Construction of an efficient genomic editing system with CRISPR/Cas9 in the vector mosquito Aedes albopictus

Aedes (Stegomyia) albopictus, also known as the Asian tiger mosquito, is a mosquito which originated in Asia. In recent years, it has become increasingly rampant throughout the world. This mosquito can transmit several arboviruses, including dengue, Zika and chikungunya viruses, and is considered a public health threat. Despite the urgent need of genome engineering to analyze specific gene functions, progress in genetical manipulation of Ae. albopictus has been slow due to a lack of efficient methods and genetic markers. In the present study, we established targeted disruptions in two genes, kynurenine hydroxylase (kh) and dopachrome conversion enzyme (yellow), to analyze the feasibility of generating visible phenotypes with genome editing by the clustered regularly interspaced short palindromic repeats (CRISPR)/ CRISPR-associated protein 9 (Cas9) system in Ae. albopictus. Following Cas9 single guide RNA ribonucleoprotein injection into the posterior end of pre-blastoderm embryos, 30%-50% of fertile survivors produced alleles that failed to complement existing kh and yellow mutations. Complete eye and body pigmentation defects were readily observed in GI pupae and adults, indicating successful generation of highly heritable mutations. We conclude that the CRISPR/Cas9-mediated gene editing system can be used mAe. albopictus and that it can be adopted as an efficient tool for genome-scale analysis and biological study.     

In silico detection of tRNA sequence features characteristic to aminoacyl-tRNA synthetase class membership

Aminoacyl tRNA synthetases (aaRS) are grouped into Class I and II based on primary and tertiary structure and enzyme properties suggesting two independent phylogenetic lineages. Analogously, tRNA molecules can also form two respective classes, based on the class membership of their corresponding aaRS. Although some aaRS-tRNA interactions are not extremely specific and require editing mechanisms to avoid misaminoacylation, most aaRS-tRNA interactions are rather stereospecific. Thus, class-specific aaRS features could be mirrored by class-specific tRNA features. However, previous investigations failed to detect conserved class-specific nucleotides. Here we introduce a discrete mathematical approach that evaluates not only class-specific 'strictly present', but also 'strictly absent' nucleotides. The disjoint subsets of these elements compose a unique partition, named extended consensus partition (ECP). By analyzing the ECP for both Class I and II tDNA sets from 50 (13 archaeal, 30 bacterial and 7 eukaryotic) species, we could demonstrate that class-specific tRNA sequence features do exist, although not in terms of strictly conserved nucleotides as it had previously been anticipated. This finding demonstrates that important information was hidden in tRNA sequences inaccessible for traditional statistical methods. The ECP analysis might contribute to the understanding of tRNA evolution and could enrich the sequence analysis tool repertoire.     

Isolation and Characterization of a Carbendazim-Degrading Rhodococcus sp. djl-6

Bacterium djl-6, capable of degrading carbendazim, was isolated by continuous enrichment culture originating from carbendazim-treated soil. The isolate was identified as Rhodococcus sp. according to its phenotypic features, physiologic and biochemical characteristics, and phylogenetic analysis. The strain could use carbendazim as sole carbon or nitrogen source. It showed a high average degradation rate of 55.56 mg · L⁻¹ · d⁻¹ in M9 medium amended with carbendazim. High-pressure liquid chromatography–mass spectrometry (HPLC-MS) analysis showed the presence of 2-aminobenzimidazole, benzimidazole, and an unknown metabolite with molecular ions (M⁺) of m/z 104.8 and 118.5. The degradation in the isolate djl-6 seems to be initiated with the cleavage of the methyl carbemate side chain, resulting in the formation of 2-aminobenzimidazole and benzimidazole. This is the first report of the intermediates benzimidazole and 2-aminobenzimidazole found together in the culture filtrate of pure bacterium.     

Vertebral artery dissection associated with viral meningitis

ABSTRACT: BACKGROUND: Vertebral artery dissection (VAD) is often associated with trauma or occurs spontaneously, inevitably causing some neurological deficits. Even though acute infection can be related to the development of spontaneous VAD (sVAD), VAD associated with viral meningitis has never been reported in the literature. CASE PRESENTATION: A 42-year-old man with fever, sore throat, and runny nose developed sudden onset of occipital headache, vertigo, transient confusion, diplopia, and ataxia. Brain stem encephalitis was diagnosed initially because the cerebrospinal fluid (CSF) study showed inflammatory changes. However, subsequent diffusion-weighted (DWI) magnetic resonance imaging of his brain demonstrated left lateral medullary infarction, and the digital subtraction angiography (DSA) confirmed VAD involving left V4 segment of the artery. Consequently, the patient was diagnosed as VAD accompanied by viral meningitis. CONCLUSION: This case suggests that viral meningitis might lead to inflammatory injury of the vertebral arterial wall, even sVAD with multiple neurological symptoms.     

Identification of differentially expressed proteins in poplar leaves induced by Marssonina brunnea f. Sp. Multigermtubi

Black spot disease in poplar is a disease of the leaf caused by fungus. The major pathogen is Marssonina brunnea f. sp. multigermtubi. To date, little is known about the molecular mechanism of poplar （M. brunnea） interaction. In order to identify the proteins related to disease resistance and understand its molecular basis, the clone ＂NL895＂ （P. euramericana CL＂NL895＂）, which is highly resistant to M. brunnea f. sp. multigermtubi, was used in this study. We used two-dimensional gel electrophoresis （2-DE） and mass spectrometry （MS） to identify the proteins in poplar leaves that were differentially expressed in response to black spot disease pathogen, M. brunnea f. sp. multigermtubi. Proteins extracted from poplar leaves at 0, 12, 24, 48, and 72 h after pathogen-inoculation were separated by 2-DE, About 500 reproducible protein spots were detected, of which 40 protein spots displayed differential expression in levels and were subjected to Matrix assisted laser desorption/ionization time of flight mass spectrometry （MALDI-TOF MS） followed by database searching. According to the function, the identified proteins were sorted into five categories, that is, protein synthesis, metabolism, defense response and unclassified proteins.     

Dependence of trans-ADP-ribosylation and nuclear glycolysis on the Arg 34-ATP complex of Zn2+ finger I of poly-ADP-ribose polymerase-1

The H-bonded complex of ATP with Arg 34 of Zn2+ finger I of poly-ADP-ribose polymerase-1 (PARP-1) determines trans-oligo-ADP-ribosylation from NAD+ to proteins other than PARP-1. This mechanism was tested in lysolecithin fractions of non-malignant and cancer cells separately and after their recombination. Cellular PARP-1 activity was recovered when the centrifugal sediment was recombined with the supernatant fraction containing cellular ADP-ribose oligomer acceptor proteins. Combination of the matrix fraction (Mx) of cancer cells (lacking OXPHOS) with its supernatant had the same PARP-1 activity as the Mx alone. The supernatant of non-malignant cells was replaced by glycolytic enzymes as ADP-ribose acceptor. The hexokinase activity of the supernatant increased when OXPHOS of intact cells was uncoupled by carbonyl cyanide 4-(trifluoro methoxy) phenylhydrazone. trans-ADP-ribosylation was demonstrated by polyacrylamide gel electrophoresis.     

Screening and identification of Shigella flexneri 2a virulence-related genes induced after invasion of epithelial cells

~~Screening and identification of Shigella flexneri 2a virulence-related genes induced after invasion of epithelial cells1. Jin, Q., Yuan, Z., Xu, J., Wang, Y., Shen, Y., Lu, W., Wang, J., Liu, H., Yang, J., Yang, P., Zhang, X., Zhang, J., Yang, G, Wu, H., Qu, D., Dong, J., Sun, L., Xue, Y, Zhao, A., Gao, Y., Zhu, J., Kan, B., Ding, K.. Chen, S., Cheng, H., Yao, Z., He, B., Chen, R., Ma, D., Qiang, B., Wen, Y, Hou, Y., Yu, J., Genome sequence of Shigella flexneri 2…     

Monoquaternary ammonium derivatives inhibit growth of protozoan parasites

The phospholipid metabolism of Plasmodium falciparum-infected erythrocytes has been shown to be an effective pharmacological target for novel chemotherapy. Thirty-seven monoquaternary ammonium derivatives analogous to choline were screened for their potential antiprotozoal activity against P. falciparum and Leishmania braziliensis. Twenty-three compounds inhibited chloroquine resistant and sensitive P. falciparum strains with inhibitory concentrations ranging from 0.001 μM to 47 μM. Among the inhibitors were six compounds with nanomolar activity containing at least one ethyl group in the polar head and a hydrophobic alkyl chain with 10 to 14 methylene groups. Four compounds also exhibited in vitro antileishmanial properties in the micromolar range.