Impact of next-generation sequencing on antimicrobial treatment in immunocompromised adults with suspected infections

BACKGROUND:Prompt pathogen identification can have a substantial impact on the optimization of antimicrobial treatment.The objective of the study was to assess the diagnostic value of next-generation sequencing(NGS)for identifying pathogen and its clinical impact on antimicrobial intervention in immunocompromised patients with suspected infections.METHODS:This was a retrospective study.Between January and August 2020,47 adult immunocompromised patients underwent NGS testing under the following clinical conditions:1)prolonged fever and negative conventional cultures;2)new-onset fever despite empiric antimicrobial treatment;and 3)afebrile with suspected infections on imaging.Clinical data,including conventional microbial test results and antimicrobial treatment before and after NGS,were collected.Data were analyzed according to documented changes in antimicrobial treatment(escalated,no change,or deescalated)after the NGS results.RESULTS:The median time from hospitalization to NGS sampling was 19 d.Clinically relevant pathogens were detected via NGS in 61.7% of patients(29/47),more than half of whom suffered from fungemia(n=17),resulting in an antimicrobial escalation in 53.2% of patients(25/47)and antimicrobial de-escalation in 0.2% of patients(1/47).Antimicrobial changes were mostly due to the identification of fastidious organisms such as Legionella,Pneumocystis jirovecii,and Candida.In the remaining three cases,NGS detected clinically relevant pathogens also detected by conventional cultures a few days later.The antimicrobial treatment was subsequently adjusted according to the susceptibility test results.Overall,NGS changed antimicrobial management in 55.3%(26/47)of patients,and conventional culture detected clinically relevant pathogens in 14.9% of the patients(7/47).CONCLUSION:With its rapid identification and high sensitivity,NGS could be a promising tool for identifying relevant pathogens and enabling rapid appropriate treatment in immunocompromised patients with suspected infections.

Diagnostic Value of Targeted Next-generation Sequencing in Pulmonary Mycobacterial Infections

Objective This study aimed to explore the diagnostic value of novel technique-targeted next-generation sequencing(tNGS)of bronchoalveolar lavage fluid(BALF)in pulmonary mycobacterial infections.Methods This retrospective study was conducted on patients who underwent bronchoscopy and tNGS,smear microscopy,and mycobacterial culture of BALF.Patients with positive Mycobacterium tuberculosis(MTB)culture or GeneXpert results were classified into the tuberculosis case group.Those diagnosed with nontuberculous mycobacteria(NTM)-pulmonary disease(NTM-PD)composed the case group of NTM-PD patients.The control group comprised patients without tuberculosis or NTM-PD.Sensitivity,specificity,and receiver operating characteristic(ROC)curves were used to evaluate the diagnostic performance.Results For tuberculosis patients with positive mycobacterial culture results,the areas under the ROC curves(AUCs)for tNGS,GeneXpert,and smear microscopy were 0.975(95%CI:0.935,1.000),0.925(95%CI:0.859,0.991),and 0.675(95%CI:0.563,0.787),respectively.For tuberculosis patients with positive GeneXpert results,the AUCs of tNGS,culture,and smear microscopy were 0.970(95%CI:0.931,1.000),0.850(95%CI:0.770,0.930),and 0.680(95%CI:0.579,0.781),respectively.For NTM-PD,the AUCs of tNGS,culture,and smear-positive but GeneXpert-negative results were 0.987(95%CI:0.967,1.000),0.750(95%CI:0.622,0.878),and 0.615(95%CI:0.479,0.752),respectively.The sensitivity and specificity of tNGS in NTM-PD patients were 100%and 97.5%,respectively.Conclusion tNGS demonstrated superior diagnostic efficacy in mycobacterial infection,indicating its potential for clinical application.

Bronchoalveolar lavage fluid metagenomic next-generation sequencing assay for identifying pathogens in lung cancer patients

Background:For patients with lung cancer,timely identification of new lung lesions as infectious or non-infectious,and accurate identification of pathogens is very important in improving OS of patients.As a new auxiliary examination,metagenomic next-generation sequencing(mNGS)is believed to be more accurate in diagnosing infectious diseases in patients without underlying diseases,compared with conventional microbial tests(CMTs).We designed this study tofind out whether mNGS has better performance in distinguishing infectious and non-infectious diseases in lung cancer patients using bronchoalveolar lavagefluid(BALF).Materials and Methods:This study was a real-world retrospective review based on electronic medical records of lung cancer patients with bronchoalveolar lavage(BAL)and BALF commercial mNGS testing as part of clinical care from 1 April 2019 through 30 April 2022 at The First Affiliated Hospital of Sun Yat-sen University.164 patients were included in this study.Patients were categorized into the pulmonary non-infectious disease(PNID)group(n=64)and the pulmonary infectious disease(PID)group(n=100)groups based onfinal diagnoses.Results:BALF mNGS increased the sensitivity rate by 60%compared to CMTs(81%vs.21%,p<0.05),whereas there was no significant difference in specificity(75%vs.98.4%,p>0.1).Among the patients with PID,bacteria were the most common cause of infection.Fungal infections occurred in 32%of patients,and Pneumocystis Yersini was most common.Patients with Tyrosine kinase inhibitors(TKIs)therapy possess longer overall survival(OS)than other anti-cancer agents,the difference between TKIs and immuno-checkpoint inhibitors(ICIs)was insignificant(median OS TKIs vs.ICIs vs.Anti-angiogenic vs.Chemo vs.Radiotherapy=76 vs.84 vs.61 vs.58 vs.60).Conclusions:our study indicates that BALF mNGS can add value by improving overall sensitivity in lung cancer patients with potential pulmonary infection,and was outstanding in identifying Pneumocystis infection.It could be able to help physicians adjust the follow-up treatment to avoid the abuse of antibiotics.

Brain abscess from oral microbiota approached by metagenomic next-generation sequencing: A case report and review of literature

BACKGROUND Brain abscess is a serious and potentially fatal disease caused primarily by microbial infection.Although progress has been made in the diagnosis and treatment of brain abscesses,the diagnostic timeliness of pathogens needs to be improved.CASE SUMMARY We report the case of a 54-year-old male with a brain abscess caused by oral bacteria.The patient recovered well after receiving a combination of metagenomic next-generation sequencing(mNGS)-assisted guided medication and surgery.CONCLUSION Therefore,mNGS may be widely applied to identify the pathogenic microor-ganisms of brain abscesses and guide precision medicine.

Metagenomic next-generation sequencing may assist diagnosis of osteomyelitis caused by Mycobacterium houstonense:A case report

BACKGROUND Mycobacterium houstonense(M.houstonense)belongs to the nontuberculous mycobacterium group.Infection caused by M.houstonense is prone to recurrence.CASE SUMMARY We present a patient who was diagnosed with osteomyelitis caused by M.houstonense and treated with a combination of cefoxitin,and amikacin combined with linezolid.CONCLUSION The emergence of metagenomic next-generation sequencing(NGS)has brought new hope for the diagnosis and treatment of listeria meningitis.NGS can analyze a large number of nucleic acid sequences in a short time and quickly determine the pathogen species in the sample.Compared with traditional cerebrospinal fluid culture,NGS can greatly shorten the diagnosis time and provide strong support for the timely treatment of patients.Regarding treatment,NGS can also play an important role.Rapid and accurate diagnosis can enable patients to start targeted treatment as soon as possible and improve the treatment effect.At the same time,by monitoring the changes in pathogen resistance,the treatment plan can be adjusted in time to avoid treatment failure.

Diagnosis of Acute Q Fever in an Elderly Patient Using Metagenomic Next-Generation Sequencing:A Case Report

Query fever(Q fever)is a globally spread zoonotic disease caused by Coxiella burnetii,commonly found in natural foci but rarely seen in Hebei Province.The clinical manifestations of Q fever are diverse and nonspecific,which often leads to missed or incorrect diagnoses in clinical practice.This article reports a case of acute Q fever diagnosed in an elderly patient using metagenomic next-generation sequencing.

Research on Next-Generation Internet Architecture

The primary problem during the evolvement of next-generation Internet is the contradiction between growing requirements for Internet and the insufficient development of network theory and technology. As the fundamental principles to guide the developing direction of Internet, the study of Internet architecture is always a focus in the research community. To address the core issue of network scalability, we propose multi-dimension scalable architecture of next-generation Internet, the main idea of which is to extend the single-dimension scalability of traditional Internet on interconnection to multi-dimension scalability of next-generation Internet. The multi-dimension scalability is composed of scale-scalability, performance-scalability, security-scalability, function-scalability, and service-scalability. We suggest five elements, namely, IPv6, authentic IPv6 addressing, scalable processing capacity of routers, end-to-end connectionless Quality-of-Service control, and 4over6 mechanism to realize the multi-dimension scalability. The current research results show that the multi-dimension scalable architecture composed of these five elements will bring great influence on next-generation Internet.

Application of next-generation sequencing in thalassemia screening:A systematic review and meta-analysis

Objective:To evaluate the value of next-generation sequencing(NGS)in the prevention and management of thalassemia.Methods:A systematic search was performed in eight databases including China Biomedical Literature Database,Chinese National Knowledge Infrastructure,Chinese Scientific Journals Database,Wanfang database,PubMed,EMBASE,Web of Science,and Cochrane Library from the inception to 1 June 2022.Stata 17.0 and Review Manager 5.4 were used for the meta-analysis.Results:Nine studies containing 14794 participants were included in the meta-analysis.Compared with the routine genetic testing(including Gap-PCR and reverse dot blot),NGS had higher detection rates in screening thalassemia(RR 1.22,95%CI 1.13-1.31,P<0.01),particularly for theα-thalassaemia mutation carriers(RR 1.24,95%CI 1.07-1.44,P<0.01).However,no significant difference was found in the screening ofβ-thalassemia(RR 1.10,95%CI 0.99-1.23,P>0.05).Conclusions:Compared with routine genetic testing,NGS had a higher detection rate in general,particularly in the detection ofα-thalassemia.

Next-generation Sequencing of MHC Class Ⅰ Genes Reveals Trans-species Polymorphism in Eutropis multifasciata and Other Species of Scincidae

The genes of the major histocompatibility complex(MHC) encode cell surface proteins that are essential for adaptive immunity. MHC genes show the most prominent genetic diversity in vertebrates,reflecting the adaptation of populations to their evolving environment, population survival and reproduction. In the present study, we used nextgeneration sequencing(NGS) to study the loci polymorphism of exon 3 of the MHC class Ⅰ genes in an ovoviviparous skink, the many-lined sun skink,Eutropis multifasciata and five other species of Scincidae, to quantify genetic variation. In addition,we genotyped the same MHC class Ⅰ genes of E.multifasciata using clone sequencing, to directly compare the effectiveness of both analytical techniques for MHC genotyping. NGS detected 20MHC class Ⅰ alleles in E. multifasciata, and 2 to 15 alleles in the other five Scincidae species. However,clone sequencing detected only 15 of those MHC class Ⅰ alleles in E. multifasciata. In addition, transspecies polymorphism of MHC class Ⅰ genes was studied by constructing a phylogenetic tree using the gene sequences obtained by NGS. Phylogenetic analysis revealed that MHC class I alleles were shared among different species of Scincidae with trans-species polymorphism, and did not exhibit specific genealogical inheritance. These results have important implications for understanding polymorphism interspecies diversity in the MHC genes of Scincidae, and the evolution of the MHC more broadly.

Pulmonary nocardiosis with bloodstream infection diagnosed by metagenomic next-generation sequencing in a kidney transplant recipient:A case report

BACKGROUND Pulmonary nocardiosis is difficult to diagnose by culture and other conventional testing,and is often associated with lethal disseminated infections.This difficulty poses a great challenge to the timeliness and accuracy of clinical detection,especially in susceptible immunosuppressed individuals.Metagenomic nextgeneration sequencing(mNGS)has transformed the conventional diagnosis pattern by providing a rapid and precise method to assess all microorganisms in a sample.CASE SUMMARY A 45-year-old male was hospitalized for cough,chest tightness and fatigue for 3 consecutive days.He had received a kidney transplant 42 d prior to admission.No pathogens were detected at admission.Chest computed tomography showed nodules,streak shadows and fiber lesions in both lung lobes as well as right pleural effusion.Pulmonary tuberculosis with pleural effusion was highly suspected based on the symptoms,imaging and residence in a high tuberculosisburden area.However,anti-tuberculosis treatment was ineffective,showing no improvement in computed tomography imaging.Pleural effusion and blood samples were subsequently sent for mNGS.The results indicated Nocardia farcinica as the major pathogen.After switching to sulphamethoxazole combined with minocycline for antinocardiosis treatment,the patient gradually improved and was finally discharged.CONCLUSION A case of pulmonary nocardiosis with an accompanying bloodstream infection was diagnosed and promptly treated before the dissemination of the infection.This report emphasizes the value of mNGS in the diagnosis of nocardiosis.mNGS may be an effective method for facilitating early diagnosis and prompt treatment in infectious diseases,which overcomes the shortcomings of conventional testing.

Construction of SNP genetic maps based on targeted next-generation sequencing and QTL mapping of vital agronomic traits in faba bean(Vicia faba L.)

Owing to the limitation of a large genome size(~13 Gb),the genetic and gene mapping studies on faba bean(Vicia faba L.)are lagging far behind those for other legumes.In this study,we selected three purified faba bean lines(Yundou 8137,H0003712,and H000572)as parents and constructed two F2 populations.These two F2 populations,namely 167 F2 plants in Pop1(Yundou 8137×H0003712)and 204 F2 plants in Pop2(H000572×Yundou 8137),were genotyped using a targeted next-generation sequencing(TNGS)genotyping platform,and two high-density single nucleotide polymorphisms(SNP)genetic linkage maps of faba bean were constructed.The map constructed from Pop1 contained 5103 SNPs with a length of 1333.31 cM and an average marker density of 0.26 cM.The map constructed from Pop2 contained 1904 SNPs with a greater length of 1610.61 cM.In these two F2 populations,QTL mapping identified 98 QTLs for 14 agronomic traits related to the flowers,pods,plant types and grains.The two maps were then merged into an integrated genetic linkage map containing 6895 SNPs,with a length of 3324.48 cM.These results not only lay the foundation for fine mapping and map-based cloning of related genes,but can also accelerate the molecular marker-assisted breeding of faba bean.

Metagenomic next-generation sequencing for pleural effusions induced by viral pleurisy:A case report

BACKGROUND Viral pleurisy is a viral infected disease with exudative pleural effusions.It is one of the causes for pleural effusions.Because of the difficult etiology diagnosis,clinically pleural effusions tend to be misdiagnosed as tuberculous pleurisy or idiopathic pleural effusion.Here,we report a case of pleural effusion secondary to viral pleurisy which is driven by infection with epstein-barr virus.Viral infection was identified by metagenomic next-generation sequencing(mNGS).CASE SUMMARY A 40-year-old male with a history of dermatomyositis,rheumatoid arthritis,and secondary interstitial pneumonia was administered with long-term oral prednisone.He presented with fever and chest pain after exposure to cold,accompanied by generalized sore and weakness,night sweat,occasional cough,and few sputums.The computed tomography scan showed bilateral pleural effusions and atelectasis of the partial right lower lobe was revealed.The pleural fluids were found to be yellow and slightly turbid after pleural catheterization.Thoracoscopy showed fibrous adhesion and auto-pleurodesis.Combining the results in pleural fluid analysis and mNGS,the patient was diagnosed as viral pleuritis.After receiving Aciclovir,the symptoms and signs of the patient were relieved.CONCLUSION Viral infection should be considered in cases of idiopathic pleural effusion unexplained by routine examination.mNGS is helpful for diagnosis.

Molecular methods for colorectal cancer screening:Progress with next-generation sequencing evolution

Currently,colorectal cancer(CRC)represents the third most common malignancy and the second most deadly cancer worldwide,with a higher incidence in developed countries.Like other solid tumors,CRC is a heterogeneous genomic disease in which various alterations,such as point mutations,genomic rearrangements,gene fusions or chromosomal copy number alterations,can contribute to the disease development.However,because of its orderly natural history,easily accessible onset location and high lifetime incidence,CRC is ideally suited for preventive intervention,but the many screening efforts of the last decades have been compromised by performance limitations and low penetrance of the standard screening tools.The advent of next-generation sequencing(NGS)has both facilitated the identification of previously unrecognized CRC features such as its relationship with gut microbial pathogens and revolutionized the speed and throughput of cataloguing CRC-related genomic alterations.Hence,in this review,we summarized the several diagnostic tools used for CRC screening in the past and the present,focusing on recent NGS approaches and their revolutionary role in the identification of novel genomic CRC characteristics,the advancement of understanding the CRC carcinogenesis and the screening of clinically actionable targets for personalized medicine.

Comprehensive next-generation sequencing reveals double primary colorectal carcinoma missed by diagnostic imaging: A case report

BACKGROUND Multiple primary colorectal carcinoma(MPCC)is a rare clinical disease,which is challenging to differentiate from metastatic disease using histopathological methods.Next-generation sequencing(NGS)has been employed to identify multiple primary cancers.CASE SUMMARY This study a rare case of a 63-year-old male patient diagnosed with MPCC by targeted NGS,which was initially missed by radiological evaluation.The patient was found to have two tumors located on the surface of the colorectum which had distinct genomic alterations.Based on wild-type KRAS detected in the unresected tumor,the patient benefited from the epidermal growth factor receptor(EGFR)inhibitor cetuximab treatment,but developed novel mutations including KIF5B-RET fusion,which provides a possible resistance mechanism to anti-EGFR therapy.CONCLUSION Our case highlights the necessity of using genetic testing for primary tumor diagnosis and the application of serial plasma circulating tumor DNA profiling for dynamic disease monitoring.

Soft tissue tuberculosis detected by next-generation sequencing:A case report and review of literature

BACKGROUND Soft tissue tuberculosis is rare and insidious,with most patients presenting with a localized enlarged mass or swelling,which may be factors associated with delayed diagnosis and treatment.In recent years,next-generation sequencing has rapidly evolved and has been successfully applied to numerous areas of basic and clinical research.A literature search revealed that the use of next-generation sequencing in the diagnosis of soft tissue tuberculosis has been rarely reported.CASE SUMMARY A 44-year-old man presented with recurrent swelling and ulcers on the left thigh.Magnetic resonance imaging suggested a soft tissue abscess.The lesion was surgically removed and tissue biopsy and culture were performed;however,no organism growth was detected.Finally,Mycobacterium tuberculosis was confirmed as the pathogen responsible for infection through next-generation sequencing analysis of the surgical specimen.The patient received a standardized anti-tuberculosis treatment and showed clinical improvement.We also performed a literature review on soft tissue tuberculosis using studies published in the past 10 years.CONCLUSION This case highlights the importance of next-generation sequencing for the early diagnosis of soft tissue tuberculosis,which can provide guidance for clinical treatment and improve prognosis.

Metagenomic next-generation sequencing in the diagnosis of neurocysticercosis:A case report

BACKGROUND The clinical symptoms and imaging manifestations of neurocysticercosis(NCC)are very different,and the difficulty and delay of clinical diagnoses may lead to an increase in mortality and disability.Rapid and accurate pathogen identification is important for the treatment of these patients.Metagenomic next-generation sequencing(mNGS)is a powerful tool to identify pathogens,especially in infections that are difficult to identify by conventional methods.CASE SUMMARY A 43-year-old male patient was admitted due to a recurrent headache for a few months.Imaging examinations showed hydrocephalus and cystic lesions,which were considered to be a central nervous system infection,but no etiology was found by routine examination.mNGS of the cerebrospinal fluid revealed high Taenia solium reads,and the positive results of a cysticercosis antibody test confirmed the infection.Combined with the patient’s clinical manifestations,the etiological evidence,and the imaging manifestation,the patient was finally diagnosed with NCC and he was prescribed dexamethasone,albendazole,neurotrophic drugs,and intracranial pressure reduction therapy.The headaches disappeared after anti-parasite treatment,and no associated symptoms recurred prior to the three-and six-month follow-up.CONCLUSION As an accurate and sensitivity detection method,mNGS can be a reliable approach for the diagnosis of NCC.

Next-generation sequencing technology for the diagnosis of Pneumocystis pneumonia in an immunocompetent female:A case report

BACKGROUND Pneumocystis pneumonia(PCP)is a serious fungal infection usually seen in patients with human immunodeficiency virus,and it is more frequently found and has a high fatality rate in immunocompromised people.Surprisingly,it rarely occurs in immunocompetent patients.However,the clinical diagnosis of this pathogen is made more difficult by the difficulty of obtaining accurate microbiological evidence with routine tests.This case reports a PCP patient with normal immune function who was diagnosed through next-generation sequencing(NGS).CASE SUMMARY A 23-year-old female who had no special disease in the past was admitted to the hospital with a persistent fever and cough.Based on the initial examination results,the patient was diagnosed with bipulmonary pneumonia,and empirical broad-spectrum antibiotic therapy was administered.However,due to the undetermined etiology,the patient's condition continued to worsen.She was transferred to the intensive care unit because of acute respiratory failure.After the diagnosis of Pneumocystis jirovecii infection through NGS in bronchoalveolar lavage fluid and treatment with trimethoprim/sulfamethoxazole and caspofungin,the patient gradually recovered and had a good prognosis.CONCLUSION This case emphasizes that,for patients with normal immune function the possibility of PCP infection,although rare,cannot be ignored.NGS plays an important role in the diagnosis of refractory interstitial pneumonia and acute respiratory failure.

Blockchain-as-a-Utility for Next-Generation Healthcare Internet of Things

The scope of the Internet of Things(IoT)applications varies from strategic applications,such as smart grids,smart transportation,smart security,and smart healthcare,to industrial applications such as smart manufacturing,smart logistics,smart banking,and smart insurance.In the advancement of the IoT,connected devices become smart and intelligent with the help of sensors and actuators.However,issues and challenges need to be addressed regarding the data reliability and protection for signicant nextgeneration IoT applications like smart healthcare.For these next-generation applications,there is a requirement for far-reaching privacy and security in the IoT.Recently,blockchain systems have emerged as a key technology that changes the way we exchange data.This emerging technology has revealed encouraging implementation scenarios,such as secured digital currencies.As a technical advancement,the blockchain network has the high possibility of transforming various industries,and the next-generation healthcare IoT(HIoT)can be one of those applications.There have been several studies on the integration of blockchain networks and IoT.However,blockchain-as-autility(BaaU)for privacy and security in HIoT systems requires a systematic framework.This paper reviews blockchain networks and proposes BaaU as one of the enablers.The proposed BaaU-based framework for trustworthiness in the next-generation HIoT systems is divided into two scenarios.The rst scenario suggests that a healthcare service provider integrates IoT sensors such as body sensors to receive and transmit information to a blockchain network on the IoT devices.The second proposed scenario recommends implementing smart contracts,such as Ethereum,to automate and control the trusted devices’subscription in the HIoT services.

Pneumocystis jirovecii diagnosed by next-generation sequencing of bronchoscopic alveolar lavage fluid: A case report and review of literature

BACKGROUND The advent of molecular targeted agents and immune checkpoint inhibitors has greatly improved the treatment of advanced renal cell carcinoma(RCC), thus significantly improving patient survival. The incidence of rare drug-related adverse events has gained increased attention.CASE SUMMARY We report a patient with advanced RCC treated with multiple lines of molecular targeted agents and immune checkpoint inhibitors, who developed a pulmonary infection after treatment with everolimus in combination with lenvatinib. Determining the pathogenic organism was difficult, but it was eventually identified as Pneumocystis jirovecii by next-generation sequencing(NGS) of bronchoscopic alveolar lavage fluid(BALF) and successfully treated with trimethoprim-sulfamethoxazole.CONCLUSION Rare pulmonary infections caused by molecular targeted agents are not uncommon in clinical practice, but their diagnosis is difficult. Evaluating BALF with NGS is a good method for rapid diagnosis of such infections.

The Internet of Things and Next-generation Public Health Information Systems

The Internet of things has particularly novel implications in the area of public health. This is due to (1) The rapid and widespread adoption of powerful contemporary Smartphone’s;(2) The increasing availability and use of health and fitness sensors, wearable sensor patches, smart watches, wireless-enabled digital tattoos and ambient sensors;and (3) The nature of public health to implicitly involve connectivity with and the acquisition of data in relation to large numbers of individuals up to population scale. Of particular relevance in relation to the Internet of Things (IoT) and public health is the need for privacy and anonymity of users. It should be noted that IoT capabilities are not inconsistent with maintaining privacy, due to the focus of public health on aggregate data not individual data and broad public health interventions. In addition, public health information systems utilizing IoT capabilities can be constructed to specifically ensure privacy, security and anonymity, as has been developed and evaluated in this work. In this paper we describe the particular characteristics of the IoT that can play a role in enabling emerging public health capabilities;we describe a privacy-preserving IoT-based public health information system architecture;and provide a privacy evaluation.