Disruption of non-classically secreted protein(MoMtp)compromised conidiation,stress homeostasis,and pathogenesis of Magnaporthe oryzae

Blast disease,caused by the hemibiotrophic ascomycete fungus,Magnaporthe oryzae,is a significant threat to sustainable rice production worldwide.Studies have shown that the blast fungus secretes vast arrays of functionally diverse proteins into the host cell for a successful disease progression.However,the final destinations of these effector proteins inside the host cell and their role in advancing fungal pathogenesis remain a mystery.Here,we reported that a putative mitochondrial targeting non-classically secreted protein(MoMtp)positively regulates conidiogenesis and appressorium maturation in M.oryzae.Moreover,MoM TP gene deletion mutant strains triggered a hypersensitive response when inoculated on rice leaves displaying that MoMtp is essential for the virulence of M.oryzae.In addition,cell wall and oxidative stress results indicated that MoMtp is likely involved in the maintenance of the structural integrity of the fungus cell.Our study also demonstrates an upregulation in the expression pattern of the MoMTP gene at all stages of infection,indicating its possible regulatory role in host invasion and the infectious development of M.oryzae.Furthermore,Agrobacterium infiltration and sheath inoculation confirmed that MoMtpGFP protein is predominantly localized in the host mitochondria of tobacco leaf and rice cells.Taken together,we conclude that MoMtp protein likely promotes the normal conidiation and pathogenesis of M.oryzae and might have a role in disturbing the proper functioning of the host mitochondria during pathogen invasion.

Life cycle and pathogenesis of hepatitis D virus:A review

Hepatitis D virus(HDV) is a defective RNA virus which requires the help of hepatitis B virus(HBV) virus for its replication and assembly of new virions. HDV genome contains only one actively transcribed open reading frame which encodes for two isoforms of hepatitis delta antigen. Post-translational modifications of small and large delta antigens(S-HDAg and L-HDAg) involving phosphorylation and isoprenylation respectively con- fer these antigens their specific properties. S-HDAg is required for the initiation of the viral genome replica- tion, whereas L-HDAg serves as a principal inhibitor of replication and is essential for the assembly of new virion particles. Immune mediation has usually been implicated in HDV-associated liver damage. The patho- genesis of HDV mainly involves interferon-α signaling inhibition, HDV-specific T-lymphocyte activation and cytokine responses, and tumor necrosis factor-alpha and nuclear factor kappa B signaling. Due to limited protein coding capacity, HDV makes use of host cel- lular proteins to accomplish their life cycle processes, including transcription, replication, post-transcriptional and translational modifications. This intimate host- pathogen interaction significantly alters cell proteome and is associated with an augmented expression of pro-inflammatory, growth and anti-apoptotic factorswhich explains severe necroinflammation and increased cell survival and an early progression to hepatocellular carcinoma in HDV patients. The understanding of the process of viral replication, HBV-HDV interactions, and etio-pathogenesis of the severe course of HDV infection is helpful in identifying the potential therapeutic targets in the virus life cycle for the prophylaxis and treatment of HDV infection and complications.

FpPDE1 function of Fsarium pseudograminearum on pathogenesis in wheat

Wheat crown rot caused by Fusarium spp. is a common disease worldwide. Both Fusarium pseudograminearum and Fusarium graminearum infect wheat crown and produce mycotoxin leading to grain loss due to white head. F. pseudograminearum （Fp） was reported in wheat from Henan Province of China a couple of years ago. The wheat crown rot （CR） caused by this new pathogen is as an emerging severe disease of wheat, which has recently expanded to several provinces in China and is, therefore, under rapid investigation. Colonization of wheat tissue by Fp is accomplished though the formation of a septated foot-shaped appressoria and generation of a penetration peg to break through the internal cells of leaf sheath. The molecular mechanism by which Fp regulates the pathogenesis on wheat host is unclear. Here, we report FpPDE1, a P-type ATPase-encoding predicted PDE1 orthologue gene of Magnaporthe oryzae, belonging to the DRS2 subfamily of aminophospholipid translocases. The gene deletion of FpPDE1 with the split-marker approach did not obviously affect hyphae growth and conidiation, but led to an attenuated virulence on wheat base stem and root. Our finding indicates that the putative aminophospholipid translocases is not essential for the infectious hyphae development in Fp.

Incorporating mucosal-associated invariant T cells into the pathogenesis of chronic liver disease

Mucosal-associated invariant T(MAIT)cells have been described in liver and nonliver diseases,and they have been ascribed antimicrobial,immune regulatory,protective,and pathogenic roles.The goals of this review are to describe their biological properties,indicate their involvement in chronic liver disease,and encourage investigations that clarify their actions and therapeutic implications.English abstracts were identified in PubMed by multiple search terms,and bibliographies were developed.MAIT cells are activated by restricted non-peptides of limited diversity and by multiple inflammatory cytokines.Diverse pro-inflammatory,anti-inflammatory,and immune regulatory cytokines are released;infected cells are eliminated;and memory cells emerge.Circulating MAIT cells are hyper-activated,immune exhausted,dysfunctional,and depleted in chronic liver disease.This phenotype lacks disease-specificity,and it does not predict the biological effects.MAIT cells have presumed protective actions in chronic viral hepatitis,alcoholic hepatitis,non-alcoholic fatty liver disease,primary sclerosing cholangitis,and decompensated cirrhosis.They have pathogenic and pro-fibrotic actions in autoimmune hepatitis and mixed actions in primary biliary cholangitis.Local factors in the hepatic microenvironment(cytokines,bile acids,gut-derived bacterial antigens,and metabolic by-products)may modulate their response in individual diseases.Investigational manipulations of function are warranted to establish an association with disease severity and outcome.In conclusion,MAIT cells constitute a disease-nonspecific,immune response to chronic liver inflammation and infection.Their pathological role has been deduced from their deficiencies during active liver disease,and future investigations must clarify this role,link it to outcome,and explore therapeutic interventions.

Pathogenesis and Immunogenicity of an Avian H9N2 Influenza Virus Isolated from Human

Objective To investigate the pathogenesis and immunogenicity of H9N2 influenza virus A/Guangzhou/333/99 （a reassortant of G1 and G9 viruses isolated from a female patient in 1999） in a mouse model of infection.Methods Mice were infected with increasing virus titers.Viral load in the lungs and trachea was determined by EID50 assay.Pulmonary histopathology was assessed by hematoxylin‐eosin staining.Anti‐HI antibody titers and T‐cell responses to viral HA were determined by ELISPOT and confirmed by flow cytometry.Results Mice presented a mild syndrome after intranasal infection with A/Guangzhou/333/99 （H9N2） influenza virus.Virus was detected in the trachea and lungs of mice harvested on days 3,6,and 9 post‐infection.A T‐cell response to viral HA was detected on day 6 and H9 HA‐specific CD 4＋ T‐cells predominated.Seroconversion was detected after 14 days and antibody persisted for at least 28 weeks.Conclusion Our results suggest that H9N2 （A/Guangzhou/333/99） can replicate in the murine respiratory tract without prior adaptation,and both humoral and cell‐mediated immunity play an important role in the immune response.

FgGyp8 as a putative FgRab1 GAP is required for growth and pathogenesis by regulating FgSnc1-mediated secretory vesicles fusion in Fusarium graminearum

Fusarium graminearum is an important plant pathogenic fungus that causes disease and yield reduction in many cereal crops, such as wheat and barley. Gyp8 stimulates GTP hydrolysis on Ypt1 in yeast. However, the functions of Gyp8 in plant pathogenic fungi are still unknown. In this study, we investigated the roles of Fg Gyp8 in F. graminearum by genetic and pathological analyses. Through gene knockout and phenotypic analyses, we found that Fg Gyp8 is required for vegetative growth in F. graminearum. The conidiation, conidial size and number of septa per conidium of ΔFggyp8 mutant are significantly reduced when compared to the wild type PH-1. Furthermore, Fg Gyp8 is crucial for pathogenicity on wheat coleoptiles and wheat heads. Fg Gyp8 contains a conserved TBC domain. Domain deletion analysis showed that the TBC domain, C-and N-terminal regions of Fg Gyp8 are all important for its biological functions in F. graminearum. Moreover, we showed that Fg Gyp8 catalyzes the hydrolysis of the GTP on Fg Rab1 to GDP in vitro, indicating that Fg Gyp8 is a GTPase-activating protein(GAP) for Fg Rab1. In addition, we demonstrated that Fg Gyp8 is required for Fg Snc1-mediated fusion of secretory vesicles with the plasma membrane in F. graminearum. Finally, we showed that Fg Gyp8 has functional redundancy with another Fg Rab1 GAP, Fg Gyp1, in F. graminearum. Taken together, we conclude that Fg Gyp8 is required for vegetative growth, conidiogenesis, pathogenicity and acts as a GAP for Fg Rab1 in F. graminearum.

Discussion on Etiology and Pathogenesis of Corona Virus Disease 2019 from“Cold-dampness and Insidious Dryness”

Based on the characteristics of the epidemic situation and the authors’understanding of the related ancient books and documents,this paper explores the etiology and pathogenesis of Corona Virus Disease 2019(COVID-19)from 5 aspects:abnormal climate in"warm winter",unique geographical location,pathogenesis evolution of cold and dampness mixed with insidious dryness,transmission and change of"triple energizer"of toxic pathogens,and game between healthy Qi and toxic pathogens.Combined with the special treatment of traditional Chinese medicine(TCM),the purpose is to make a modest contribution to curbing the epidemic situation with TCM.

Nascent Polypeptide-Associated Complex Involved in the Development and Pathogenesis of Fusarium graminearum on Wheat

Reliable knowledge on pathogenic agents contributes to effective plant protection.For most plant pathogens,maintaining protein homeostasis(proteostasis)is essential for unfolding the cellular functions to survive and thrive.However,the fungal proteins involved in proteostasis remain poorly characterized in the process of pathogenesis.In this study,we characterized the function of the nascent polypeptideassociated complex(NAC)in Fusarium graminearum(F.graminearum)(FgNAC),one of the top 10 fungal pathogens with predominant scientific/economic importance.We found that FgNACa,a subunit of FgNAC,manifests high structural and functional similarity to its homologous counterparts in yeast and other species.The mutants of F.graminearum lacking NACa are viable but suffer significant defects in vegetative growth,conidial production,and pathogenesis.In addition,we show here that FgNACa can interact with another subunit of NAC(FgNACb)in a yeast-two-hybrid assay.The subcellular localization results show that FgNACa and FgNACb are predominantly localized in the cytoplasm.Future studies should focus on deciphering the mechanism by which NAC orchestrates protein biogenesis and consequentially modulates development and pathogenesis.

Pathogenesis and drug resistance mechanism of Burkholderia pseudomallei

Burkholderia pseudomallei is the pathogen that causes melioidosis.Melioidosis has a long duration of chronic infection,atypical clinical manifestations at acute onset,and is prone to life-threatening complications and poor prognosis.Understanding the pathogenesis and drug resistance mechanism of Burkholderia pseudomallei will effectively help the diagnosis and treatment of the disease and improve the prognosis.This review focuses on the extracellular movement of Burkholderia pseudomallei in host cells,the way of infecting host cells,virulence factors,and drug resistance mechanisms(efflux pumps,changes in target sites,etc.).This study provides a possible direction for the early diagnosis,treatment and control of melioidosis caused by this bacterium.

The Wheat Pathogenesis Related Protein (TdPR1.2) Ensures Contrasting Behaviors to <i>E. coli</i>Transformant Cells under Stress Conditions

The pathogenesis-related proteins 1 (PR-1) gene family play important roles in the plant metabolism in response to biotic and abiotic stresses. The wheat TdPR1.2 has been previously isolated and characterized. Here we showed by bio-informatic analysis that TdPR1.2 contains six cysteine residues that are conserved between all PR-1 proteins tested. Using ScanProsite tool, we found that TdPR1.2 structure has a CRISP family signature 1 and 2 located at the C-terminal part of the protein. Those two domains are conserved in many identified PR1.2 proteins in plants. Moreover, SignalIP-5.0 analysis revealed that TdPR1.2 contains a putative signal peptide formed by 25 amino acids at the N-terminal extremity. The presence of this signal peptide suggested that the mature proteins will be secreted after the cleavage of the signal sequence. Further, we investigate the role of the TdPR1.2 proteins in the growth of Escherichia coli transformants cells under different abiotic stresses. Our results showed that the full-length form of TdPR1.2 enhanced tolerance of E. coli against salt and osmotic stress but not to KCl. Moreover, TdPR1.2 protein confers bacterial tolerance to heavy metals in solid and liquid mediums. Based on these results, we suggest that the TdPR1.2 protein could play an important role in response to abiotic stress conditions.

Exploration on the Treatment of Upper Airway Cough Syndrome Based on the Pathogenesis of Hidden Pathogen(Evil)

Upper airway cough syndrome refers to a clinical syndrome with chronic cough as the main manifestation caused by the reverse flow of secretions from various upper airway diseases such as allergic rhinitis,rhino-sinusitis,adenoid hypertrophy and other parts of the nose and pharynx.Professor Yin Dan believes that the main disease of upper airway cough syndrome is due to cold fluid retained in lung,blood stasis endogenous,compound feeling of wind evil.The pathogenesis is external wind evil and the hidden pathogen together to cause the disease.In the treatment,the warm cold fluid retention is the core,accompanied by the activation of blood and wind,so that the wind evil can go away,the cold evil can be warmed,the phlegm can be removed,and the blood stasis can be dispersed,so as to achieve the purpose of treating the upper airway cough syndrome of the children’s cold fluid retention in lung.

Immunopathogenesis and immunomodulatory therapy for myocarditis

Myocarditis is an inflammatory cardiac disease characterized by the destruction of myocardial cells, infiltration of interstitial inflammatory cells, and fibrosis, and is becoming a major public health concern. The aetiology of myocarditis continues to broaden as new pathogens and drugs emerge. The relationship between immune checkpoint inhibitors, severe acute respiratory syndrome coronavirus 2, vaccines against coronavirus disease-2019, and myocarditis has attracted increased attention. Immunopathological processes play an important role in the different phases of myocarditis, affecting disease occurrence, development, and prognosis. Excessive immune activation can induce severe myocardial injury and lead to fulminant myocarditis,whereas chronic inflammation can lead to cardiac remodelling and inflammatory dilated cardiomyopathy. The use of immunosuppressive treatments, particularly cytotoxic agents, for myocarditis, remains controversial. While reasonable and effective immunomodulatory therapy is the general trend. This review focuses on the current understanding of the aetiology and immunopathogenesis of myocarditis and offers new perspectives on immunomodulatory therapies.

Amyloid,tau,pathogen infection and antimicrobial protection in Alzheimer’s disease-conformist,nonconformist,and realistic prospects for AD pathogenesis

Background:Alzheimer’s disease(AD)is a fatal disease that threatens the quality of life of an aging population at a global scale.Various hypotheses on the etiology of AD have been developed over the years to guide efforts in search of therapeutic strategies.Main body:In this review,we focus on four AD hypotheses currently relevant to AD onset:the prevailing amyloid cascade hypothesis,the well-recognized tau hypothesis,the increasingly popular pathogen(viral infection)hypothesis,and the infection-related antimicrobial protection hypothesis.In briefly reviewing the main evidence supporting each hypothesis and discussing the questions that need to be addressed,we hope to gain a better understanding of the complicated multi-layered interactions in potential causal and/or risk factors in AD pathogenesis.As a defining feature of AD,the existence of amyloid deposits is likely fundamental to AD onset but is insufficient to wholly reproduce many complexities of the disorder.A similar belief is currently also applied to hyperphosphorylated tau aggregates within neurons,where tau has been postulated to drive neurodegeneration in the presence of pre-existing Aβplaques in the brain.Although infection of the central nerve system by pathogens such as viruses may increase AD risk,it is yet to be determined whether this phenomenon is applicable to all cases of sporadic AD and whether it is a primary trigger for AD onset.Lastly,the antimicrobial protection hypothesis provides insight into a potential physiological role for Aβpeptides,but how Aβ/microbial interactions affect AD pathogenesis during aging awaits further validation.Nevertheless,this hypothesis cautions potential adverse effects in Aβ-targeting therapies by hindering potential roles for Aβin anti-viral protection.Conclusion:AD is a multi-factor complex disorder,which likely requires a combinatorial therapeutic approach to successfully slow or reduce symptomatic memory decline.A better understanding of how various causal and/or risk factors affecting disease onset and progression will enhance the likelihood of conceiving effective treatment paradigms,which may involve personalized treatment strategies for individual patients at varying stages of disease progression.

Molecular basis of pathogenesis of postharvest pathogenic Fungi and control strategy in fruits: progress and prospect

The disease caused by pathogenic fungi is the main cause of postharvest loss of fresh fruits.The formulation of disease control strategies greatly depends on the understanding of pathogenic mechanism of fungal pathogens and control strategy.In recent years,based on the application of various combinatorial research methods,some pathogenic genes of important postharvest fungal pathogens in fruit have been revealed,and their functions and molecular regulatory networks of virulence have been explored.These progresses not only provide a new perspective for understanding the molecular basis and regulation mechanism of pathogenicity of postharvest pathogenic fungi,but also are beneficial to giving theoretical guidance for the creation of new technologies of postharvest disease control.Here,we synthesized these recent advances and illustrated conceptual frameworks,and identified several issues on the focus of future studies.

Seed priming with Pseudomonas putida isolated from rhizosphere triggers innate resistance against Fusarium wilt in tomato through pathogenesis-related protein activation and phenylpropanoid pathway

Pathogenesis-related(PR)proteins are one of the major and preliminary proteins accumulated as a defense against biotic stress.This defense response can be induced by using beneficial rhizobacteria,which has been studied in various host-pathogen interactions.In the present study,eleven Pseudomonas isolates were assessed for their potential to ferment sorbitol,reduce nitrate,and produce mycolytic enzymes,1-aminocyclopropane-l-carboxylic acid(ACC)deaminase,phenazine antibiotics,and N-acyl homoserine lactones(AHLs).All isolates were tested against the host-specific pathogen Fusarium oxysporum MTCC1755 in tomato under greenhouse conditions,and shortlisted isolates were tested for their rhizosphere competence.In-vitro test results showed that the isolates were able to produce mycolytic enzymes,including protease,lipase,chitinase,cellulase,and amylase,and the antibiotic phenazine and were negative for pyoluteorin.All the isolates except two were positive for ACC deaminase production.Greenhouse results showed that the isolates M80,M96,and T109 significantly reduced symptoms of Fusarium wilt.Extended greenhouse tests under autoclaved and unautoclaved soil conditions showed that M80,M96,and T109 were excellent rhizosphere competitors and were identified as Pseudomonas putida.In brief,the defense-specific biochemical variations in the host could describe the improved defense against Fusarium wilt occurring in the primed plants.These three Pseudomonas strains could be used as potential biocontrol agents,along with their rhizosphere competence.

Efficiency and effectiveness of systems for the treatment of domestic wastewater based on subsurface flow constructed wetlands in Jarabacoa, Dominican Republic

Constructed wetlands(CwW)are well known nature-based systems for water treatment.This study evaluated the efficiency and effectiveness of seven domestic wastewater treatment systems based on horizontal flow CWs in Jarabacoa,the Dominican Republic.The results showed that the CWs were efficient in reducing the degree of contamination of wastewater to levels below the Dominican wastewater discharge standards for parameters such as the 5-day biochemical oxygen demand(BOD5)and chemical oxygen demand,but not for the removal of phosphorus and fecal coliforms.In addition,a horizontal flow subsurface wetland in the peri-urban area El Dorado was evaluated in terms of the performance of wastewater treatment in tropical climatic conditions.The concentrations of heavy metals,such as zinc,copper,chromium,and iron,were found to decrease in the effluent of the wetland,and the concentrations for nickel and manganese tended to increase.The levels of heavy metals in the effluent were lower than the limit values of the Dominican wastewater discharge standards.The construction cost of these facilities was around 200 USD per population equivalent,similar to the cost in other countries in the same region.This study suggested some solutions to the improved performance of CWs:selection of a microbial flora that guarantees the reduction of nitrates and nitrites to molecular nitrogen,use of endemic plants that bioaccumulate heavy metals,combination of constructed wetlands with filtration on activated carbon,and inclusion of water purification processes that allow to evaluate the reuse of treated water.

Control of highly pathogenic avian influenza through vaccination

The stamping-out strategy has been used to control highly pathogenic avian influenza viruses in many countries,driven by the belief that vaccination would not be successful against such viruses and fears that avian influenza virus in vaccinated birds would evolve more rapidly and pose a greater risk to humans.In this review,we summarize the successes in controlling highly pathogenic avian influenza in China and make suggestions regarding the requirements for vaccine selection and effectiveness.In addition,we present evidence that vaccination of poultry not only eliminates human infection with avian influenza virus,but also significantly reduces and abolishes some harmful characteristics of avian influenza virus.

A novel pathogen Fusarium cuneirostrum causing common bean(Phaseolus vulgaris)root rot in China

Several fungal pathogens cause root rot of common bean,among which Fusarium spp.are the most common pathogens causing Fusarium root rot(FRR)worldwide.FRR has been becoming an increasingly severe disease of common bean in China,but the species of Fusarium spp.have remained unclear.Thus,this study was performed to identify the pathogen causing common bean root rot in Liangcheng County,Inner Mongolia,China.Nineteen Fusarium-like isolates were obtained after pathogen isolation and purification.The pathogenicity test indicated that eight isolates caused severe disease symptoms on common bean,while 11 other isolates were not pathogenic.The eight pathogenic isolates,FCL1–FCL8,were identified as Fusarium cuneirostrum by morphological characterization and phylogenetic analysis using partial sequences of EF-1α,ITS,28S,and IGS regions.Host range test showed that the representative F.cuneirostrum isolate FCL3 was also pathogenic to mung bean,while not pathogenic to adzuki bean,chickpea,cowpea,faba bean,pea,and soybean.Moreover,50 common bean and 50 mung bean cultivars were screened for resistance to FRR,and seven highly resistant or resistant cultivars of common bean were identified,while no resistant cultivars of mung bean were screened.This study revealed that F.cuneirostrum was one of common bean FRR pathogens in Inner Mongolia and it could induce mung bean root rot as well.To our knowledge,this is the first report of F.cuneirostrum causing FRR of common bean in China.

Investigation of the epidemiology,pathogenicity and immunogenicity of Bordetella bronchiseptica isolated from cats and dogs in China from 2021 to 2023

Bordetella bronchiseptica(Bb)is recognized as a leading cause of respiratory diseases in dogs and cats.However,epidemiological data on Bb in dogs and cats in China are still limited,and there is no commercially available vaccine.Live vaccines containing Bb that are widely used abroad are generally efective but can establish latency and potentially reactivate to cause illness in some immunodefcient vaccinated recipients,raising safety concerns.In this study,34 canine-derived and two feline-derived Bb strains were isolated from 1809 canine and 113 feline nasopharyngeal swab samples collected from eight provinces in China from 2021 to 2023.The PCR results showed that the percentage of positive Bb was 22.94%(441/1922),and more than 90%of the Bb isolates had four virulence factor-encoding genes(VFGs),namely,fhaB,prn,betA and dnt.All the isolated strains displayed a multidrug-resistant phenotype.The virulence of 10 Bb strains isolated from dogs with respiratory symptoms was tested in mice,and we found that eight isolates were highly virulent.Furthermore,the eight Bb isolates with high virulence were inactivated and intramuscularly injected into mice,and three Bb strains(WH1218,WH1203 and WH1224)with the best protective efcacy were selected.Dogs immunized with these three strains exhibited strong protection against challenge with the Bb feld strain WH1218.Ultimately,the WH1218 strain with the greatest protection in dogs was selected as the vaccine candidate.Dogs and cats that received a vaccine containing 109 CFU of the inactivated WH1218 strain showed complete protection against challenge with the Bb feld strain WH1218.This study revealed that Bb is an important pathogen that causes respiratory diseases in domestic dogs and cats in China,and all the isolates exhibited multidrug resistance.The present work contributes to the current understanding of the prevalence,antimicrobial resistance,and virulence genes of Bb in domestic dogs and cats.Additionally,our results suggest that the WH1218 strain is a promising candidate safe and efcacious inactivated Bb vaccine.

Modern approaches for detection of volatile organic compounds in metabolic studies focusing on pathogenic bacteria:Current state of the art

Pathogenic microorganisms produce numerous metabolites,including volatile organic compounds(VOCs).Monitoring these metabolites in biological matrices(e.g.,urine,blood,or breath)can reveal the presence of specific microorganisms,enabling the early diagnosis of infections and the timely implementation of tar-geted therapy.However,complex matrices only contain trace levels of VOCs,and their constituent com-ponents can hinder determination of these compounds.Therefore,modern analytical techniques enabling the non-invasive identification and precise quantification of microbial VOCs are needed.In this paper,we discuss bacterial VOC analysis under in vitro conditions,in animal models and disease diagnosis in humans,including techniques for offline and online analysis in clinical settings.We also consider the advantages and limitations of novel microextraction techniques used to prepare biological samples for VOC analysis,in addition to reviewing current clinical studies on bacterial volatilomes that address inter-species in-teractions,the kinetics of VOC metabolism,and species-and drug-resistance specificity.