Landscape of global urban environmental resistome and its association with local socioeconomic and medical status.

Antimicrobial resistance (AMR) poses a critical threat to global health and development, with environmental factors-particularly in urban areas-contributing significantly to the spread of antibiotic resistance genes (ARGs). However, most research to date has been conducted at a local level, leaving significant gaps in our understanding of the global status of antibiotic resistance in urban environments. To address this issue, we thoroughly analyzed a total of 86,213 ARGs detected within 4,728 metagenome samples, which were collected by the MetaSUB International Consortium involving diverse urban environments in 60 cities of 27 countries, utilizing a deep-learning based methodology. Our findings demonstrated the strong geographical specificity of urban environmental resistome, and their correlation with various local socioeconomic and medical conditions. We also identified distinctive evolutionary patterns of ARG-related biosynthetic gene clusters (BGCs) across different countries, and discovered that the urban environment represents a rich source of novel antibiotics. Our study provides a comprehensive overview of the global urban environmental resistome, and fills a significant gap in our knowledge of large-scale urban antibiotic resistome analysis.

High-throughput screening of small-molecules libraries identified antibacterials against clinically relevant multidrug-resistant A. baumannii and K. pneumoniae.

BACKGROUND: The current pipeline for new antibiotics fails to fully address the significant threat posed by drug-resistant Gram-negative bacteria that have been identified by the World Health Organization (WHO) as a global health priority. New antibacterials acting through novel mechanisms of action are urgently needed. We aimed to identify new chemical entities (NCEs) with activity against Klebsiella pneumoniae and Acinetobacter baumannii that could be developed into a new treatment for drug-resistant infections. METHODS: We developed a high-throughput phenotypic screen and selection cascade for generation of hit compounds active against multidrug-resistant (MDR) strains of K. pneumoniae and A. baumannii. We screened compound libraries selected from the proprietary collections of three pharmaceutical companies that had exited antibacterial drug discovery but continued to accumulate new compounds to their collection. Compounds from two out of three libraries were selected using "eNTRy rules" criteria associated with increased likelihood of intracellular accumulation in Escherichia coli. FINDINGS: We identified 72 compounds with confirmed activity against K. pneumoniae and/or drug-resistant A. baumannii. Two new chemical series with activity against XDR A. baumannii were identified meeting our criteria of potency (EC50 ≤50 µM) and absence of cytotoxicity (HepG2 CC50 ≥100 µM and red blood cell lysis HC50 ≥100 µM). The activity of close analogues of the two chemical series was also determined against A. baumannii clinical isolates. INTERPRETATION: This work provides proof of principle for the screening strategy developed to identify NCEs with antibacterial activity against multidrug-resistant critical priority pathogens such as K. pneumoniae and A. baumannii. The screening and hit selection cascade established here provide an excellent foundation for further screening of new compound libraries to identify high quality starting points for new antibacterial lead generation projects. FUNDING: BMBF and GARDP.

The efficacy of slow-rate ventriculolumbar perfusion chemotherapy for leptomeningeal carcinomatosis: a phase II study.

PURPOSE: This study aimed to evaluate the symptomatic response and side effects of ventriculolumbar perfusion (VLP) methotrexate chemotherapy with a low perfusion rate in patients with leptomeningeal metastasis. METHODS: Patients in a single-arm, two-stage phase II trial based on Simon's minimax design received VLP with a reduced (15 cc/h) perfusion rate with the purpose of decreasing constitutional side effects such as nausea/vomiting, insomnia, and confusion. The primary outcome was control of increased intracranial pressure (ICP). The secondary outcome was an occurrence of side effects. The results were compared with those of a previous trial of VLP with a 20-cc/h perfusion rate. RESULTS: Total 90 patients were enrolled. Out of 65 patients with increased ICP, 32 achieved normalized ICP after VLP chemotherapy (bias-adjusted response rate = 51%). The incidence of moderate-to-severe nausea/vomiting was reduced to 46% from 64% in the previous study, and that of sleep disturbance was increased to 13% from 9%, but both failed to reach statistical significance. The incidence of moderate-to-severe confusion was significantly reduced to 12% from 23% in the previous study (p = 0.04). Median overall survival was better among patients with controlled ICP than among those who remained with increased ICP (193 days vs. 94 days, p = 0.013). CONCLUSION: Compared with a higher perfusion rate, the low perfusion rate failed to provide non-inferior ICP control or improved side effects, except for confusion. The relationship between VLP perfusion rate and ICP control needs to be evaluated in future trials adjusting for bias from uncompleted protocol due to poor general condition.

A portable smartphone-based colorimetric sensor that utilizes dual amplification for the on-site detection of airborne bacteria.

Over the past few years, infections caused by airborne pathogens have spread worldwide, infecting several people and becoming an increasingly severe threat to public health. Therefore, there is an urgent need for developing airborne pathogen monitoring technology for use in confined environments to enable epidemic prevention. In this study, we designed a colorimetry-based bacterial detection platform that uses a clustered regularly interspaced short palindromic repeat-associated protein 12a system to amplify signals and a urease enzyme to induce color changes. Furthermore, we have developed a smartphone application that can distinguish colors under different illumination conditions based on the HSV model and detect three types of disease-causing bacteria. Even synthetic oligomers of a few picomoles of concentration and genomic DNA of airborne bacteria smaller than several nanograms can be detected with the naked eye and using color analysis systems. Furthermore, in the air capture model system, the bacterial sample generated approximately a 2-fold signal difference compared with that in the control group. This colorimetric detection method can be widely applied for public safety because it is easy to use and does not require complex equipment.

AcrAB-TolC, a major efflux pump in Gram negative bacteria: toward understanding its operation mechanism.

Antibiotic resistance (AR) is a silent pandemic that kills millions worldwide. Although the development of new therapeutic agents against antibiotic resistance is in urgent demand, this has presented a great challenge, especially for Gram-negative bacteria that have inherent drug-resistance mediated by impermeable outer membranes and multidrug efflux pumps that actively extrude various drugs from the bacteria. For the last two decades, multidrug efflux pumps, including AcrAB-TolC, the most clinically important efflux pump in Gram-negative bacteria, have drawn great attention as strategic targets for re-sensitizing bacteria to the existing antibiotics. This article aims to provide a concise overview of the AcrAB-TolC operational mechanism, reviewing its architecture and substrate specificity, as well as the recent development of AcrAB-TolC inhibitors. [BMB Reports 2023; 56(6): 326-334].

PTD-FGF2 Attenuates Elastase Induced Emphysema in Mice and Alveolar Epithelial Cell Injury.

Aberrant communication in alveolar epithelium is a major feature of inflammatory response for the airway remodeling leading to chronic obstructive pulmonary disease (COPD). In this study, we investigated the effect of protein transduction domains (PTD) conjugated Basic Fibroblast Growth Factor (FGF2) (PTD-FGF2) in response to cigarette smoke extract (CSE) in MLE-12 cells and porcine pancreatic elastase (PPE)-induced emphysematous mice. When PPE-induced mice were intraperitoneally treated with 0.1-0.5 mg/kg PTD-FGF2 or FGF2, the linear intercept, infiltration of inflammatory cells into alveoli and pro-inflammatory cytokines were significantly decreased. In western blot analysis, phosphorylated protein levels of c-Jun N-terminal Kinase 1/2 (JNK1/2), extracellular signal-regulated kinase (ERK1/2) and p38 mitogen-activated protein kinases (MAPK) were decreased in PPE-induced mice treated PTD-FGF2. In MLE-12 cells, PTD-FGF2 treatment decreased reactive oxygen species (ROS) production and further decreased Interleukin-6 (IL-6) and IL-1b cytokines in response to CSE. In addition, phosphorylated protein levels of ERK1/2, JNK1/2 and p38 MAPK were reduced. We next determined microRNA expression in the isolated exosomes of MLE-12 cells. In reverse transcription-polymerase chain reaction (RT-PCR) analysis, level of let-7c miRNA was significantly increased while levels of miR-9 and miR-155 were decreased in response to CSE. These data suggest that PTD-FGF2 treatment plays a protective role in regulation of let-7c, miR-9 and miR-155 miRNA expressions and MAPK signaling pathways in CSE-induced MLE-12 cells and PPE-induced emphysematous mice.

Conductive Thread-Based Immunosensor for Pandemic Influenza A (H1N1) Virus Detection.

Infectious agents such as viruses pose significant threats to human health, being transmitted via direct contact as well as airborne transmission without direct contact, thus requiring rapid detection to prevent the spread of infectious diseases. In this study, we developed a conductive thread-based immunosensor (CT-IS), a biosensor to easily detect the presence of airborne viruses. CT-IS utilizes an antibody that specifically recognizes the HA protein of the pandemic influenza A (pH1N1) virus, which is incorporated into the conductive thread. The antigen-antibody interaction results in increased strain on the conductive thread in the presence of the pH1N1 virus, resulting in increased electrical resistance of the CT-IS. We evaluated the performance of this sensor using the HA protein and the pH1N1 virus, in addition to samples from patients infected with the pH1N1 virus. We observed a significant change in resistance in the pH1N1-infected patient samples (positive: n = 11, negative: n = 9), whereas negligible change was observed in the control samples (patients not infected with the pH1N1 virus; negative). Hence, the CT-IS is a lightweight fiber-type sensor that can be used as a wearable biosensor by combining it with textiles, to detect the pH1N1 virus in a person's vicinity.

Polydiacetylene-based hydrogel beads as colorimetric sensors for the detection of biogenic amines in spoiled meat.

Ingesting large quantities of biogenic amines (BAs), which are released from spoiled foods, can have adverse side effects on the human body. Herein, we developed a colorimetric sensor using polydiacetylene (PDA)-based hydrogel beads that change color upon binding with BAs, thereby conveniently checking whether food is spoiled due to improper storage and distribution. The colorimetric sensor is fabricated by mixing PDA liposomes with an alginate solution. PDA undergoes a color change from blue to red when exposed to various external stimuli. In addition, alginate bestows the hydrogel with a three-dimensional porous structure, affording a large surface area. The PDA-based hydrogel beads can visually confirm the presence of BAs in solution or vapor form. Cadaverine and propylamine were rapidly detected with distinct color changes in the solution and vapor phases, respectively. The spoilage of pork meat at room temperature could be detected after two days as a 40.84% red chromatic shift.

A simple protein histidine kinase activity assay for high-throughput inhibitor screening.

Bacterial two-component systems (TCSs), which typically consist of a sensor histidine kinase (HK) and a response regulator (RR), have been investigated as attractive antibacterial drug targets. Unfortunately, current HK activity assays based on the quantification of autophosphorylated HKs are hampered by the instability of the phosphohistidine (pHis) product, rendering them ill-suited for high-throughput screenings. To address this challenge, we developed a simple HK activity assay using readily available reagents, which we have termed AUDECY (AUtophosphorylation-DEphosphorylation CYcle assay). Instead of trying to preserve the fragile pHis, we deliberately decomposed it with a pHis-specific phosphatase to constitute an ATPase-like cycle for convenient colorimetric measurements. This kinetic assay was successfully employed for the kinetic characterization of E. coli EnvZ and for high-throughput inhibitor screening of vancomycin-resistant Enterococcus faecium (VRE) VanS, of which histidine kinase activity was hardly detectable with conventional methods. Through the screening, we identified OSU-03012, a potent VanS HK inhibitor, which sensitized VRE toward vancomycin, highlighting the potential of AUDECY in HK inhibitor discovery.

Saliency guided data augmentation strategy for maximally utilizing an object's visual information.

Among the various types of data augmentation strategies, the mixup-based approach has been particularly studied. However, in existing mixup-based approaches, object loss and label mismatching can occur if random patches are utilized when constructing augmented images, and additionally, patches that do not contain objects might be included, which degrades performance. In this paper, we propose a novel augmentation method that mixes patches in a non-overlapping manner after they are extracted from the salient regions in an image. The suggested method can make effective use of object characteristics, because the constructed image consists only of visually important regions and is robust to noise. Since the patches do not occlude each other, the semantically meaningful information in the salient regions can be fully utilized. Additionally, our method is more robust to adversarial attack than the conventional augmentation method. In the experimental results, when Wide ResNet was trained on the public datasets, CIFAR-10, CIFAR-100 and STL-10, the top-1 accuracy was 97.26%, 83.99% and 82.40% respectively, which surpasses other augmentation methods.

Metagenomic characterization of bacterial community and antibiotic resistance genes found in the mass transit system in Seoul, South Korea.

Mass transit systems, including subways and buses, are useful environments for studying the urban microbiome, as the vast majority of populations in urban areas use public transportation. Microbial communities in urban environments include both human- and environment-associated bacteria that play roles in health and pathogen transmission. In this study, we used shotgun metagenomic sequencing to profile microbial communities sampled from various surfaces found in subway stations and bus stops within the Seoul mass transit system. The metagenomic approach and network analysis were used to investigate broad-spectrum antibiotic resistance genes (ARGs) and their co-occurrence patterns. We uncovered 598 bacterial species in 76 samples collected from various surfaces within the Seoul mass transit system. All samples were dominated by the potential human pathogen Salmonella enterica (40 %) and the human skin bacterium Cutibacterium acnes (19 %). Significantly abundant biomarkers detected in subway station samples were associated with bacteria typically found in the human oral cavity and respiratory tract, whereas biomarkers detected in bus stop samples were associated with bacteria commonly found in soil, water, and plants. Temperature and location had significant effects on microbial community structure and diversity. In total, 41 unique ARG subtypes were identified, associated with single-drug or multidrug resistance to clinically important and extensively used antibiotics, including aminoglycosides, carbapenem, glycopeptide, and sulfonamides. We revealed that Seoul subway stations and bus stops possess unique microbiomes containing potential human pathogens and ARGs. These findings provide insights for refining location-specific responses to reduce exposure to potentially causative agents of infectious diseases, improving public health.

Pharmacological perturbation of thiamine metabolism sensitizes Pseudomonas aeruginosa to multiple antibacterial agents.

New therapeutic concepts are critically needed for carbapenem-resistant Pseudomonas aeruginosa, an opportunistic pathogen particularly recalcitrant to antibiotics. The screening of around 230,000 small molecules yielded a very low hit rate of 0.002% after triaging for known antibiotics. The only novel hit that stood out was the antimetabolite oxythiamine. Oxythiamine is a known transketolase inhibitor in eukaryotic cells, but its antibacterial potency has not been reported. Metabolic and transcriptomic analyses indicated that oxythiamine is intracellularly converted to oxythiamine pyrophosphate and subsequently inhibits several vitamin-B1-dependent enzymes, sensitizing the bacteria to several antibiotic and non-antibiotic drugs such as tetracyclines, 5-fluorouracil, and auranofin. The positive interaction between 5-fluorouracil and oxythiamine was confirmed in a murine ocular infection model, indicating relevance during infection. Together, this study revealed a system-level significance of thiamine metabolism perturbation that sensitizes P. aeruginosa to multiple small molecules, a property that could inform on the development of a rational drug combination.

Isothermal amplification-mediated lateral flow biosensors for in vitro diagnosis of gastric cancer-related microRNAs.

MicroRNAs (miRNAs) are important diagnostic and prognostic biomarkers for various tumors. Currently, many diagnostic systems have been developed to detect miRNAs, but simple techniques for detecting miRNAs are still required. Recently, we reported that the expression of miRNA-135b is upregulated in gastric epithelial cells during gastric inflammation and carcinogenesis. Our aim was to develop an in vitro diagnostic platform to analyze the expression of gastric cancer-related biomarkers in the blood. The diagnostic platform comprised an isothermal amplification-based lateral flow biosensor (IA-LFB) that enables easy diagnosis of gastric cancer through visual observation. In this platform, trace amounts of biomarkers are isothermally amplified through rolling circle amplification (RCA), and the amplified product is grafted to the LFB. The performance of the IA-LFB was confirmed using RNAs extracted from in vitro and in vivo models. The platform could detect target miRNAs within 3 h with excellent sensitivity and selectivity. In particular, the IA-LFB could detect the overexpression of gastric cancer-related markers (miRNA-135b and miRNA-21) in RNAs extracted from the blood of patients with various stages (stages 1-4) of gastric cancer compared to that in healthy volunteers. Therefore, IA-LFB is a simple and sensitive in vitro diagnostic system for detecting gastric cancer-related biomarkers and can contribute to the early diagnosis and prognosis monitoring of gastric cancer. Furthermore, this technology can be applied to systems that can detect multiple biomarkers related to various diseases (such as infectious and genetic diseases).

miRNA sensing hydrogels capable of self-signal amplification for early diagnosis of Alzheimer's disease.

Alzheimer's disease (AD), one of the leading senile disorders in the world, causes severe memory loss and cognitive impairment. To date, there is no clear cure for AD. However, early diagnosis and monitoring can help mitigate the effects of this disease. In this study, we reported a platform for diagnosing early-stage AD using microRNAs (miRNAs) in the blood as biomarkers. First, we selected an appropriate target miRNA (miR-574-5p) using AD model mice (4-month-old 5XFAD mice) and developed a hydrogel-based sensor that enabled high-sensitivity detection of the target miRNA. This hydrogel contained catalytic hairpin assembly (CHA) reaction-based probes, leading to fluorescence signal amplification without enzymes and temperature changes, at room temperature. This sensor exhibited high sensitivity and selectivity, as evidenced by its picomolar-level detection limit (limit of detection: 1.29 pM). Additionally, this sensor was evaluated using the plasma of AD patients and non-AD control to validate its clinical applicability. Finally, to use this sensor as a point-of-care-testing (POCT) diagnostic system, a portable fluorometer was developed and verified for feasibility of application.

FRET-based hACE2 receptor mimic peptide conjugated nanoprobe for simple detection of SARS-CoV-2.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has led to a pandemic of acute respiratory disease, namely coronavirus disease (COVID-19). This disease threatens human health and public safety. Early diagnosis, isolation, and prevention are important to suppress the outbreak of COVID 19 given the lack of specific antiviral drugs to treat this disease and the emergence of various variants of the virus that cause breakthrough infections even after vaccine administration. Simple and prompt testing is paramount to preventing further spread of the virus. However, current testing methods, namely RT-PCR, is time-consuming. Binding of the SARS-CoV-2 spike (S) glycoprotein to human angiotensin-converting enzyme 2 (hACE2) receptor plays a pivotal role in host cell entry. In the present study, we developed a hACE2 mimic peptide beacon (COVID19-PEB) for simple detection of SARS-CoV-2 using a fluorescence resonance energy transfer system. COVID19-PEB exhibits minimal fluorescence in its ''closed'' hairpin structure; however, in the presence of SARS-CoV-2, the specific recognition of the S protein receptor-binding domain by COVID19-PEB causes the beacon to assume an ''open'' structure that emits strong fluorescence. COVID19-PEB can detect SARS-CoV-2 within 3 h or even 50 min and exhibits strong fluorescence even at low viral concentrations, with a detection limit of 4 × 103 plaque-forming unit/test. Furthermore, in SARS-CoV-2-infected patient samples confirmed using polymerase chain reaction, COVID19-PEB accurately detected the virus. COVID19-PEB could be developed as a rapid and accurate diagnostic tool for COVID-19.

Metabolite profiling of a Sargassum micracanthum methanol extract with in vitro efficacy against human head and neck squamous cell carcinoma aggressiveness.

OBJECTIVE: Extracts from the brown algae Sargassum micracanthum have documented anti-viral, anti-oxidant, and anti-inflammatory activities as well as potential anti-tumor efficacy against several cancer types. Here, we evaluated the inhibitory effect and molecular mechanisms of methanol extract of S. micracanthum (MESM) on the aggressiveness of human head and neck squamous cell carcinoma (HNSCC) using in vitro cell culture-based models. DESIGN: To test the potential efficacy of MESM on the migratory and invasive properties of HNSCC cells, we used wound healing, transwell cell migration and invasion assays. Proteome profiling and functional in silico analysis were applied to investigate the possible modes of action by MESM. We also examined the metabolite profiling of MESM using gas chromatography/mass spectrometry. RESULTS: MESM inhibited the motility of human HNSCC cell lines as well as invasiveness without influencing cell survival. Proteome profiling identified 19 oncogenic proteins significantly downregulated by MESM treatment. Protein-protein interaction network and gene ontology analyses revealed that Tie2 and associated angiogenic signaling pathway components were significantly enriched among these downregulated oncogenic proteins, which was confirmed by validating the reduced Tie2 expression in MESM treatment groups. Metabolite profiling of MESM identified six-carbon sugar alcohols such as D-sorbitol and/or D-mannitol as the main bioactive compounds. D-sorbitol and D-mannitol effectively reduced Tie2 expression and the aggressiveness of human HNSCC cell lines. CONCLUSIONS: These findings suggest that six-carbon sugar alcohols in MESM have promising anti-cancer efficacy for the treatment of human HNSCC and further identify Tie2 signaling components as potential treatment targets.

Electrospun Nanofibrous Membrane-Based Colorimetric Device for Rapid and Simple Screening of Amphetamine-Type Stimulants in Drinks.

With the growth of drug-facilitated crimes, prevention has become increasingly important. Although various drug detection technologies exist, most focus on postconsumption detection. However, the prevention of drug-facilitated crimes requires technology for the quick and easy detection of amphetamine-type stimulants (ATSs) before ingestion. Herein, drug screening kits (DSKs) were developed for the simple detection of ATSs in drinks. The DSKs consisted of polydiacetylene nanofiber-based paper sensors fabricated by electrospinning with 10,12-pentacosadiynoic acid (PCDA) and PCDA-dopamine as sensing materials that can bind ATSs via hydrogen bonding and π-π interactions. Dropping a drink on the DSK provided an immediate visual indication of the presence of ATSs. When ATSs were present in the drink, the color of the DSK clearly changed from blue to red, with the increase in red intensity being more than twofold greater than that observed when water alone was tested. Notably, the result could be confirmed by the naked eye without any analytical instrumentation. A color change indicating the presence of ATSs was successfully observed in various alcoholic and nonalcoholic drinks. These results indicate the potential of DSKs for preventing drug-facilitated crimes caused by unwanted drug intake.

An RNA Interference (RNAi) Toolkit and Its Utility for Functional Genetic Analysis of Leishmania (Viannia).

RNA interference (RNAi) is a powerful tool whose efficacy against a broad range of targets enables functional genetic tests individually or systematically. However, the RNAi pathway has been lost in evolution by a variety of eukaryotes including most Leishmania sp. RNAi was retained in species of the Leishmania subgenus Viannia, and here we describe the development, optimization, and application of RNAi tools to the study of L. (Viannia) braziliensis (Lbr). We developed vectors facilitating generation of long-hairpin or "stem-loop" (StL) RNAi knockdown constructs, using GatewayTM site-specific recombinase technology. A survey of applications of RNAi in L. braziliensis included genes interspersed within multigene tandem arrays such as quinonoid dihydropteridine reductase (QDPR), a potential target or modulator of antifolate sensitivity. Other tests include genes involved in cell differentiation and amastigote proliferation (A600), and essential genes of the intraflagellar transport (IFT) pathway. We tested a range of stem lengths targeting the L. braziliensis hypoxanthine-guanine phosphoribosyl transferase (HGPRT) and reporter firefly luciferase (LUC) genes and found that the efficacy of RNAi increased with stem length, and fell off greatly below about 128 nt. We used the StL length dependency to establish a useful 'hypomorphic' approach not possible with other gene ablation strategies, with shorter IFT140 stems yielding viable cells with compromised flagellar morphology. We showed that co-selection for RNAi against adenine phosphoryl transferase (APRT1) using 4-aminopyrazolpyrimidine (APP) could increase the efficacy of RNAi against reporter constructs, a finding that may facilitate improvements in future work. Thus, for many genes, RNAi provides a useful tool for studying Leishmania gene function with some unique advantages.

Microfluidic device for one-step detection of breast cancer-derived exosomal mRNA in blood using signal-amplifiable 3D nanostructure.

Metastasis attributed to approximately 90% of cancer-related deaths; hence, the detection of metastatic tumor-derived components in the blood assists in determining cancer recurrence and patient survival. Microfluidic-based sensors facilitate analysis of small fluid volumes and represent an accurate, rapid, and user-friendly method of field diagnoses. In this study, we have developed a microfluidic chip-based exosomal mRNA sensor (exoNA-sensing chip) for the one-step detection of exosomal ERBB2 in the blood by integrating a microfluidic chip and 3D-nanostructured hydrogels. The exoNA-sensing chip is a vacuum-driven power-free microfluidic chip that can accurately control the flow of trace fluids (<100 µL). The sensing part of the exoNA-sensing chip includes 3D-nanostructured hydrogels capable of detecting ERBB2 and a reference gene by amplifying a fluorescent signal via an enzyme-free catalytic hairpin assembly reaction at room temperature. This hydrogel offers a detection limit of 58.3 fM with good selectivity for target sequences. The performance of the exoNA-sensing chip was evaluated by testing in vitro and in vivo samples and was proven to be effective for cancer diagnosis and liquid biopsies.

Annotating unknown species of urban microorganisms on a global scale unveils novel functional diversity and local environment association.

In urban ecosystems, microbes play a key role in maintaining major ecological functions that directly support human health and city life. However, the knowledge about the species composition and functions involved in urban environments is still limited, which is largely due to the lack of reference genomes in metagenomic studies comprises more than half of unclassified reads. Here we uncovered 732 novel bacterial species from 4728 samples collected from various common surface with the matching materials in the mass transit system across 60 cities by the MetaSUB Consortium. The number of novel species is significantly and positively correlated with the city population, and more novel species can be identified in the skin-associated samples. The in-depth analysis of the new gene catalog showed that the functional terms have a significant geographical distinguishability. Moreover, we revealed that more biosynthetic gene clusters (BGCs) can be found in novel species. The co-occurrence relationship between BGCs and genera and the geographical specificity of BGCs can also provide us more information for the synthesis pathways of natural products. Expanded the known urban microbiome diversity and suggested additional mechanisms for taxonomic and functional characterization of the urban microbiome. Considering the great impact of urban microbiomes on human life, our study can also facilitate the microbial interaction analysis between human and urban environment.