Terbinafine is a novel and selective activator of the two-pore domain potassium channel TASK3

Two-pore domain potassium channels (K2Ps) are characterized by their four transmembrane domain and two-pore topology. They carry background (or leak) potassium current in a variety of cell types. Despite a number of important roles there is currently a lack of pharmacological tools with which to further probe K2P function. We have developed a cell-based thallium flux assay, using baculovirus delivered TASK3 (TWIK-related acid-sensitive K+ channel 3, KCNK9, K2P9.1) with the aim of identifying novel, selective TASK3 activators. After screening a library of 1000 compounds, including drug-like and FDA approved molecules, we identified Terbinafine as an activator of TASK3. In a thallium flux assay a pEC50 of 6.2 ( ±0.12) was observed. When Terbinafine was screened against TASK2, TREK2, THIK1, TWIK1 and TRESK no activation was observed in thallium flux assays. Several analogues of Terbinafine were also purchased and structure activity relationships examined. To confirm Terbinafine's activation of TASK3 whole cell patch clamp electrophysiology was carried out and clear potentiation observed in both the wild type channel and the pathophysiological, Birk-Barel syndrome associated, G236R TASK3 mutant. No activity at TASK1 was observed in electrophysiology studies. In conclusion, we have identified the first selective activator of the two-pore domain potassium channel TASK3

A conceptual framework for nomenclatural stability and validity of medically important fungi: a proposed global consensus guideline for fungal name changes supported by ABP, ASM, CLSI, ECMM, ESCMID-EFISG, EUCAST-AFST, FDLC, IDSA, ISHAM, MMSA, and MSGERC

The rapid pace of name changes of medically important fungi is creating challenges for clinical laboratories and clinicians involved in patient care. We describe two sources of name change which have different drivers, at the species versus the genus level. Some suggestions are made here to reduce the number of name changes. We urge taxonomists to provide diagnostic markers of taxonomic novelties. Given the instability of phylogenetic trees due to variable taxon sampling, we advocate to maintain genera at the largest possible size. Reporting of identified species in complexes or series should where possible comprise both the name of the overarching species and that of the molecular sibling, often cryptic species. Because the use of different names for the same species will be unavoidable for many years to come, an open access online database of the names of all medically important fungi, with proper nomenclatural designation and synonymy, is essential. We further recommend that while taxonomic discovery continues, the adaptation of new name changes by clinical laboratories and clinicians be reviewed routinely by a standing committee for validation and stability over time, with reference to an open access database, wherein reasons for changes are listed in a transparent way

Signals and Sensory Mechanisms that Impact Campylobacter jejuni-Host Interactions

Campylobacter jejuni is a leading cause of bacterial diarrheal disease worldwide and a frequent commensal organism of the intestinal tract of poultry and other agriculturally-important animals. Upon infection of the avian host, C. jejuni likely responds to external stimuli present within the intestinal tract to establish commensalism. The sensing mechanisms and subsequent physiological responses by C. jejuni can be crucial for initial growth and colonization and long-term persistence within the infected host. However, how many of the signals and sensing mechanisms affecting C. jejuni biology are not fully understood. In this work, I explored signal transduction mechanisms and possible in vivo signals that may influence the colonization capacity of C. jejuni. One method C. jejuni employs to monitor environmental stimuli are two-component regulatory systems (TCSs). I analyzed the potential of C. jejuni Cjj81176_1484 (Cjj1484) and Cjj81176_1483 (Cjj1483) to encode a cognate TCS that influences expression of genes possibly important for C. jejuni growth and colonization. Through transcriptome analysis, I discovered that the regulons of the Cjj1484 histidine kinase and the Cjj1483 response regulator contain many common genes, which suggests these proteins likely form a cognate TCS. I found that this TCS generally functions to repress expression of specific proteins with roles in metabolism, iron/heme acquisition, and respiration. Furthermore, the TCS repressed expression of Cjj81176_0438 and Cjj81176_0439, which had previously been found to encode a gluconate dehydrogenase complex required for commensal colonization of the chick intestinal tract. However, the TCS and other specific genes whose expression is repressed by the TCS were not required for colonization of chicks. I observed that the Cjj1483 response regulator binds target promoters both in unphosphorylated and phosphorylated forms and influences expression of some specific genes independently of the Cjj1484 histidine kinase. I propose that this TCS may sense signals found in the host intestinal tract, wherein repression of genes may be relieved. In addition to characterizing the Cjj1484/Cjj1483 TCS, I explored the role of metabolites that are commonly found in the intestines -- organic acids and short chain fatty acids (SCFAs) -- in C. jejuni commensal colonization. C. jejuni has both acetate and lactate utilization pathways, as well as for acetate production. I observed that acetogenesis mutants incapable of producing acetate were deficient for colonization of the avian intestinal tract early during infection, but not at later points during infection. Furthermore, I found that an acetogenesis mutant was impaired during growth in a defined media containing solely amino acids and organic acids as carbon sources. Transcriptome analysis of the acetogenesis mutant identified the SCFA-induced regulon which contains metabolically important genes, many of which have been implicated in C. jejuni colonization and virulence. In addition, I found that peb1C, which was downregulated in the acetogenesis mutant, was important for colonization of the chick ceca. I further confirmed in vitro that physiological concentrations of the SCFAs acetate and butyrate activated expression of the SCFA-induced regulon whereas the organic acid lactate repressed these genes. I found that in vivo expression of the SCFA-induced regulon was highest in regions of the intestinal tract where SCFAs are present in the greatest concentration. Furthermore, butyrate counteracted the inhibitory effects of lactate when the two compounds were combined in culture in vitro. I propose that C. jejuni senses the concentration of SCFAs and organic acids to discriminate between different regions of the intestinal tract and to coordinate expression of colonization genes in the preferred niche for colonization. In effect, SCFA sensing and signaling allows C. jejuni to home to appropriate sites of the host for colonization and long-term persistence

Severe Infective Endocarditis Caused by Bartonella rochalimae

A 22-year-old man from Guatemala sought care for subacute endocarditis and mycotic brain aneurysm after living in good health in the United States for 15 months. Bartonella rochalimae, a recently described human and canine pathogen, was identified by plasma microbial cell-free DNA testing. The source of infection is unknown

Combinatorial Regulation of fmgD by MrpC2 and FruA during Myxococcus xanthus Development▿

Upon starvation, a dense population of rod-shaped Myxococcus xanthus bacteria coordinate their movements to construct mounds in which some of the cells differentiate to spherical spores. During this process of fruiting body formation, short-range C-signaling between cells regulates their movements and the expression of genes important for sporulation. C-signaling activates FruA, a transcription factor that binds cooperatively with another transcription factor, MrpC2, upstream of the fmgA and fmgBC promoters, activating transcription. We have found that a third C-signal-dependent gene, herein named fmgD, is subject to combinatorial control by FruA and MrpC2. The two proteins appear to bind cooperatively upstream of the fmgD promoter and activate transcription. FruA binds proximal to the fmgD promoter, as in the fmgBC promoter region, whereas MrpC2 binds proximal to the fmgA promoter. A novel feature of the fmgD promoter region is the presence of a second MrpC2 binding site partially overlapping the promoter and therefore likely to mediate repression. The downstream MrpC2 site appears to overlap the FruA site, so the two transcription factors may compete for binding, which in both cases appears to be cooperative with MrpC2 at the upstream site. We propose that binding of MrpC2 to the downstream site represses fmgD transcription until C-signaling causes the concentration of active FruA to increase sufficiently to outcompete the downstream MrpC2 for cooperative binding with the upstream MrpC2. This would explain why fmgD transcription begins later during development and is more dependent on C-signaling than transcription of fmgA and fmgBC

Penicillium citrinum: Opportunistic pathogen or idle bystander? A case analysis with demonstration of galactomannan cross-reactivity

We present a case of an immunocompromised woman with fever, pulmonary infiltrates and multiple bronchoalveolar lavage (BAL) cultures positive for Penicillium citrinum with a concomitant high BAL galactomannan level. We report the results of Aspergillus galactomannan testing performed on culture supernatants from her P. citrinum strain that confirmed the suspected cross-reactivity. Finally, we discuss the clinical significance and antifungal susceptibility of P. citrinum in our case and review the literature

Microbiota-Derived Short-Chain Fatty Acids Modulate Expression of Campylobacter jejuni Determinants Required for Commensalism and Virulence.

Campylobacter jejuni promotes commensalism in the intestinal tracts of avian hosts and diarrheal disease in humans, yet components of intestinal environments recognized as spatial cues specific for different intestinal regions by the bacterium to initiate interactions in either host are mostly unknown. By analyzing a C. jejuni acetogenesis mutant defective in converting acetyl coenzyme A (Ac-CoA) to acetate and commensal colonization of young chicks, we discovered evidence for in vivo microbiota-derived short-chain fatty acids (SCFAs) and organic acids as cues recognized by C. jejuni that modulate expression of determinants required for commensalism. We identified a set of C. jejuni genes encoding catabolic enzymes and transport systems for amino acids required for in vivo growth whose expression was modulated by SCFAs. Transcription of these genes was reduced in the acetogenesis mutant but was restored upon supplementation with physiological concentrations of the SCFAs acetate and butyrate present in the lower intestinal tracts of avian and human hosts. Conversely, the organic acid lactate, which is abundant in the upper intestinal tract where C. jejuni colonizes less efficiently, reduced expression of these genes. We propose that microbiota-generated SCFAs and lactate are cues for C. jejuni to discriminate between different intestinal regions. Spatial gradients of these metabolites likely allow C. jejuni to locate preferred niches in the lower intestinal tract and induce expression of factors required for intestinal growth and commensal colonization. Our findings provide insights into the types of cues C. jejuni monitors in the avian host for commensalism and likely in humans to promote diarrheal disease.IMPORTANCECampylobacter jejuni is a commensal of the intestinal tracts of avian species and other animals and a leading cause of diarrheal disease in humans. The types of cues sensed by C. jejuni to influence responses to promote commensalism or infection are largely lacking. By analyzing a C. jejuni acetogenesis mutant, we discovered a set of genes whose expression is modulated by lactate and short-chain fatty acids produced by the microbiota in the intestinal tract. These genes include those encoding catabolic enzymes and transport systems for amino acids that are required by C. jejuni for in vivo growth and intestinal colonization. We propose that gradients of these microbiota-generated metabolites are cues for spatial discrimination between areas of the intestines so that the bacterium can locate niches in the lower intestinal tract for optimal growth for commensalism in avian species and possibly infection of human hosts leading to diarrheal disease

Diagnostic challenges of prolonged post-treatment clearance of Plasmodium nucleic acids in a pre-transplant autosplenectomized patient with sickle cell disease

Abstract Background Autosplenectomy, as a result of sickle cell disease, is an important risk factor for severe malaria. While molecular methods are helpful in providing rapid and accurate infection detection and species identification, the effect of hyposplenism on result interpretation during the course of infection should be carefully considered. Case presentation A 32-year old autosplenectomized Nigerian male with severe sickle cell disease was referred to the National Institutes of Health for allogenic hematopoietic stem cell transplant. Despite testing negative for malaria by both smear and PCR 2 weeks after arrival in the USA, the patient developed fever and diffuse bilateral lower rib cage and upper abdominal pain 2 weeks later and subsequently tested positive for Plasmodium falciparum. Parasitaemia was tracked over time by microscopy and nucleic acid tests to evaluate the therapeutic response in the setting of hyposplenism. The patient showed prompt resolution of patent infection by microscopy but remained positive by molecular methods for > 30 days after treatment initiation. Conclusion While molecular testing can provide sensitive Plasmodium nucleic acid detection, the persistence of Plasmodium nucleic acids following adequate treatment in functionally asplenic patients can lead to a diagnostic dilemma. In such patients, clinical response and peripheral blood smears should guide patient management following treatment. Nonetheless, in pre-transplant patients at high-risk for pre-existing Plasmodium infections, highly sensitive molecular assays can be useful to rule out infection prior to transplantation