Molecular Biology Reports最新文献

Background: Next generation sequencing (NGS) remains underutilized in clinical microbiology applications despite providing broad pathogen spectrum detection superior to other molecular methods. This is primarily because of lower sensitivity of metagenomic NGS (mNGS) compared to PCR, lengthy turn-around times, cost, and complexity of data analysis. Capture sequencing is a technique that can mitigate some of the limitations of mNGS. Using probes that are engineered to selectively bind and pull down desired nucleic acids, capture sequencing enriches for targets of interest and can result in up to a 10,000-fold increase in sensitivity compared to mNGS. In this study, we describe the application of capture sequencing on Oxford Nanopore Technology's portable sequencer, the MinION MK1C.

Methods: We examined the performance of VirCapSeq-VERT and TBDCapSeq, two distinct capture sequencing assays that target vertebrate viruses and tick-borne pathogens, respectively. Both assays were originally established on the Illumina platform. To enable sequencing on the MinION instrument, we developed a modified hybrid workflow using our established library preparation and capture protocol for Illumina, followed by the addition of the ONT sequencing adaptor. In tests using contrived and clinical samples, we compared sensitivity thresholds and sequencing output, including pathogen genome coverage and relevant read counts.

Results: The addition of capture enrichment to MinION NGS provided significant improvement in pathogen detection when compared to mNGS. Assessment of assay performance on pathogen-positive samples revealed equivalent sensitivity on the MinION MK1C and Illumina NextSeq. We found that the elevated read counts and sequencing depth generated by Illumina NGS were offset by the greater read length obtained on the MinION MK1C and resulted in comparable pathogen genome coverage between the two platforms.

Conclusion: This study demonstrates the utility for employment of VirCapSeq and TBDCapSeq on different sequencing platforms and suggest the potential of the MinION platform for broad-spectrum clinical diagnostics.

Background: Geniotrigona thoracica is one of the commonest stingless bee species for pollination services and commercial meliponiculture in Thailand, Malaysia, and Indonesia.

Methods and results: Its complete mitochondrial genome exhibits genome duplication, heteroplasmy and gene rearrangement. It comprises a 16,045-bp "canonical" mitogenome; and a 32,092-bp genome comprising two segments, the 16,045-bp "canonical" genome and its "duplicated-rearranged" genome. The "canonical" genome contains 39 genes-13 protein-coding genes (PCGs), 2 rRNA genes, and 24 tRNA genes (trnK and trnM are duplicated). All the 13 PCGs as well as the two rRNA genes and 19 tRNA genes are located on the majority (J) strand. The "duplicated-rearranged" genome comprises a short segment of tRNA genes in close proximity to the control region (trnM-trnK-trnI-trnA-trnK-trnM-control region) and a long inverted segment of 33 genes (13 PCGs, 2 rRNAs and 18 tRNAs of the nad2 to trnS2 segment of the "canonical" genome). Based on 15 mt-genes and 13 PCGs, G. thoracica forms a subclade with the lineage consisting of Heterotrigona and Lepidotrigona.

Conclusions: This phylogenetic relationship concurs with earlier findings based on over 2500 ultra-conserved element (UCE) loci as well as mitochondrial and nuclear genes, indicating mitogenome is suitable for taxon differentiation and phylogenetic study. In addition, the geographic isolates of G. thoracica from Malaysia and Thailand are genetically variable.

Bacterial outer membrane vesicles (OMVs) were previously considered merely as waste products of bacterial metabolism. But its inherent stability, immunogenicity, ability to be highly regulated, and targeting potential are turning them into a platform with revolutionary potential in tumor drug delivery. Tumor remains a major challenge in medicine. Even though traditional therapies are effective to some extent, they are often accompanied by intolerable side effects. Furthermore, conventional treatments have low targeting efficiency, and tumor cells almost always develop resistance to them. OMVs are naturally tiny vesicles released by Gram-negative bacteria. They possess the ability to deliver drugs very efficiently and at the same time stimulate the host immune system. As a result, they can overcome limitations such as poor targeting and drug resistance. OMVs have huge potential not only in tumor therapy but also in the development of vaccines and drug delivery systems. Nevertheless, reviews focusing solely on their role in tumor drug delivery systems are still quite rare. This review thoroughly analyzes the immunological basis of bacterial OMVs, the engineering strategies, and their antitumor applications with a focus on how their innate immunogenicity and highly customizable features can be exploited to develop precision antitumor platforms. It essentially offers a comprehensive and systematic theoretical framework for realizing and elevating the latent use of bacterial OMVs in tumor therapy.

Human immunodeficiency virus (HIV) remains a persistent global health burden, as combination antiretroviral therapy (ART) achieves sustained viral suppression but fails to eliminate long-lived latent reservoirs. Stem cell-based therapeutic strategies have emerged as transformative approaches with the potential to induce durable remission and, ultimately, a functional cure. Clinical proof-of-concept has been established through allogeneic hematopoietic stem cell transplantation (HSCT) using CCR5Δ32/Δ32 donor cells, demonstrating that durable resistance to viral entry can result in prolonged HIV remission. Building on these landmark observations, recent advances in autologous gene-edited hematopoietic stem and progenitor cells and induced pluripotent stem cell (iPSC)-derived immune effectors have accelerated the development of scalable, patient-specific interventions. The convergence of stem cell biology with precision genome-editing platforms, including CRISPR-Cas9, transcription activator-like effector nucleases (TALENs), and zinc finger nucleases (ZFNs), has enabled targeted disruption of viral entry pathways and host dependency factors, while offering new strategies to address viral latency and immune reconstitution. Despite significant challenges related to treatment-associated toxicity, manufacturing complexity, long-term safety, and ethical considerations, rapid progress in cellular engineering and translational immunology continues to advance the field toward curative outcomes. This review critically synthesizes recent progress in stem cell-based HIV therapeutics, elucidates the underlying mechanistic frameworks, evaluates emerging clinical and preclinical evidence, and outlines future directions required to achieve a durable functional cure.

Background: The IGF-1 (insulin-like growth factor) and CAST (calpastatin) genes have been reported to influence several economically important productivity traits in sheep. This study aimed to investigate the relationship between the genotypes of these two genes, identified by PCR-RFLP (Polymerase Chain Reaction-Restriction Fragment Length Polymorphism) analysis, and growth-related characteristics in Anatolian Merino and Akkaraman sheep.

Methods and results: In addition to live weight (LW), morphometric measurements including rump height (RH), withers height (WH), body length (BL), chest depth (CD), chest width (CW), and chest circumference (CC) were recorded. IGF-1 genotypes were identified using HaeIII digestion, revealing three genotypes (CC, CG, and GG), while CAST genotypes were determined by MspI digestion, yielding two genotypes (MM and MN). Associations between genotypes and traits were evaluated through analysis of variance and CHAID regression tree analysis, and breed comparisons were performed using t-tests. Anatolian Merino sheep exhibited higher LW, BL, CW, and CC, whereas Akkaraman sheep showed greater RH and WH. No significant association between IGF-1 genotypes and morphometric traits was observed in Anatolian Merino sheep; however, in Akkaraman sheep, the IGF-1 GG genotype was significantly associated with higher LW, RH, WH, CW, and CC. CAST genotypes were not related to any of the examined traits, and regression tree analysis supported these results.

Conclusions: Overall, the results suggest that IGF-1 genotypes, particularly GG, could be valuable for marker-assisted selection (MAS) programmes targeting live weight and body measurements in sheep.