Development and validation of novel PCR primers for identification of plasmid-mediated colistin resistance (mcr) genes in various environmental settings

Antibiotic resistance is considered one of the biggest threats to public health and has become a major concern for governments and international organizations. Combating this problem starts with improving global surveillance of antibiotic resistance genes (ARGs) and applying standardized protocols, both in a clinical and environmental context, in agreement with the One Health approach. Exceptional efforts should be directed to controlling ARGs conferring resistance to Critically Important Antimicrobials (CIA). In this study, a systematic literature review to synthesize data on the identification of mcr genes using a PCR technique was performed. Additionally, a novel set of PCR primers for mcr-1 – mcr-9 genes detection was proposed. The developed primers were in silico and experimentally validated by comparison with mcr-specific PCR primers reported in the literature. This validation, besides being a proof-of-concept for primers’ usefulness, provided insight into the distribution of mcr genes in municipal wastewater, clay and river sediments, glacier moraine, manure, seagulls and auks feces and daphnids from four countries. This analysis proved that commonly used primers may deliver false results, and some mcr genes may be overlooked in tested samples. Newly-developed PCR primers turned out to be relevant for the screening of mcr genes in various environments.info:eu-repo/semantics/publishedVersio

5-epi-Incensole: synthesis, X-ray crystal structure and absolute configuration by means of ECD and VCD studies in solution and solid state

Incensole 1 and its acetate 2, found in incense, demonstrate interesting biological activities. Incensole acetate 2 was prepared on a large scale by employing the Paul and Jauch protocol from the crude extracts of Boswellia papyrifera Hochst. 5-epi-Incensole 3, obtained as colorless crystals, was prepared from incensole acetate via three steps; deacetylation, oxidation and reduction. The structure of 5-epi-incensole 3 was elucidated by means of spectroscopic data analysis, and the absolute configuration was established by single crystal X-ray analysis in combination with electronic and vibrational circular dichroism. In particular, the applicability of the solid-state ECD/TDDFT protocol to a compound with only two non-conjugated alkene chromophores was verified

Genetic Profile and Clinical Implications of Hepatoblastoma and Neuroblastoma Coexistence in a Child

The aim of the following case report is to provide a description of the coexistence of two independent tumors in a child. A 9-month-old male was referred to Department of Pediatric Oncology and Hematology with hepatic tumor present on ultrasound imaging and symptoms of enlarged abdominal circumference. Physical examination revealed a palpable epigastric mass and the imaging techniques showed a tumor of the left hepatic lobe measuring 11 × 6.5 × 8.9 cm with pancreas infiltration, distant metastases in both lungs and abnormal lesion in the left adrenal gland. Basing on histopathological examination, after a core-needle biopsy, hepatoblastoma (HBL) (mixed epithelial-mesenchymal subtype) was diagnosed. The α-fetoprotein level was 112 993 ng/ml. Elevated values of normetanephrine, 3-methoxytyramine as well as neuron-specific enolase were observed. Due to the clinical picture and diagnosis, the patient was qualified to preoperative chemotherapy according to the SIOPEL-3 protocol, followed by SIOPEL-4 protocol for the high-risk patients. After undergoing preoperative chemotherapy, imaging tests revealed regression of hepatic tumor and no focal pulmonary masses, while regression of adrenal gland mass was not completed. The patient was qualified for left hemihepatectomy with left adrenalectomy. Histopathological examination of liver specimen confirmed the HBL diagnosis. However, in left adrenal gland and paraaortic lymph nodes the residual neuroblastoma (NBL) cells were detected. Whole exome sequencing (WES) was utilized to identify disease-associated germline mutations. WES revealed a novel germline insertion variant in TWIST1 (p.Gly86dup), along with the potentially pathogenic non-synonymous variants in NF1 (p.Val2511Ile), RAF1 (p.Leu445Arg), and WHSC1 (p.Ser4Asn) genes. Currently, 6 months after completion of treatment according to the SIOPEL-4 protocol, the patient is in good general condition, without any signs, and symptoms of relapse of both neoplasms. The coexistence of two different primary childhood malignancies is rarely seen. So far, only one case of synchronous HBL and NBL has been reported. However, for the first time therapeutic process was successful. A specific signature of rare germline mutations can be proposed as a predisposing factor to synchronous HBL and NBL occurrence

Complementarity of Electronic and Vibrational Circular Dichroism Spectroscopy in Structure Determination of Vic-Amino Alcohols

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Induced circularly polarized luminescence for revealing DNA binding with fluorescent dyes

To the best of our knowledge this is the first report on the application of induced circularly polarized luminescence (CPL) for sensing the binding of fluorescent dyes to double stranded DNA. Using Thiazole Orange (TO) and 4',6-diamidino-2-phenylindole (DAPI) as models, we show utility and limitations of CPL in DNA binding studies. The results obtained indicate that CPL can be used as a new chiroptical tool for this purpose, however, special attention while recording CPL data must be used, in order to exclude measurement artefacts caused by linear polarization components

Circular Dichroism Imaging: Mapping the Local Supramolecular Order in Thin Films of Chiral Functional Polymers

We introduce the concept of circular dichroism (CD) imaging CDi, which is made possible by the use of the highly collimated light beam of synchrotron radiation (SR) only available at Diamond Light Source B23 beamline for surface scanning of solid samples. At the moment, the space resolution is about 1 mm2, but there are prospects to increase it to at least 0.1 mm2 in the near future. We apply SR-CDi to thin films of conjugated polymers, which give rise to large chiral aggregates. Electronic CD is able to reveal and characterize these supramolecular structures; thus, SR-CDi provides the ultimate complement to microscopy for investigating local domains with different molecular order and to fill a gap between a purely morphological knowledge and an intimate insight into the molecular level. As a corollary of this proof-of-concept work, we showed that such a new technique may provide rational and successful protocols to obtain homogeneous active layers for the fabrication of optoelectronic devices

Phytotoxic Metabolites Produced by Diaporthella cryptica, the Causal Agent of Hazelnut Branch Canker

From the culture filtrates of Diaporthella cryptica, an emerging hazelnut pathogen, 2-hydroxy-3-phenylpropanoate methyl ester and its 3-(4-hydroxyphenyl) and 3-(1H-indol-3-yl) analogues, named crypticins A-C, were isolated together with the well-known tyrosol. Crypticins A-C were identified by spectroscopic (essentially nuclear magnetic resonance and high-resolution electrospray ionization mass spectrometry) methods. The R absolute configuration (AC) of crypticin A was determined by comparing its optical rotation and electronic circular dichroism (ECD) spectrum with those of papuline, the methyl ester of (-)(S)-phenyllactic acid isolated as the main phytotoxin of Pseudomonas syringae pv. papulans, responsible for apple blister spot. The ACs of crypticins B and C were determined by time-dependent density functional theory calculations of their ECD spectra. Papuline and the new metabolites herein isolated, except tyrosol, were tested at 1 mg/mL on cork oak, grapevine, hazelnut, and holm oak leaves using the leaf puncture assay. They were also tested on tomato cuttings at 0.5 and 0.05 mg/mL. In the leaf puncture assay, none of the compounds was found to be active. Crypticin C and papuline were active in the tomato cutting assay. Additionally, crypticin C displayed moderate inhibitory effect against Phytophthora cambivora. © 2018 American Chemical Society

Pyriculins A and B, two monosubstituted hex-4-ene-2,3-diols and other phytotoxic metabolites produced by Pyricularia grisea isolated from buffelgrass (Cenchrus ciliaris)

Pyricularia grisea has been identified as a foliar pathogen on buffelgrass (Cenchrus ciliaris) in North America and was studied as a potential source of phytotoxins for buffelgrass control. Two monosubstituted hex-4-ene-2,3-diols, named pyriculins A and B, were isolated from its culture filtrate organic extract together with (10S,11S)-(â\u88\u92)-epipyriculol, trans-3,4-dihydro-3,4,8-trihydroxy-1(2H)-napthalenone, and (4S)-(+)-isosclerone. Pyriculins A and B were characterized by spectroscopic (essentially nuclear magnetic resonance [NMR], High-resolution electrospray ionization mass spectrometry [HRESIMS]) and chemical methods such as (4E)-1-(4-hydroxy-1,3-dihydroisobenzofuran-1-yl)hex-4-ene-2,3-diols. The relative and absolute configuration of these compounds was determined by a combination of spectroscopic (NMR, electronic circular dichroism [ECD]) and computational tools. When bioassayed in a buffelgrass coleoptile and radicle elongation test, (10S,11S)-(â\u88\u92)-epipyriculol proved to be the most toxic compound. Seed germination was much reduced and slowed with respect to the control and radicles failed to elongate. All five compounds delayed germination, but only (10S,11S)-(â\u88\u92)-epipyriculol was able to prevent radicle development of buffelgrass seedlings. It had no effect on coleoptile elongation, while the other four compounds caused significantly increased coleoptile development relative to the control

Phytotoxic Metabolites Produced by <i>Diaporthella cryptica</i>, the Causal Agent of Hazelnut Branch Canker

From the culture filtrates of <i>Diaporthella cryptica</i>, an emerging hazelnut pathogen, 2-hydroxy-3-phenylpropanoate methyl ester and its 3-(4-hydroxyphenyl) and 3-(1<i>H</i>-indol-3-yl) analogues, named crypticins A–C, were isolated together with the well-known tyrosol. Crypticins A–C were identified by spectroscopic (essentially nuclear magnetic resonance and high-resolution electrospray ionization mass spectrometry) methods. The <i>R</i> absolute configuration (AC) of crypticin A was determined by comparing its optical rotation and electronic circular dichroism (ECD) spectrum with those of papuline, the methyl ester of (−)­(<i>S</i>)-phenyllactic acid isolated as the main phytotoxin of <i>Pseudomonas syringae</i> pv. <i>papulans</i>, responsible for apple blister spot. The ACs of crypticins B and C were determined by time-dependent density functional theory calculations of their ECD spectra. Papuline and the new metabolites herein isolated, except tyrosol, were tested at 1 mg/mL on cork oak, grapevine, hazelnut, and holm oak leaves using the leaf puncture assay. They were also tested on tomato cuttings at 0.5 and 0.05 mg/mL. In the leaf puncture assay, none of the compounds was found to be active. Crypticin C and papuline were active in the tomato cutting assay. Additionally, crypticin C displayed moderate inhibitory effect against <i>Phytophthora cambivora</i>