Vulvovaginal Trichosporonosis

Objective: Isolation of Trichosporon species from vaginal secretions is a rare event, and no data are available on its pathogenic role. A case series is presented to determine the pathogenic role of Trichosporon species in vulvovaginal infections. Methods: We performed a retrospective chart review of patients seen in the W.S.U. Vaginitis Clinic in order to identify patients from whom Trichosporon species were isolated. Results: Between 1986 and 2001, a total of 13 patients had a total of 18 positive vaginal cultures for Trichosporon species. All 18 vaginal isolates were T. inkin. In general, positive vaginal cultures were accompanied by low yeast colony counts. Four out of 18 positive T. inkin cultures were obtained from visits by asymptomatic patients. Of the remaining 14 positive T. inkin cultures from patients with symptoms, nine out of 14 cultures had other diagnoses (Candida albicans, six cases; bacterial vaginosis, two cases; Trichomonas, one case). Five positive T. inkin cultures were obtained from visits at which patients had symptoms and no associated diagnosis. In only one of the five episodes could we establish a clear pathogenic role for Trichosporon. In this case the patient was treated with boric acid and had resolution of symptoms and a negative culture at follow-up. In-vitro susceptibility tests revealed that T. inkin was resistant to flucytosine and susceptible to all topical and oral azoles. Conclusions: T. inkin is occasionally found in vulvovaginal cultures and is usually a non-pathogen. Transient colonization tended to occur in women, usually of African—American origin, with major perturbations in vaginal flora (bacterial vaginosis and trichomoniasis) and increased pH. Pathogenic consequences of Trichosporon colonization appear to be rare

Inhibiting the oncogenic translation program is an effective therapeutic strategy in multiple myeloma

Published in final edited form as: Sci Transl Med. 2017 May 10; 9(389). https://doi.org/10.1126/scitranslmed.aal2668.Multiple myeloma (MM) is a frequently incurable hematological cancer in which overactivity of MYC plays a central role, notably through up-regulation of ribosome biogenesis and translation. To better understand the oncogenic program driven by MYC and investigate its potential as a therapeutic target, we screened a chemically diverse small-molecule library for anti-MM activity. The most potent hits identified were rocaglate scaffold inhibitors of translation initiation. Expression profiling of MM cells revealed reversion of the oncogenic MYC-driven transcriptional program by CMLD010509, the most promising rocaglate. Proteome-wide reversion correlated with selective depletion of short-lived proteins that are key to MM growth and survival, most notably MYC, MDM2, CCND1, MAF, and MCL-1. The efficacy of CMLD010509 in mouse models of MM confirmed the therapeutic relevance of these findings in vivo and supports the feasibility of targeting the oncogenic MYC-driven translation program in MM with rocaglates

Design, construction and characterization of a set of insulated bacterial promoters

We have generated a series of variable-strength, constitutive, bacterial promoters that act predictably in different sequence contexts, span two orders of magnitude in strength and contain convenient sites for cloning and the introduction of downstream open-reading frames. Importantly, their design insulates these promoters from the stimulatory or repressive effects of many 5′- or 3′-sequence elements. We show that different promoters from our library produce constant relative levels of two different proteins in multiple genetic contexts. This set of promoters should be a useful resource for the synthetic-biology community

Engineering transcription factors with novel DNA-binding specificity using comparative genomics

The transcriptional program for a gene consists of the promoter necessary for recruiting RNA polymerase along with neighboring operator sites that bind different activators and repressors. From a synthetic biology perspective, if the DNA-binding specificity of these proteins can be changed, then they can be used to reprogram gene expression in cells. While many experimental methods exist for generating such specificity-altering mutations, few computational approaches are available, particularly in the case of bacterial transcription factors. In a previously published computational study of nitrogen oxide metabolism in bacteria, a small number of amino-acid residues were found to determine the specificity within the CRP (cAMP receptor protein)/FNR (fumarate and nitrate reductase regulatory protein) family of transcription factors. By analyzing how these amino acids vary in different regulators, a simple relationship between the identity of these residues and their target DNA-binding sequence was constructed. In this article, we experimentally tested whether this relationship could be used to engineer novel DNA–protein interactions. Using Escherichia coli CRP as a template, we tested eight designs based on this relationship and found that four worked as predicted. Collectively, these results in this work demonstrate that comparative genomics can inform the design of bacterial transcription factors

Circular Single-Stranded Synthetic DNA Delivery Vectors for MicroRNA

Single-stranded (ss) circular oligodeoxynucleotides were previously found to undergo rolling circle transcription (RCT) by phage and bacterial RNA polymerases (RNAPs) into tandemly repetitive RNA multimers. Here, we redesign them to encode minimal primary miRNA mimics, with the long term aim of intracellular transcription followed by RNA processing and maturation via endogenous pathways. We describe an improved method for circularizing ss synthetic DNA for RCT by using a recently described thermostable RNA ligase, which does not require a splint oligonucleotide to juxtapose the ligating ends. In vitro transcription of four templates demonstrates that the secondary structure inherent in miRNA-encoding vectors does not impair their RCT by RNAPs previously shown to carry out RCT. A typical primary-miRNA rolling circle transcript was accurately processed by a human Drosha immunoprecipitate, indicating that if human RNAPs prove to be capable of RCT, the resulting transcripts should enter the endogenous miRNA processing pathway in human cells. Circular oligonucleotides are therefore candidate vectors for small RNA delivery in human cells, which express RNAPs related to those tested here

Evaluation of the bacterial diversity of Pressure ulcers using bTEFAP pyrosequencing

<p>Abstract</p> <p>Background</p> <p>Decubitus ulcers, also known as bedsores or pressure ulcers, affect millions of hospitalized patients each year. The microflora of chronic wounds such as ulcers most commonly exist in the biofilm phenotype and have been known to significantly impair normal healing trajectories.</p> <p>Methods</p> <p>Bacterial tag-encoded FLX amplicon pyrosequencing (bTEFAP), a universal bacterial identification method, was used to identify bacterial populations in 49 decubitus ulcers. Diversity estimators were utilized and wound community compositions analyzed in relation to metadata such as Age, race, gender, and comorbidities.</p> <p>Results</p> <p>Decubitus ulcers are shown to be polymicrobial in nature with no single bacterium exclusively colonizing the wounds. The microbial community among such ulcers is highly variable. While there are between 3 and 10 primary populations in each wound there can be hundreds of different species present many of which are in trace amounts. There is no clearly significant differences in the microbial ecology of decubitus ulcer in relation to metadata except when considering diabetes. The microbial populations and composition in the decubitus ulcers of diabetics may be significantly different from the communities in non-diabetics.</p> <p>Conclusions</p> <p>Based upon the continued elucidation of chronic wound bioburdens as polymicrobial infections, it is recommended that, in addition to traditional biofilm-based wound care strategies, an antimicrobial/antibiofilm treatment program can be tailored to each patient's respective wound microflora.</p

Multiple mechanisms disrupt the let-7 microRNA family in neuroblastoma

Poor prognosis in neuroblastoma is associated with genetic amplification of MYCN. MYCN is itself a target of let-7, a tumour suppressor family of microRNAs implicated in numerous cancers. LIN28B, an inhibitor of let-7 biogenesis, is overexpressed in neuroblastoma and has been reported to regulate MYCN. Here we show, however, that LIN28B is dispensable in MYCN-amplified neuroblastoma cell lines, despite de-repression of let-7. We further demonstrate that MYCN messenger RNA levels in amplified disease are exceptionally high and sufficient to sponge let-7, which reconciles the dispensability of LIN28B. We found that genetic loss of let-7 is common in neuroblastoma, inversely associated with MYCN amplification, and independently associated with poor outcomes, providing a rationale for chromosomal loss patterns in neuroblastoma. We propose that let-7 disruption by LIN28B, MYCN sponging, or genetic loss is a unifying mechanism of neuroblastoma development with broad implications for cancer pathogenesis.United States. National Institutes of Health (R01GM107536)Alex's Lemonade Stand FoundationHoward Hughes Medical InstituteBoston Children's Hospital. Manton Center for Orphan Disease ResearchNational Institute of General Medical Sciences (U.S.) (T32GM007753

Molecular cloning and expression of the biodegradative threonine dehydratase gene ( tdc ) of Escherichia coli K12

The biodegradative threonine dehydratase gene ( tdc ) of Escherichia coli was cloned by isolating a dehydratase-negative mutant after Tn5 mutagenesis, cloning the tdc ::Tn5 DNA into pBR322 and then replacing the Tn5 element on the plasmid in vivo. Subcloning and nucleotide sequence data revealed two distinct procaryotic promoterlike elements each containing a potential CAP-binding site and AT-rich regions, and a Shine-Dalgarno sequence. One of these putative promoters, P 2 , was located immediately upstream from the tdc coding region, and a second, P 1 , was approximately 1 kilobase upstream from P 2 . Deletion of the potential CAP-binding site from P 1 prevented tdc gene expression. However, removal of P 2 and a large segment of the upstream DNA had no discernible effect on dehydratase synthesis. A 936-base pair open reading frame was found between P 1 and the tdc coding region, which produced a polypeptide of about 32 kilodaltons. The data suggest that P 1 , and not P 2 , is necessary for tdc gene expression, and that the DNA sequences coding for the 32 KD polypeptide and threonine dehydratase are part of a single transcriptional unit.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/47562/1/438_2004_Article_BF00425676.pd