High-Throughput RNA Extraction from Citrus Tissues for the Detection of Viroids.

High-throughput nucleic acid extraction is critical for the implementation of modern viroid detection assays. Successful large-scale nursery, field surveys, and other regulatory, quarantine, or research diagnostic programs are increasingly dependent on high-throughput tissue pulverization and nucleic acid extraction protocols. Magnetic bead-based approaches using semi-automated robotic equipment allow high-throughput extraction and purification of high-quality uniform total nucleic acids for each individual sample. Here, we describe a streamlined and optimized protocol for citrus tissue processing and RNA extraction that can be used for downstream applications such as viroid detection by reverse transcription-quantitative polymerase chain reaction.

Plasmonic Gold Nanoisland Film for Bacterial Theranostics.

Plasmonic nanomaterials have been intensively explored for applications in biomedical detection and therapy for human sustainability. Herein, plasmonic gold nanoisland (NI) film (AuNIF) was fabricated onto a glass substrate by a facile seed-mediated growth approach. The structure of the tortuous gold NIs of the AuNIF was demonstrated by scanning electron microscopy and energy-dispersive X-ray spectroscopy. Based on the ultraviolet-visible spectrum, the AuNIF revealed plasmonic absorption with maximum intensity at 624 nm. With the change to the surface topography created by the NIs, the capture efficiency of Escherichia coli (E. coli) by the AuNIF was significantly increased compared to that of the glass substrate. The AuNIF was applied as a surface-enhanced Raman scattering (SERS) substrate to enhance the Raman signal of E. coli. Moreover, the plasmonic AuNIF exhibited a superior photothermal effect under irradiation with simulated AM1.5 sunlight. For photothermal therapy, the AuNIF also displayed outstanding efficiency in the photothermal killing of E. coli. Using a combination of SERS detection and photothermal therapy, the AuNIF could be a promising platform for bacterial theranostics.

Hemoglobin-Conjugated Gold Nanoclusters for Qualitative Analysis of Haptoglobin Phenotypes.

Designing a facile and rapid detection method for haptoglobin (Hp) phenotypes in human blood plasma is urgently needed to meet clinic requirements in hemolysis theranostics. In this work, a novel approach to qualitatively analyze Hp phenotypes was developed using a fluorescent probe of gold nanoclusters (AuNCs). Hemoglobin-conjugated (Hb)-AuNCs were successfully synthesized with blue-green fluorescence and high biocompatibility via one-pot synthesis. The fluorescence of Hb-AuNCs comes from the ligand-metal charge transfer between surface ligands of Hb and the gold cores with high oxidation states. The biocompatibility assays including cell viability and fluorescence imaging, demonstrated high biocompatibility of Hb-AuNCs. For the qualitative analysis, three Hp phenotypes in plasma, Hp 1-1, Hp 2-1, and Hp 2-2, were successfully discriminated according to changes in the fluorescence intensity and peak position of the maximum intensity of Hb-AuNCs. Our work provides a practical method with facile and rapid properties for the qualitative analysis of three Hp phenotypes in human blood plasma.

Full genome characterization of 12 citrus tatter leaf virus isolates for the development of a detection assay.

Citrus tatter leaf virus (CTLV) threatens citrus production worldwide because it induces bud-union crease on the commercially important Citrange (Poncirus trifoliata × Citrus sinensis) rootstocks. However, little is known about its genomic diversity and how such diversity may influence virus detection. In this study, full-length genome sequences of 12 CTLV isolates from different geographical areas, intercepted and maintained for the past 60 years at the Citrus Clonal Protection Program (CCPP), University of California, Riverside, were characterized using next generation sequencing. Genome structure and sequence for all CTLV isolates were similar to Apple stem grooving virus (ASGV), the type species of Capillovirus genus of the Betaflexiviridae family. Phylogenetic analysis highlighted CTLV's point of origin in Asia, the virus spillover to different plant species and the bottleneck event of its introduction in the United States of America (USA). A reverse transcription quantitative polymerase chain reaction assay was designed at the most conserved genome area between the coat protein and the 3'-untranslated region (UTR), as identified by the full genome analysis. The assay was validated with different parameters (e.g. specificity, sensitivity, transferability and robustness) using multiple CTLV isolates from various citrus growing regions and it was compared with other published assays. This study proposes that in the era of powerful affordable sequencing platforms the presented approach of systematic full-genome sequence analysis of multiple virus isolates, and not only a small genome area of a small number of isolates, becomes a guideline for the design and validation of molecular virus detection assays, especially for use in high value germplasm programs.

Antimicrobial Gold Nanoclusters: Recent Developments and Future Perspectives.

Bacterial infections have caused serious threats to public health due to the antimicrobial resistance in bacteria. Recently, gold nanoclusters (AuNCs) have been extensively investigated for biomedical applications because of their superior structural and optical properties. Great efforts have demonstrated that AuNCs conjugated with various surface ligands are promising antimicrobial agents owing to their high biocompatibility, polyvalent effect, easy modification and photothermal stability. In this review, we have highlighted the recent achievements for the utilizations of AuNCs as the antimicrobial agents. We have classified the antimicrobial AuNCs by their surface ligands including small molecules (< 900 Daltons) and macromolecules (> 900 Daltons). Moreover, the antimicrobial activities and mechanisms of AuNCs have been introduced into two main categories of small molecules and macromolecules, respectively. In accordance with the advancements of antimicrobial AuNCs, we further provided conclusions of current challenges and recommendations of future perspectives of antimicrobial AuNCs for fundamental researches and clinical applications.

Complete nucleotide sequence and genome organization of an endornavirus from bottle gourd (Lagenaria siceraria) in California, U.S.A.

The full-length nucleotide sequence and genome organization of an Endornavirus isolated from ornamental hard shell bottle gourd plants (Lagenaria siceraria (Molina) Standl.) in California (CA), USA tentatively named L. siceraria endornavirus-California (LsEV-CA) was determined. The LsEV-CA genome was 15088 bp in length, with a G + C content of 36.55 %. The lengths of the 5' and 3' untranslated regions were 111 and 52 bp, respectively. The genome of LsEV-CA contained one large ORF encoding a 576 kDa polyprotein. The predicted protein contains two glycosyltransferase motifs, as well as RNA-dependent RNA polymerase and helicase domains. LsEV-CA was detected in healthy-looking field-grown gourd plants, as well as plants expressing yellows symptoms. It was also detected in non-symptomatic greenhouse-grown gourd seedlings grown from seed obtained from the same field sites. These preliminary data indicate that LsEV-CA is likely not associated with the gourd-yellows syndrome observed in the field.