the cost-effective automation of highly multiplexed assays for detection and identification of genetic targets.

Objectives: The therapeutic goal for patients with rheumatoid arthritis (RA) is clinical remission. This is best achieved by early diagnosis and appropriate therapeutic intervention. RA is associated with dysregulation of T-cell subsets (naïve, regulatory (Treg) and inflammation-related cells (IRC)) early in the disease. Our aim was to test the hypothesis that T-cell subset quantification can predict the achievement of clinical remission with early treatment in RA., Methods: T-cell subsets were quantified in 108 drug-naïve, early RA patients commencing methotrexate (MTX) or MTX+antitumor necrosis factor (anti-TNF) and in 105 healthy controls (HC). The primary outcome assessed was remission (DAS28<2.6). A pilot study used frozen cells (38 patients and 35 HCs, see online supplementary material) and was validated with fresh blood (70 patients and 70 HCs)., Results: Immune dysregulation in early RA was confirmed with an association between age and reduced naïve cells compared with HCs (p=0.006), a lower age-adjusted Treg and higher IRC frequency (p=0.001). Anticitrullinated peptide antibody (ACPA) positivity was associated with lower naïve (p=0.031) and Treg frequencies (p=0.039). In 50 patients treated with MTX, ACPA/age-adjusted analysis demonstrated that higher naïve cell frequency (relative to HC) was associated with remission (OR 5.90 (1.66 to 20.98), p=0.006, sensitivity/specificity 62%/79%, Positive Predictive Value (PPV)/Negative Predictive Value (NPV) 66%/76%). Remission with MTX+anti-TNF (n=20) was not found to be associated with naïve cell frequency, and for patients with reduced naïve cells the remission rate increased from 24% (MTX) to 42% (MTX+anti-TNF)., Conclusions: Baseline T-cell subset analysis has a value in predicting early RA remission with first therapy with MTX. Immunological analysis could be used in conjunction with clinical/serological features to predict response to MTX and help select the most appropriate therapy at disease presentation., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.)

Met is the receptor of hepatocyte growth factor (HGF), a cytoprotective cytokine. Disturbing the equilibrium between Met and its ligand may lead to inappropriate cell survival, accumulation of genetic abnormalities and eventually, malignancy. Abnormal activation of the HGF/Met axis is established in solid tumours and in chronic haematological malignancies, including myeloma, acute myeloid leukaemia, chronic myelogenous leukaemia (CML), and myeloproliferative neoplasms (MPNs). The molecular mechanisms potentially responsible for the abnormal activation of HGF/Met pathways are described and discussed. Importantly, inCML and in MPNs, the production of HGF is independent of Bcr-Abl and JAK2V617F, the main molecular markers of these diseases. In vitro studies showed that blocking HGF/Met function with neutralizing antibodies or Met inhibitors significantly impairs the growth of JAK2V617F-mutated cells. With personalised medicine and curative treatment in view, blocking activation of HGF/Met could be a useful addition in the treatment of CML and MPNs for those patients with high HGF/MET expression not controlled by current treatments (Bcr-Abl inhibitors in CML; phlebotomy, hydroxurea, JAK inhibitors in MPNs).

Background: Group B streptococcal infections are the leading cause of sepsis and meningitis in newborns. A rapid and reliable method for the detection of this pathogen at the time of delivery is needed for the early treatment of neonates. Isothermal amplification techniques such as recombinase polymerase amplification have advantages relative to PCR in terms of the speed of reaction and simplicity., Methods: We studied the clinical performance of recombinase polymerase amplification for the screening of group B streptococci in vaginal/anal samples from 50 pregnant women. We also compared the limit of detection and the analytical specificity of this isothermal assay to real-time PCR (RT-PCR)., Results: Compared to RT-PCR, the recombinase polymerase amplification assay showed a clinical sensitivity of 96% and a clinical specificity of 100%. The limit of detection was 98 genome copies and the analytical specificity was 100% for a panel of 15 bacterial and/or fungal strains naturally found in the vaginal/anal flora. Time-to-result for the recombinase polymerase amplification assay was <20 min compared to 45 min for the RT-PCR assay; a positive sample could be detected as early as 8 min., Conclusions: We demonstrate the potential of isothermal recombinase polymerase amplification assay as a clinically useful molecular diagnostic tool that is simple and faster than PCR/RT-PCR. Recombinase polymerase amplification offers great potential for nucleic acid-based diagnostics at the point of care.

Autoantibodies have been associated with human pathologies for a long time, particularly with autoimmune diseases (AIDs). Rheumatoid factor (RF) is known since the late 1930s to be associated with rheumatoid arthritis (RA). The discovery of anticitrullinated protein antibodies in the last century has changed this and other posttranslational modifications (PTM) relevant to RA have since been described. Such PTM introduce neoepitopes in proteins that can generate novel autoantibody specificities. The recent recognition of these novel specificities in RA provides a unique opportunity to understand human B-cell development in vivo. In this paper, we will review the three of the main classes of PTMs already associated with RA: citrullination, carbamylation, and oxidation. With the advancement of research methodologies it should be expected that other autoantibodies against PTM proteins could be discovered in patients with autoimmune diseases. Many of such autoantibodies may provide significant biomarker potential.

RA is a complex disease that develops as a series of events often referred to as disease continuum. RA would benefit from novel biomarker development for diagnosis where new biomarkers are still needed (even if progresses have been made with the inclusion of ACPA into the ACR/EULAR 2010 diagnostic criteria) and for prognostic notably in at risk of evolution patients with autoantibody-positive arthralgia. Risk biomarkers for rapid evolution or cardiovascular complications are also highly desirable. Monitoring biomarkers would be useful in predicting relapse. Finally, predictive biomarkers for therapy outcome would allow tailoring therapy to the individual. Increasing numbers of cytokines have been involved in RA pathology. Many have the potential as biomarkers in RA especially as their clinical utility is already established in other diseases and could be easily transferable to rheumatology. We will review the current knowledge's relation to cytokine used as biomarker in RA. However, given the complexity and heterogeneous nature of RA, it is unlikely that a single cytokine may provide sufficient discrimination; therefore multiple biomarker signatures may represent more realistic approach for the future of personalised medicine in RA.

We first determined the analytical specificity and ubiquity (i.e., the ability to detect all or most strains) of a Clostridium perfringens-specific real-time PCR (rtPCR) assay based on the cpa gene (cpa rtPCR) by using a bacterial strain panel composed of C. perfringens and non-C. perfringens Clostridium strains. All non-C. perfringens Clostridium strains tested negative, whereas all C. perfringens strains tested positive with the cpa rtPCR, for an analytical specificity and ubiquity of 100%. The cpa rtPCR assay was then used to confirm the identity of 116 putative C. perfringens isolates recovered after filtration of water samples and culture on mCP agar. Colonies presenting discordant results between the phenotype on mCP agar and cpa rtPCR were identified by sequencing the 16S rRNA and cpa genes. Four mCP(-)/rtPCR(+) colonies were identified as C. perfringens, whereas 3 mCP(+)/rtPCR(-) colonies were identified as non-C. perfringens. The cpa rtPCR was negative with all 51 non-C. perfringens strains and positive with 64 of 65 C. perfringens strains. Finally, we compared mCP agar and a CRENAME (concentration and recovery of microbial particles, extraction of nucleic acids, and molecular enrichment) procedure plus cpa rtPCR (CRENAME + cpa rtPCR) for their abilities to detect C. perfringens spores in drinking water. CRENAME + cpa rtPCR detected as few as one C. perfringens CFU per 100 ml of drinking water sample in less than 5 h, whereas mCP agar took at least 25 h to deliver results. CRENAME + cpa rtPCR also allows the simultaneous and sensitive detection of Escherichia coli and C. perfringens from the same potable water sample. In itself, it could be used to assess the public health risk posed by drinking water potentially contaminated with pathogens more resistant to disinfection.

DNA hybridization is a universal and specific mechanism for the recognition of biological targets. Some cationic polythiophene transducers sensitive to DNA structure have been previously utilized to detect such biomolecules. Further characterization of these systems indicates that both DNA sequence composition and length modulate the biosensor performance. It appears that different repeated sequence patterns cause different conformational changes of the polythiophene, from a more relaxed form to an extremely rigid one. A length difference between the DNA oligonucleotide probe and target has a detrimental effect on the fluorescent signal, but it can be attenuated by changing the sequence composition of the protruding target sequence. This demonstrates that the nature of DNA can be critical for hybridization-based detection systems., (Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The microbiological quality of 165 1 litre well water samples collected in the Québec City region was assessed by culture-based methods (mFC agar, Chromocult coliform agar, Colilert(®), MI agar, Chromocult enterococci, Enterolert™, and mEI agar) and by a molecular microbiology strategy, dubbed CRENAME-rtPCR, developed for the detection of Escherichia coli, Enterococcus spp., Enterococcus faecalis/faecium, and Bacillus atrophaeus subsp. globigii. In these drinking water samples, approved culture-based methods detected E. coli at rates varying from 1.8 to 3.6% and Enterococcus spp. at rates varying from 3.0 to 11.5%, while the molecular microbiology approach for E. coli was found to be as efficient, detecting contamination in 3.0% of samples. In contrast, CRENAME-rtPCR detected Enterococcus spp. in 27.9% of samples while the E. faecalis/faecium molecular assay did not uncover a single contaminated sample, thereby revealing a discrepancy in the coverage of waterborne enterococcal species detected by classical and molecular microbiology methods. The validation of the CRENAME-E. coli rtPCR test as a new tool to assess the quality of drinking water will require larger scale studies elaborated to demonstrate its equivalence to approved methods.

We sequenced the evolutionarily conserved genes 16S rRNA, atpD, tuf, and recA from Streptococcus pseudopneumoniae, Streptococcus pneumoniae, Streptococcus mitis, and Streptococcus oralis. Phylogenetic analysis revealed that recA provided good resolution between these species, including discrimination of the novel species S. pseudopneumoniae. By contrast, the more conserved 16S rRNA, tuf and atpD are not sufficiently discriminatory. Therefore, recA sequences were used to develop a real-time PCR assay with a locked nucleic acid-mediated TaqMan probe for the specific detection and identification of S. pseudopneumoniae. The PCR assay showed excellent specificity and a detection limit of <10 genome copies for the detection and identification of S. pseudopneumoniae strains, which makes it a promising tool for molecular identification and epidemiological studies. In conclusion, this article describes for the first time a PCR assay for the specific identification of S. pseudopneumoniae., (© 2011 The Authors. Clinical Microbiology and Infection © 2011 European Society of Clinical Microbiology and Infectious Diseases.)

Histone deacetylase inhibitors (HDACIs) are in advanced clinical development as cancer therapeutic agents. However, first generation HDACIs such as butyrate and valproate are simple short chain aliphatic compounds with moieties resembling acetyl groups, and have a broad spectrum of activity against HDACs. More complex second generation HDACIs undergoing clinical trials, such as the benzamide group compounds MS-275 and MGCD0103, are specific primarily for HDAC1 and HDAC2. To expand the repertoire of available HDACIs and HDAC specificities we created a novel benzamide-based compound named MI-192. When tested against purified recombinant HDACs, MI-192 had marked selectivity for the class I enzymes, HDAC2 and HDAC3. Screening in the NCI60 screen demonstrated that MI-192 had greatly enhanced efficacy against cells of leukaemic origin. When tested in culture against the acute myeloid leukaemic cell lines U937, HL60 and Kasumi-1, MI-192 induced differentiation and was cytotoxic through promotion of apoptosis. MI-192 therefore justifies further investigation and development as a potential therapeutic agent for use in leukaemia., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Three enterococcal isolates, CCRI-16620, CCRI-16986(T) and CCRI-16985(T), originating from water were characterized using morphological, biochemical and molecular taxonomic methods. 16S rRNA gene sequence analysis classified all three strains in the Enterococcus faecalis species group. The phylogenetic tree of 16S rRNA gene sequences showed that the three isolates form two separate branches. The first branch is represented by strains CCRI-16620 and CCRI-16986(T) and the second branch by strain CCRI-16985(T). Further sequence analysis of the housekeeping genes rpoA (encoding RNA polymerase α subunit), pheS (phenylalanyl-tRNA synthase), tufA (elongation factor Tu) and atpD (ATP synthase β-subunit) as well as the results of amplified fragment length polymorphism (AFLP) DNA fingerprinting and DNA-DNA hybridization experiments confirmed the distinct status of these strains. Moreover, biochemical tests allowed phenotypic differentiation of the strains from the other species of the E. faecalis species group. On the basis of the results obtained, the names Enterococcus ureasiticus sp. nov. (type strain CCRI-16986(T) = CCUG 59304(T) = DSM 23328(T) = LMG 26304(T)) and Enterococcus quebecensis sp. nov. (type strain CCRI-16985(T) = CCUG 59306(T) = DSM 23327(T) = LMG 26306(T)) are proposed for the two hitherto undescribed species.