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We conducted patient chart reviews and whole-genome sequencing of wound specimens containing presumptive Corynebacterium ulcerans from Vancouver, British Columbia, Canada, during July 2019-July 2023. Sequencing confirmed 8/14 isolates were C. ramonii and identified 2 distinct clusters. Molecular methods should be used to clinically differentiate potential toxin-producing Corynebacterium spp. [ABSTRACT FROM AUTHOR]

Background Shigella sonnei has caused sexually transmitted enteric infections in men who have sex with men (MSM) in Vancouver. We recently observed a high rate of multidrug-resistant (MDR) S. sonnei bacteremia among persons experiencing homelessness (PEH). We aimed to describe the wider epidemiology, clinical outcomes, and genomics of S. sonnei infections over time. Methods A retrospective review of 163 patients with S. sonnei infections was undertaken from 2015 to 2022. We collected demographic, clinical, and microbiological data over 2 time periods: historical (2015–2020) and recent (2021–2022). Severe shigellosis definition included hospitalization, bacteremia, or death. Whole-genome sequencing was performed to identify genotype, infer relatedness, and predict antimicrobial resistance. Results S. sonnei infections increased from 8.3 (historical period) to 56.5 (recent period) cases/year. Over time, the primary population characteristics associated with shigellosis shifted from MSM (45; 98%) to PEH (86; 77%). The population intersection between MSM and PEH historically and recently was similar and occurred in 3 (6%) and 10 (9%) of patients, respectively. Severe shigellosis was significantly higher in the recent versus historical period (69 [61%] vs 7 [14%]; P <.001). A dominant clone of MDR S. sonnei , 3.6.1.1.2 (CipR.MSM5), emerged with resistance to all first- and second-line agents, yet with susceptibility to ceftriaxone. Conclusions We observed a substantial increase in severe shigellosis and shift from sexually transmitted S. sonnei infections in MSM to likely environmental transmission among PEH. More severe disease associated with the 3.6.1.1.2 clone of MDR S. sonnei in PEH could be a result of underlying vulnerabilities of the affected population. [ABSTRACT FROM AUTHOR]

A robust surveillance system for early identification of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) is of critical public health value. VOCs have demonstrated in vitro [...]

Since the introduction of the diphtheria toxoid vaccine, cases of diphtheria caused by toxigenic strains of Corynebacterium diphtheriae have decreased (1). More recently, however, C. diphtheriae appears to have reemerged, [...]

Background and Aim: Patients suspected of Alpha 1-Antitrypsin (A1AT) abnormality based on low serum concentration are routinely confirmed through polymerase chain reaction (PCR) testing of peripheral blood. Genotyping formalin- fixed paraffin-embedded (FFPE) tissue is a novel approach that could aid in detecting variant A1AT. We performed qPCR on FFPE liver explants with Periodic Acid Schiff after Diastase (PASD)- and A1AT-positive globules to confirm and estimate the frequency of A1AT deficiency in transplant cases. Materials and Methods: Eighteen (12.68%) of 142 patients with end-stage liver disease showed PASD/A1AT positive globules. FFPE of the explants was tested through qPCR to detect S and Z alleles. A second age- and sex-matched control group consisting of five liver transplant patients with negative globules was included in the study. Results: qPCR assay was successful with all the samples meeting QC parameters. All patients included in the study elucidated Z allele variants; 2 homozygous (11.1%) and 16 heterozygous (88.9%). The control group demonstrated normal wild-type MM allele. Conclusion: Screening for A1AT deficiency using serum levels is not sufficiently sensitive to detect deficiency, especially in carriers. If A1AT testing was not performed preoperatively and the risk is high based on the PASD/ A1AT-positive globules in the explants, then molecular testing of FFPE tissue can be a viable method for confirming the diagnosis. [ABSTRACT FROM AUTHOR]

Objectives To utilize long-read nanopore sequencing (R10.4.1 flowcells) for WGS of a cluster of MDR Shigella sonnei , specifically characterizing genetic predictors of antimicrobial resistance (AMR). Methods WGS was performed on S. sonnei isolates identified from stool and blood between September 2021 and October 2022. Bacterial DNA from clinical isolates was extracted on the MagNA Pure 24 and sequenced on the GridION utilizing R10.4.1 flowcells. Phenotypic antimicrobial susceptibility testing was interpreted based on CLSI breakpoints. Sequencing data were processed with BugSeq, and AMR was assessed with BugSplit and ResFinder. Results Fifty-six isolates were sequenced, including 53 related to the cluster of cases. All cluster isolates were identified as S. sonnei by sequencing, with global genotype 3.6.1.1.2 (CipR.MSM5), MLST 152 and PopPUNK cluster 3. Core genome MLST (cgMLST, examining 2513 loci) and reference-based MLST (refMLST, examining 4091 loci) both confirmed the clonality of the isolates. Cluster isolates were resistant to ampicillin (bla TEM-1), trimethoprim/sulfamethoxazole (dfA1 , dfrA17 ; sul 1, sul 2), azithromycin (ermB , mphA) and ciprofloxacin (gyrA S83L, gyrA D87G, parC S80I). No genomic predictors of resistance to carbapenems were identified. Conclusions WGS with R10.4.1 enabled rapid sequencing and identification of an MDR S. sonnei community cluster. Genetic predictors of AMR were concordant with phenotypic antimicrobial susceptibility testing. [ABSTRACT FROM AUTHOR]

Background: To support the implementation of high-throughput pipelines suitable for SARS-CoV-2 sequencing and analysis in a clinical laboratory, we developed an automated sample preparation and analysis workflow. Methods: We used the established ARTIC protocol with approximately 400 bp amplicons sequenced on Oxford Nanopore's MinION. Sequences were analyzed using Nextclade, assigning both a clade and quality score to each sample. Results: A total of 2179 samples on twenty-five 96-well plates were sequenced. Plates of purified RNA were processed within 12 h, sequencing required up to 24 h, and analysis of each pooled plate required 1 h. The use of samples with known threshold cycle (Ct) values enabled normalization, acted as a quality control check, and revealed a strong correlation between sample Ct values and successful analysis, with 85% of samples with Ct < 30 achieving a "good" Nextclade score. Less abundant samples responded to enrichment with the fraction of Ct > 30 samples achieving a "good" classification rising by 60% after addition of a post-ARTIC PCR normalization. Serial dilutions of 3 variant of concern samples, diluted from approximately Ct = 16 to approximately Ct = 50, demonstrated successful sequencing to Ct = 37. The sample set contained a median of 24 mutations per sample and a total of 1281 unique mutations with reduced sequence read coverage noted in some regions of some samples. A total of 10 separate strains were observed in the sample set, including 3 variants of concern prevalent in British Columbia in the spring of 2021. Conclusions: We demonstrated a robust automated sequencing pipeline that takes advantage of input Ct values to improve reliability. [ABSTRACT FROM AUTHOR]

Kang, Hyung Sub, Dirk Kerstan, Long-Jun Dai, Gordon Ritchie, and Gary A. Quamme. Adenosine modulates [Mg.sup.2+] uptake in distal convoluted tubule cells via [A.sub.1] and [A.sub.2] purinoceptors. Am J Physiol Renal Physiol 281: F1141-F1147, 2001.--tk;1Adenosine plays a role in the control of water and electrolyte reabsorption in the distal tubule. As the distal convoluted tubule is important in the regulation of renal [Mg.sup.2+] balance, we determined the effects of adenosine on cellular [Mg.sup.2+] uptake in this segment. The effect of adenosine was studied on immortalized mouse distal convoluted tubule (MDCT) cells, a model of the intact distal convoluted tubule. The rate of [Mg.sup.2+] uptake was measured with fluorescence techniques using mag-fura 2. To assess [Mg.sup.2+] uptake, MDCT cells were first [Mg.sup.2+] depleted to 0.22 [+ or -] 0.01 mM by being cultured in [Mg.sup.2+]-free media for 16 h and then placed in 1.5 mM Mg[Cl.sub.2]; next, changes in intracellular [Mg.sup.2+] concentration ([[Mg.sup.2+]][.sub.i]) were determined. [[Mg.sup.2+]][.sub.i] returned to basal levels, 0.53 [+ or -] 0.02 mM, with a mean refill rate, d([[Mg.sup.2+]][.sub.i])/dt, of 137 [+ or -] 16 nM/s. Adenosine stimulates basal [Mg.sup.2+] uptake by 41 [+ or -] 10%. The selective [A.sub.1] purinoceptor agonist [N.sup.6]-cyclopentyladenosine (CPA) increased intracellular [Ca.sup.2+] and decreased parathyroid hormone (PTH)-stimulated cAMP formation and PTH-mediated [Mg.sup.2+] uptake. On the other hand, the selective [A.sub.2] receptor agonist 2- [p-(2-carbonyl-ethyl)-phenylethylamino]-5'-N-ethyl-carboxamidoadenosine (CGS) stimulated [Mg.sup.2+] entry in a concentration-dependent fashion. CGS increased cAMP formation and the protein kinase A inhibitor RpcAMPS inhibited CGS-stimulated [Mg.sup.2+] uptake. Selective inhibition of phospholipase C, protein kinase C, or mitogen-activated protein kinase enzyme cascades with U-73122, Ro-31-8220, and PD-98059, respectively, diminished [A.sub.2] agonist-mediated [Mg.sup.2+] entry. Aldosterone potentiated CGS-mediated [Mg.sup.2+] entry, and elevation of extracellular [Ca.sup.2+] diminished CGS-responsive cAMP formation and [Mg.sup.2+] uptake. Accordingly, MDCT cells possess both [A.sub.1] and [A.sub.2] purinoceptor subtypes with intracellular signaling typical of these respective receptors. We conclude that adenosine has dual effects on [Mg.sup.2+] uptake in MDCT cells through separate [A.sub.1] and [A.sub.2] purinoceptor pathways. intracellular magnesium; fluorescence; intracellular calcium transients; intracellular adenosine 3'5-cyclic monophosphate; immortalized mouse distal convoluted tubule cells Received 16 March 2001; accepted in final form 25 July 2001