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Objectives IMP-type carbapenemases are rarely detected in Europe and limited information is available to guide the treatment of infections caused by carbapenemase-producing Enterobacterales (CPE) producing these carbapenemases. Accurate antimicrobial susceptibility testing (AST) results are essential for optimal antibiotic management. Here we report discrepancies in AST of IMP-producing Enterobacterales (IMP-CPE) complicating the management of severe sepsis. Methods Antimicrobial susceptibilities were analysed by in-house VITEK® 2, Etest and broth microdilution (BMD). Carbapenemase-encoding genes were detected by PCR. Whole-genome sequencing (WGS) was performed using an Illumina MiSeq platform. Results Minimum inhibitory concentrations (MICs) determined by VITEK® 2 for Enterobacter hormaechei and Klebsiella oxytoca blood culture isolates were ≥16 mg/L for meropenem and ≤0.5 mg/L for ertapenem. In contrast, Etest analysis and BMD returned MICs of 2 mg/L and 1 mg/L, respectively. Both isolates tested positive for IMP carbapenemase-encoding genes by PCR. WGS revealed that both isolates carried the same blaIMP-4 gene. Based on VITEK® 2 susceptibilities, initial treatment was with tigecycline and amikacin. After subsequent deterioration, the patient was successfully treated with ertapenem and amikacin. Conclusion This case highlights that automated AST by VITEK® 2 can over-report meropenem resistance for IMP carbapenemase-producers compared with Etest and BMD. Clinicians need to be cautious deciding against carbapenem treatment based on VITEK® 2 susceptibility testing results for IMP-positive Enterobacterales. Tigecycline was inferior to carbapenem treatment for pyelonephritis caused by isolates expressing IMP carbapenemases, however specific evidence guiding the treatment of these infections is lacking.

A Gram-negative rod from the Yersinia genus was isolated from a clinical case of yersiniosis in the United Kingdom. Long read sequencing data from an Oxford Nanopore Technologies (ONT) MinION in conjunction with Illumina HiSeq reads were used to generate a finished quality genome of this strain. Overall Genome Related Index (OGRI) of the strain was used to determine that it was a novel species within Yersinia , despite biochemical similarities to Yersinia enterocolitica . The 16S ribosomal RNA gene accessions are MN434982-MN434987 and the accession number for the complete and closed chromosome is CP043727. The type strain is SRR7544370T (=NCTC 14382T/=LMG 31573T).

In recent years, the concept of bacteria-mediated cancer therapy has gained significant attention as an alternative to conventional therapy. The focus has been on non-typhoidal Salmonella (NTS), particularly S. Typhimurium, for its anti-cancer properties, however, other NTS serovars such as Salmonella Oslo, which are associated with foodborne illnesses could potentially be effective anti-cancer agents. Here, we report the draft genome sequence of Salmonella Oslo isolated from seafood and its laboratory generated auxotrophic mutant.

A Gram-stain-negative, rod-shaped strain isolated from pig-production environments was identified as a new species within the genus Yersinia using multifaceted genomic and biochemical approaches. The genome of this strain was closed using a hybrid assembly approach combining both high accuracy short read sequencing data with long read sequencing technology. Phylogenetic analysis of the 16S rRNA gene showed ~98 % similarity to Yersinia kristensenii and ~98 % similarity to Yersinia enterocolitica . Average nucleotide identity (OrthoANI) values were calculated as 85.79 % to Y. kristensenii ATCC 33638T and 85.73 % to Y. enterocolitica ATCC 9610T thereby providing evidence that this isolate should be considered as a novel species. The type strain is CFS1934T (=NCTC 14222T=LMG 31076T).

Nontyphoidal Salmonella species are globally disseminated pathogens and are the predominant cause of gastroenteritis. The pathogenesis of salmonellosis has been extensively studied using in vivo murine models and cell lines, typically challenged with Salmonella enterica serovar Typhimurium. Although S. enterica serovars Enteritidis and Typhimurium are responsible for most of the human infections reported to the Centers for Disease Control and Prevention (CDC), several other serovars also contribute to clinical cases of salmonellosis. Despite their epidemiological importance, little is known about their infection phenotypes. Here, we report the virulence characteristics and genomes of 10 atypical S. enterica serovars linked to multistate foodborne outbreaks in the United States. We show that the murine RAW 264.7 macrophage model of infection is unsuitable for inferring human-relevant differences in nontyphoidal Salmonella infections, whereas differentiated human THP-1 macrophages allowed these isolates to be further characterized in a more human-relevant context.

Cationic antimicrobial peptides have raised interest as attractive alternatives to classical antibiotics, and also have utility in preventing food spoilage. We set out to enrich cationic antimicrobial peptides from milk hydrolysates using gels containing various ratios of anionic pectin/alginate. All processes were carried out with food-grade materials in order to suggest food-safe methods suited for producing food ingredients or supplements. Hydrolysed caseinate peptides retained in the gel fraction, identified by mass spectrometry, were enriched for potential antimicrobial peptides, as judged by a computational predictor of antimicrobial activity. Peptides retained in a 60:40 pectin:alginate gel fraction had a strong antimicrobial effect against 8 tested bacterial strains with a minimal inhibitory concentration of 1.5–5 mg/mL, while the unfractionated hydrolysate only had a detectable effect in one of the eight strains. Among 110 predicted antimicrobial peptides in the gel fraction, four are known antimicrobial peptides, HKEMPFPK, TTMPLW, YYQQKPVA and AVPYPQR. These results highlight the potential of pectin/alginate food-gels based processes as safe, fast, cost-effective methods to separate and enrich for antimicrobial peptides from complex food protein hydrolysates.

One mcr-1 -carrying Salmonella enterica serovar Indiana strain D90, was identified from 1320 Salmonella enterica isolates from poultry slaughterhouse in 2012 in China. The objective of this study was to verify the transferability of the mcr-1 gene and also completely characterize the sequence of the strain at the whole-genome level. Broth matting assays were carried out to detect the transferability and whole-genome sequencing (WGS) of S. enterica serovar Indiana D90 was performed using the PacBio RS II system. Open reading frames were assigned using Rapid Annotation using Subsystem Technology (RAST) and analysed by BLASTn and BLASTp. Salmonella Pathogenisity Islands (SPIs) were annotated by SPIFinder platform. The complete genome sequence of S. enterica serovar Indiana D90 contained a circular 4,779,514-bp chromosome and four plasmids. Genome analysis and sequencing revealed that 24 multi-drug resistance (MDR) genes were located on plasmids. The largest plasmid pD90-1, was found to be of an IncHI2/HI2A/Q1/N type that encoded a bla CTX-M-65 gene along with 20 additional antimicrobial resistance genes. A 60.5-kbp IncI2 plasmid pD90-2 contained a nikA - nikB - mcr-1 genetic structure, that can be successfully transferred to E. coli and S. enterica serovar Typhimurium at low transfer rates. Interestingly, comparative sequence analysis revealed the plasmids pD90-1 and pD90-2 showed considerable nucleotide similarity to pHNSHP45-2 and pHNSHP45, respectively. Moreover, the genome and the plasmid pD90-2 also showed high similarity to one carbapenem resistant S. enterica serovar Indiana strain, C629 and its plasmid pC629, respectively. This is the first report of the complete nucleotide sequence of one mcr-1 -carrying MDR S. enterica serovar Indiana strain.

In recent years, the concept of bacteria-mediated cancer therapy has gained significant attention as an alternative to conventional therapy. The focus has been on non-typhoidal Salmonella (NTS), particularly S. Typhimurium, for its anti-cancer properties, however, other NTS serovars such as Salmonella Oslo, which are associated with foodborne illnesses could potentially be effective anti-cancer agents. Here, we report the draft genome sequence of Salmonella Oslo isolated from seafood and its laboratory generated auxotrophic mutant.

The factors influencing the virulence of P. aeruginosa in the development of invasive infection remain poorly understood. Here, we investigated the role of the host microenvironment in shaping pathogen virulence and investigated the mechanisms involved. Comparing seven paired genetically indistinguishable clinical bloodstream and peripheral isolates of P. aeruginosa, we demonstrate that isolates derived from bloodstream infections are more virulent than their peripheral counterparts (p = 0.025). Bloodstream and peripheral isolates elicited similar NF-kB responses in a THP-1 monocyte NF-kappaB reporter cell line implicating similar immunogenicity. Proteomic analysis by mass spectrometry identified multiple virulence and virulence-related factors including LecA and RpoN in significantly greater abundance in the bacterial supernatant from the bloodstream isolate in comparison to that from the corresponding peripheral isolate. Investigation by qPCR revealed that control of expression of these virulence factors was not due to altered levels of transcription. Based on these data, we hypothesize a post-transcriptional mechanism of virulence regulation in P. aeruginosa bloodstream infections influenced by surrounding microenvironmental conditions. Science Foundation Ireland

Klebsiella pneumoniae is an important nosocomial pathogen with an extraordinary resistant phenotype due to a combination of acquired resistant-elements and efflux mechanisms. In this study a detailed molecular characterization of 11 K. pneumoniae isolates of clinical origin was carried out. Eleven clinical isolates were tested for their susceptibilities, by disk diffusion and broth microdilution and interpreted according to CLSI guidelines. Efflux activity was determined by measuring the extrusion of ethidium bromide and biofilm formation was assessed following static growth in Müeller-Hinton and minimal media M9 broths at two temperatures and time points. Template DNA from all 11 isolates was extracted and sequenced. The study collection was found to be resistant to several (extended-spectrum beta-lactam) ESBL-type compounds along with several (fluoro)quinolones (FQ). Resistance to tetracycline accounted for 55% of the study collection (n = 6) and three of the 11 isolates were resistance to carbapenems. Genotyping identified blaCTX-M-15 (82%), blaSHV-12 (55%), and blaTEM-1B (45%) ESBL encoding genes and FQ resistance was associated the presence of the oqxAB operon, identified in 10 of the 11 isolates and qnrB gene in one isolate. The polymorphisms detected in the quinolone resistance-determining regions (QRDRs) were associated with isolates of the clonal group CG15. Sequence types (ST) identified were representative of previously described clonal groups including CG258 (n = 7), CG15 (n = 3), and CG147 (n = 1). Plasmid replicon type databases were queried indicating the presence of IncFII and IncFIB replicon types in the majority of the isolates (91%), followed by IncFIA (45%), and IncR (45%). Two of the 11 isolates were found positive for yersiniabactin siderophore-encoding genes. No differences in the ability to efflux ethidium bromide were identified. Biofilm formation was stronger when the isolates were grown under stressed conditions at 37°C for a period up to 96 h. These data confirm the fact that well-recognized clonal groups of K. pneumoniae of importance to human health carries a diverse repertoire of antimicrobial resistance determinants, particularly related to critically important drugs in the ESBL and FQ classes. The capacity of most isolates to form strong biofilms, when stressed under laboratory-simulated conditions, supports the risk to human health associated with nosocomial infections deriving from indwelling medical devices.

The synthesis of several diorganotin(IV) dicarboxylate compounds, including acetates and nicotinates as well as diorganotin(IV) dichloride complexes of the ligands phen, dione and dppz were undertaken. Several difficulties in either the syntheses or complexation reactions with the organic ligands were encountered. The diorganotin(IV) dichloride complexes of the ligands (R2SnCl2·L where R = Me, n-Bu or Ph and L = phen, dione or dppz) were tested against Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa. The dibutyltin(IV) derivatives exhibited the broadest range of activity in comparison to the dimethyltin(IV) or diphenyltin(IV) derivatives. The addition of the nicotinate group did not promote activity against any of the bacteria. Furthermore, only in the case of Ph2SnCl2·dione was there improved activity compared to the organic ligand itself.

Here, we report the draft genome sequence of the lactose-negative, rifampin-resistant, Escherichia coli strain 26R 793. This isolate has been widely used in conjugation experiments as a general recipient strain.

Blown pack spoilage (BPS) is a major issue for the beef industry. Etiological agents of BPS involve members of a group of Clostridium species, including Clostridium estertheticum which has the ability to produce gas, mostly carbon dioxide, under anaerobic psychotrophic growth conditions. This spore-forming bacterium grows slowly under laboratory conditions, and it can take up to 3 months to produce a workable culture. These characteristics have limited the study of this commercially challenging bacterium. Consequently information on this bacterium is limited and no effective controls are currently available to confidently detect and manage this production risk. In this study the complete genome of C. estertheticum DSM 8809 was determined by SMRT® sequencing. The genome consists of a circular chromosome of 4.7 Mbp along with a single plasmid carrying a potential tellurite resistance gene tehB and a Tn3-like resolvase-encoding gene tnpR. The genome sequence was searched for central metabolic pathways that would support its biochemical profile and several enzymes contributing to this phenotype were identified. Several putative antibiotic/biocide/metal resistance-encoding genes and virulence factors were also identified in the genome, a feature that requires further research. The availability of the genome sequence will provide a basic blueprint from which to develop valuable biomarkers that could support and improve the detection and control of this bacterium along the beef production chain.

Identification of Enterobacteriaceae harbouring the plasmid-mediated transferable colistin resistance gene mcr-1 presents a new challenge to public health. The aim of this study was to characterise multidrug-resistant Shiga toxin-producing Escherichia coli (STEC) harbouring the mcr-1 gene on plasmids cultured from pigs in China. Using CHROMagar™ ECC plates combined with stx gene detection by PCR, 93 STEC were recovered from 326 faecal, 351 small intestine content and 326 colon content samples taken from healthy pigs in 2011 and 2012 in China. This study, in which ten colistin-resistant isolates with minimum inhibitory concentrations (MICs) of 8-12 mg/L were identified and found to be positive by PCR for the mcr-1 gene, is a follow-up to an earlier investigation. Plasmid profiling by S1-nuclease digestion followed by pulsed-field gel electrophoresis (PFGE) identified several high-molecular-weight plasmids and these were typed by PCR-based replicon typing (PBRT). Two of the ten isolates, namely STEC-CQ09 (O116:H11/CC23/ST88) and CQ10 (O2:H32/ST3628), were selected for further study as described in this report.

To provide a review of the medical literature and practical experience in the diagnosis, history, and physical examination of the spine with emphasis on the administration of spinal injections and their role in the treatment of painful spinal conditions.

Congress has recently focused on the complex relationship between federal student aid, states' funding appropriations for higher education, and institutional tuition and fee levels. Fueling this focus is the ongoing cost shift in public higher education, from states to students and families, as well as to the federal government via student aid programs. This shift in who pays for education is primarily a consequence of gradual state disinvestment in public higher education. As a result, college officials have compensated for the loss of state dollars with a combination of cost-cutting measures, reductions in student enrollment, and an increased reliance on student tuition and fee revenues. The shift in higher education funding, from states to students—driven by insufficient, and in many cases, sharply reduced state appropriations for higher education—has placed more pressure on federal lawmakers to expand existing student aid programs. Increased federal investment in student aid has helped negate the impact of rising tuition and fees. The trend in states' disinvestment in public higher education is especially problematic considering the burgeoning student enrollments occurring in many states. As a result, recent federal legislation has included financial incentives for state lawmakers to maintain specified minimum funding levels for public higher education in order to dissuade states from substantially reducing their appropriation commitments. In establishing these "Maintenance of Effort" (MOE) provisions that determine minimum funding thresholds states must meet in order to receive specified federal funds, Congress intended for these federal monies to supplement state resources aimed at supporting institutions and students, not supplant states' fiscal commitments to higher education.

Building a comprehensive knowledge base of the association of Listeria monocytogenes isolates across national food chains, clinical cases, and environments can play a key role in helping control the incidence of listeriosis. Today, many food chains cross national borders and are often shared by neighboring countries. This study characterized L. monocytogenes isolated from food samples in Northern Ireland and investigated whether similarities in the population and associations of L. monocytogenes strains exist in the neighboring countries of Northern Ireland and the Republic of Ireland, which together constitute the island of Ireland. Listeria monocytogenes isolates were characterized using serotyping and pulsed-field gel electrophoresis subtyping. This data was then interrogated against existing data for the Republic of Ireland, to identify any shared trends in the ecology and contamination patterns of L. monocytogenes strains. The results of this study indicated that contaminated food products often shared L. monocytogenes strains with other products. A total of six different strain subtypes were identified among 18 contaminated products. Overall strain diversity in positive samples was low, with no sample yielding more than one L. monocytogenes strain, as determined by pulsed-field gel electrophoresis subtyping. When comparisons against an Irish strain database were performed, many related strain subtypes were also shared by a variety of sources in the Republic of Ireland. This study highlights the potential benefits that a whole-island surveillance approach may present to food safety and public health in both Northern Ireland and the Republic of Ireland.

A quantitative, peripherally accessible biomarker for neuropathic pain has great potential to improve clinical outcomes. Based on the premise that peripheral and central immunity contribute to neuropathic pain mechanisms, we hypothesized that biomarkers could be identified from the whole blood of adult male rats, by integrating graded chronic constriction injury (CCI), ipsilateral lumbar dorsal quadrant (iLDQ) and whole blood transcriptomes, and pathway analysis with pain behavior. Correlational bioinformatics identified a range of putative biomarker genes for allodynia intensity, many encoding for proteins with a recognized role in immune/nociceptive mechanisms. A selection of these genes was validated in a separate replication study. Pathway analysis of the iLDQ transcriptome identified Fcγ and Fcε signaling pathways, among others. This study is the first to employ the whole blood transcriptome to identify pain biomarker panels. The novel correlational bioinformatics, developed here, selected such putative biomarkers based on a correlation with pain behavior and formation of signaling pathways with iLDQ genes. Future studies may demonstrate the predictive ability of these biomarker genes across other models and additional variables.

The synaptopodin 2 gene can be differently spliced, resulting in three mRNAs with varying 3′ ends coding for different C-termini (De Ganck et al 2008). The calculated molecular weights of these proteins are 117, 119, and 136 kDa. These isoforms require the expression of exons 1–3. However, only a single protein product has been detected in mammalian muscle lacking the product of these first exons.We extracted four fesselin isoforms from avian smooth muscle tissue. These include the first isolated 79 and 103 kDa isoforms (Leinweber et al. 1999). The newly detected isoforms migrated on SDS gels with apparent molecular masses of 140 and 160 kDa. In contrast to our initial assumption that the 79 kDa was a proteolytic product of the 103 kDa protein we now show that they are different spliceforms. The 79 kDa isoform forms the core of synaptopodin 2. The other isoforms have different extensions at either the N- or C-terminal regions.The different isoforms were differentially extracted by different buffers. Extractions were most complete under conditions that depolymerized both actin and intermediate filaments. Surprisingly although fesselin binds to actin and myosin none of the four different isoforms was extracted with the acto-myosin-complex. The extraction data suggest that fesselin functions in actin filament organization rather than in regulation of actin-myosin interactions. In contrast to smooth muscle tissue we detected one isoform of fesselin in skeletal and heart muscle tissue in agreement with the findings in mammalian tissue. In avian skeletal muscle we observed a 79 kDa isoform and in heart muscle a 170 kDa isoform. The reason for the differential expression of fesselin in different muscle types is unknown.