Diagnosis and treatment of refractory infectious diseases using nanopore sequencing technology:Three case reports

BACKGROUND Infectious diseases are still one of the greatest threats to human health,and the etiology of 20%of cases of clinical fever is unknown;therefore,rapid identification of pathogens is highly important.Traditional culture methods are only able to detect a limited number of pathogens and are time-consuming;serologic detection has window periods,false-positive and false-negative problems;and nucleic acid molecular detection methods can detect several known pathogens only once.Three-generation nanopore sequencing technology provides new options for identifying pathogens.CASE SUMMARY Case 1:The patient was admitted to the hospital with abdominal pain for three days and cessation of defecation for five days,accompanied by cough and sputum.Nanopore sequencing of the drainage fluid revealed the presence of orallike bacteria,leading to a clinical diagnosis of bronchopleural fistula.Cefoperazone sodium sulbactam treatment was effective.Case 2:The patient was admitted to the hospital with fever and headache,and CT revealed lung inflammation.Antibiotic treatment for Streptococcus pneumoniae,identified through nanopore sequencing of cerebrospinal fluid,was effective.Case 3:The patient was admitted to our hospital with intermittent fever and an enlarged neck mass that had persisted for more than six months.Despite antibacterial treatment,her symptoms worsened.The nanopore sequencing results indicate that voriconazole treatment is effective for Aspergillus brookii.The patient was diagnosed with mixed cell type classical Hodgkin's lymphoma with infection.CONCLUSION Three-generation nanopore sequencing technology allows for rapid and accurate detection of pathogens in human infectious diseases.

Application of Biological Nanopore Sequencing Technology in the Detection of Microorganisms

Environmental pollution and the spread of pathogenic microorganisms pose a significant threat to the health of humans and the planet.Thus,understanding and detecting microorganisms is crucial for maintaining a healthy living environment.Nanopore sequencing is a single-molecule detection method developed in the 1990s that has revolutionized various research fields.It offers several advantages over traditional sequencing methods,including low cost,label-free,time-saving detection speed,long sequencing reading,real-time monitoring,convenient carrying,and other significant advantages.In this review,we summarize the technical principles and characteristics of nanopore sequencing and discuss its applications in amplicon sequencing,metagenome sequencing,and whole-genome sequencing of environmental microorganisms,as well as its in situ application under some special circumstances.We also analyze the advantages and challenges of nanopore sequencing in microbiology research.Overall,nanopore sequencing has the potential to greatly enhance the detection and understanding of microorganisms in environmental research,but further developments are needed to overcome the current challenges.

Polymer translocation through nanopore under external electric field:dissipative particle dynamics study

The DNA sequencing technology has achieved a leapfrog development in recent years. As a new generation of the DNA sequencing technology, nanopore sequenc- ing has shown a broad application prospect and attracted vast research interests since it was proposed. In the present study, the dynamics of the electric-driven translocation of a homopolymer through a nanopore is investigated by the dissipative particle dynam- ics （DPD）, in which the homopolymer is modeled as a worm-like chain （WLC）. The DPD simulations show that the polymer chain undergoes conformation changes during the translocation process. The different structures of the polymer in the translocation process, i.e., single-file, double folded, and partially folded, and the induced current block- ades are analyzed. It is found that the current blockades have different magnitudes due to the polymer molecules traversing the pore with different folding conformations. The nanoscale vortices caused by the concentration polarization layers （CPLs） in the vicinity of the sheet are also studied. The results indicate that the translocation of the polymer has the effect of eliminating the vortices in the polyelectrolyte solution. These findings are expected to provide the theoretical guide for improving the nanopore sequencing tech- nique.

Transcriptomic Analysis and Comparison of the Gene Expression Profiles in Fast- and Slow-Growing Pearl Oysters Pinctada fucata martensii

The pearl oyster Pinctada fucata martensii is an economically valuable shellfish that is cultured for seawater pearl pro-duction,which mainly depends on oyster growth.However,the growth mechanisms of the pearl oyster are still poorly understood.In this study,oysters were grouped with relative growth rate,including fast-growing(FG)group and slow-growing(SG)group.Oxford Nanopore Technologies(ONT)long-read sequencing was applied to investigate the molecular mechanisms involved in the growth of this species.Five alternative splicing(AS)types were analyzed in both FG and SG groups,which include alternative 3’splice site,alternative 5’splice site,exon skipping,intron retention,and mutually exclusive exon.Transcriptome analysis showed that four of five different AS events(excluding mutually exclusive exons)occurred more frequently in FG than in SG oysters,and the five main AS types exhibited different characteristics.The AS events that were detected may be involved in growth,and the difference in ex-pression of AS events between FG and SG oysters may be involved in the mechanism underlying the difference in growth.Fifty dif-ferentially expressed genes(DEGs)were identified between the FG and SG oysters.The results showed that 40 genes were signifi-cantly up-regulated in FG oysters,while 10 genes were significantly down-regulated in SG oyster.Several genes related to nutrient metabolism,shell formation,and immunity were more highly expressed in FG oysters than in SG oysters.In summary,FG oysters exhibited higher metabolic and biomineralization activities and had a more powerful immune system than SG oysters.These results provide insight into the growth of P.f.martensii that can be used to improve breeding programs.