Members of the interferon regulatory factor (IRF) gene family are crucial regulators of type I interferon signaling, which may play a role in the resistance of glioma to immune checkpoint blockade. However, the expression profiles, potential functions, and clinical significance of IRF family members remain largely unknown. Here, we examined IRF transcript levels and clinicopathological data from glioma patients using several bioinformatic databases, including ONCOMINE, GEPIA, TCGA, and cBioPortal. We found that IRF1, IRF2, IRF5, IRF8 and IRF9 were significantly upregulated in glioma compared to normal brain tissue. Higher IRF1, IRF2, IRF3, IRF4, IRF5, IRF7, IRF8 and IRF9 mRNA levels correlated with more advanced tumor grades and poorer outcomes. Moreover, although IRFs mutation rates were low (ranging from 0.5% to 2.3%) in glioma patients, genetic alterations in IRFs were associated with more favorable patient survival. Functional analysis showed that IRFs participated in glioma pathology mainly through multiple inflammation- and immunity-related pathways. Additionally, correlations were identified between IRFs and infiltration of immune cells within glioma tissues. Collectively, these results indicate that IRF family members, including IRF1, IRF2, IRF5, IRF8 and IRF9, may serve as prognostic biomarkers and indicators of immune status in glioma patients. Glioma is the most prevalent primary malignancy in the human brain and is characterized by high recurrence and lethality rates . AccordiRecent reports suggest that type I interferon signaling, which is regulated by interferon regulatory factors (IRFs), plays an important role in glioma resistance to immune checkpoint blockade , 6. The Recent advances in gene sequencing technology have enabled comprehensive analysis of IRF family members with existing bioinformatic tools. In this study, we performed an in-depth exploration of the expression patterns of IRF family members in glioma and evaluated their potential as prognostic biomarkers with the goal of improving molecular diagnosis and prognostic prediction for glioma patients.p = 0.008); two additional studies by Liang and Bredel found that IRF1 expression was increased 2.225- and 2.151-fold, respectively, in glioblastoma. IRF2 transcript levels were also higher in glioblastoma than normal brain tissues in two datasets from TCGA . The results of Sun’s study suggested that IRF5 was increased 2.125-fold in diffuse astrocytoma (p = 1.33E-4) and 2.180-fold in anaplastic astrocytoma (p = 3.20E-5). Moreover, studies by Lee, Ramaswamy, and Bredel all found that IRF8 and IRF9 levels were significantly increased in glioblastoma or anaplastic oligoastrocytoma , IRF2 (p = 6.80E-18), IRF3 (p = 1.70E-23), IRF5 (p = 2.30E-08), IRF7 (p = 1.90E-29), IRF8 (p = 0.029), IRF9 (p = 4.80E-05) expression and pathological grade (p = 0.064). No correlation between IRF6 expression and pathological grade was observed (p = 0.49). For all of the identified correlations, expression increased as tumors progressed, suggesting that IRFs may play a role in glioma tumorigenesis and progression.Relationships between IRF family member expression and clinicopathological parameters of glioma patients were examined using data from the TCGA database. Among the 260 grade II, 267 grade III, and 173 grade IV glioma patients, significant correlations were observed between IRF1 , IRF2 (p = 2.22E-16), IRF3 (p = 1.43E-15), IRF4 (p = 0.004), IRF5 p = 5.84E-12), IRF7 (p = 6.85E-28), IRF8 (p = 0.001), and IRF9 (p = 0.000) transcript levels had significantly longer overall survival times   to 20.48% (17/83); mutations, deep deletions, and amplification were the most common types of alteration . For eac.237E-6) .We next explored potential co-expression among IRF family genes using data from TCGA glioma dataset. Pearson's correlation results revealed significant positive correlations between the following IRFs: IRF1 with IRF2 (r = 0.48) and IRF7 (r = 0.52); IRF2 with IRF1, IRF5 (r = 0.54), and IRF8 (r = 0.43); IRF3 with IRF7 (r = 0.41); IRF5 with IRF7 (r = 0.41) and IRF8 (R = 0.73); IRF7 with IRF9 (r = 0.74) . Little p < 1.0E-16; Next, we conducted a network analysis to examine potential internal interactions among IRF family genes as well as external interactions with other functionally related genes. PPI network analysis using STRING software revealed close protein-protein associations among the IRF family genes with 9 nodes, 32 edges, and an average node degree of 7.11  and 11 KEGG items were enriched. p < 0.01). Similar results were obtained for GBM. In addition, IRF1, IRF2, IRF6, and IRF9 were positively correlated with CD8+ T cell infiltration in LGG. However, IRF1, IRF5, IRF6, IRF7, IRF8, and IRF9 expression were negatively correlated with CD8+ T cell infiltration in GBM.Since the IRF family may regulate glioma progression and prognosis by participating in a wide range of inflammatory and immune responses, we undertook a comprehensive analysis of tumor immune infiltrates using the TIMER database. The results are shown in p = 0.002), CD8+ T cells (p = 0.042), IRF1 expression (p = 0.001), and IRF8 expression (p = 0.000) were significantly associated with the prognosis of LGG patients, while CD4+ T cells, dendritic cells (p =0.002), IRF1 expression (p = 0.01), IRF7 expression (p = 0.007), and IRF8 expression (p = 0.028) were associated with the prognosis of GBM patients , another online tool that contains RNA sequence expression data from 518 LGG samples, 163 GBM samples, and 207 normal brain samples [t-tests; p < 0.05 and fold change >2 were considered significant.We used a two-step analysis to assess IRF family member expression patterns in glioma patients. First, we examined mRNA level data from ONCOMINE, the largest public microarray database for genome-wide expression analysis . Data we samples . IRF expp-value <0.05 was considered significant.Correlations between IRF family member expression and clinicopathological characteristics and prognosis in glioma patients were evaluated using data derived from The Cancer Genome Atlas (TCGA) database, which contains both sequencing and clinical records for over 30 types of human cancers . A totalThe molecular characteristics and internal/external interactions of IRF family members were explored with multiple tools. Genetic alterations and their associations with patient prognosis were evaluated using cBioPortal, an online tool for visualization and analysis of multidimensional cancer genomics data . Co-exprp < 0.05. Additionally, associations between IRF family members and the activity of cancer pathways, including TSC/mTOR, RTK, RAS/MAPK, PI3K/AKT, hormone ER, hormone AR, EMT, DNA damage response, cell cycle, and apoptosis pathways, were explored with GSCALite, a web-based platform for gene set cancer analysis [To examine biological functions, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were conducted for IRF family members and functionally related genes using DAVID software (version 6.8) . Biologianalysis .The TIMER (Tumor Immune Estimation Resource) database is an immune infiltrate analysis tool for systematic evaluation of the different immune cells that infiltrate tumor tissue and their clinical significance . In our


In the SIDERO-CR-2014–2016 surveillance study, European clinical isolates comprising carbapenem-non-susceptible (CarbNS) Enterobacterales and MDR non-fermenters were tested against cefiderocol and comparators.Many carbapenem-resistant (CR) Gram-negative (GN) pathogens exhibit MDR, meaning few therapeutic options are available for CR-GN infections. Cefiderocol, a siderophore cephalosporin, has demonstrated Cefiderocol MICs were determined using iron-depleted CAMHB, and comparators using CAMHB, per recommended CLSI methodology. Carbapenemase gene profiles were determined using PCR.N = 870) from 23 European countries comprised CarbNS Enterobacterales (n = 457), MDR Pseudomonas aeruginosa (n = 177) and MDR Acinetobacter baumannii (n = 236). The most common carbapenemases were KPC (52%), OXA-48-like (19%), VIM (14%) and NDM (8%) in Enterobacterales, VIM (41%) in P. aeruginosa and OXA-23-like (57%) and OXA-24/40-like (37%) in A. baumannii. Most carbapenemase-producing isolates (65%) co-carried ESBLs. Approximately half of P. aeruginosa isolates were negative for carbapenemases, compared with 10% of Enterobacterales and 3% of A. baumannii. A similar proportion of Enterobacterales were susceptible to cefiderocol  compared with comparator antimicrobial agents, including colistin  and ceftazidime/avibactam . Of P. aeruginosa isolates, 98.3% were susceptible to cefiderocol (100% of VIM producers), similar to colistin (100%). Against A. baumannii, 94.9% had cefiderocol MIC ≤2 mg/L and 93.6% of isolates were susceptible to colistin.Isolates (Cefiderocol demonstrated potent activity against CarbNS and MDR GN bacteria, including non-fermenters and a wide variety of MBL- and serine-β-lactamase-producing strains. Globally, the incidence of carbapenem-resistant (CR) GN bacteria is increasing,Cefiderocol is a novel siderophore cephalosporin, which was developed for the treatment of MDR GN bacteria, including those resistant to carbapenems. Cefiderocol has recently been approved in Europe for the treatment of infections due to aerobic GN organisms in adults with limited treatment options,,,Escherichia coli and Klebsiella pneumoniae, as well as meropenem-resistant Pseudomonas aeruginosa and Acinetobacter baumannii.The structure of cefiderocol is based around a cephalosporin backbone with the addition of a catechol moiety at the 3-position side chain. The cephalosporin core enables cefiderocol to act like other cephalosporins, binding primarily to PBPs and killing bacterial cells by inhibition of peptidoglycan cell wall biosynthesis. Cefiderocol differs from other cephalosporins in that the catechol moiety chelates ferric (Fe-III) iron, mimicking natural siderophores, allowing cefiderocol to exploit the bacteria’s own active receptor-mediated iron transport system to cross the outer membrane.in vitro conditions are essential in order to mimic the hypoferremic conditions encountered by bacteria in the human body during infection.,,in vivo efficacySusceptibility testing of cefiderocol by broth microdilution requires an iron-depleted medium to promote the natural production of siderophores by bacterial cells. Iron-depleted In the SIDERO-CR surveillance study, CR and MDR clinical isolates of GN bacteria collected from patients between 2014 and 2016 were tested against cefiderocol and comparators using recommended CLSI broth microdilution methodology.JAC-AMR Online).,Full methodology for the SIDERO-CR study and molecular characterization using PCR has been published previously (see n = 457), defined as having a meropenem MIC of ≥2 mg/L. Isolates of P. aeruginosa (n = 177) and A. baumannii (n = 236) were included if they demonstrated an amikacin-resistant (MIC ≥32 mg/L), ciprofloxacin-resistant (MIC ≥4 mg/L) and imipenem-resistant (MIC ≥16 mg/L) MDR phenotype.Included in the SIDERO-CR-2014–2016 European test set were carbapenem-non-susceptible (CarbNS) phenotypes of Enterobacterales isolates (A. baumannii (CLSI breakpoint ≤8 mg/L),,P. aeruginosa are considered susceptible to cefiderocol at MIC ≤2 mg/L (resistant >2 mg/L).MICs were determined for cefiderocol, cefepime, ceftazidime/avibactam, ceftolozane/tazobactam, ciprofloxacin, colistin and meropenem by broth microdilution according to CLSI guidelines.in vitro samples were anonymized.Ethics approval was not required as all N = 870) were from Italy [217 (24.9%)], Greece [128 (14.7%)] and Russia [91 (10.5%)].The majority of SIDERO-CR-2014–2016 European isolates (A. baumannii [135/385 (35.1%)] and P. aeruginosa (106/385 [27.5%]). K. pneumoniae (120/385 [31.2%]) was the most common Enterobacterales species in RTIs , followed by intra-abdominal infections , surgical site infections  and bloodstream infections . K. pneumoniae was the most common pathogen in UTIs [59/157 (37.6%)], SSIs [57/89 (64.0%)] and BSIs [47/85 (55.3%)] while A. baumannii was the most common in IAIs [46/125 (36.8%)].Respiratory tract infections (RTIs) were the most common isolate source [385 (44.3%)]; the majority were non-fermenters [241/385 (62.6%)], consisting of Is Table . Urinaryin vitro activity against a variety of CarbNS-GN bacteria in SIDERO-CR-2014–2016 European isolates. Overall, 772/870 (88.7%) had a cefiderocol MIC of ≤2 mg/L; 547/634 (86.3%) of Enterobacterales and P. aeruginosa isolates were susceptible to cefiderocol and 224/236 (94.9%) of A. baumannii isolates had a cefiderocol MIC of ≤2 mg/L.Cefiderocol exhibited 90 = 4 mg/L) (TableK. pneumoniae isolates (n = 332), 82.8% were susceptible to cefiderocol, while 71.7% were susceptible to colistin and 88.9% to ceftazidime/avibactam.Of CarbNS Enterobacterales, 81.6% were cefiderocol susceptible (MIC/L) Table. The pro90 was 1 mg/L against MDR P. aeruginosa and 98.3% of isolates were cefiderocol susceptible (MIC ≤2 mg/L). Colistin demonstrated a similar level of activity (100% susceptible) to cefiderocol, while other comparators were active against <35% of isolates.The cefiderocol MICA. baumannii, the cefiderocol MIC90 was 1 mg/L and 94.9% of isolates had a cefiderocol MIC of ≤2 mg/L. Of the comparators, only colistin demonstrated activity (93.6% susceptible). The MIC ranges for cefiderocol were 0.004 − 8 mg/L for P. aeruginosa and 0.03 − 32 mg/L for K. pneumoniae, while the range against A. baumannii was somewhat wider (0.015 to >64 mg/L).Against MDR P. aeruginosa isolates non-susceptible to cefiderocol (MIC >2 mg/L) was 13.7% (87/634); these isolates consisted mainly of CarbNS K. pneumoniae (n = 57) and Enterobacter cloacae (n = 18) , Italy (n = 2), Denmark (n = 1), Portugal (n = 1) and Turkey (n = 1)].In total, the proportion of Enterobacterales and ) Figure . There wn = 107; cefiderocol MIC90 = 4 mg/L; range: 0.06 − 32 mg/L), as did 97.8% of ceftolozane/tazobactam-resistant P. aeruginosa  and all colistin-resistant isolates  , 76.9% were cefiderocol susceptible; all 15 colistin-resistant A. baumannii isolates had cefiderocol MICs of ≤2 mg/L.Notably, 66.4% of ceftazidime/avibactam-resistant Enterobacterales remained susceptible to cefiderocol (L) Figure. Of coliP. aeruginosa isolates, VIM (41.2%) was the most common carbapenemase. OXA-23-like (57.2%) and OXA-24/40-like (37.3%) producers accounted for the majority of A. baumannii isolates. Most isolates produced multiple β-lactamases, e.g. 34/37 (91.9%) NDM-1 Enterobacterales also harboured ESBLs and OSBLs. A proportion of all isolates, phenotypically non-susceptible to meropenem, were negative for carbapenemase genes, with a greater proportion of P. aeruginosa (49.7%) being carbapenemase negative compared with Enterobacterales (9.8%) and A. baumannii (3.4%).In total, 28 subclasses of carbapenemase were identified, along with 37 ESBL (e.g. CTX-M) and older-spectrum β-lactamase  subclasses,n = 157) and Greece (n = 97) provided the most CarbNS Enterobacterales isolates. Most of these isolates produced KPC  or VIM ; however, NDM was less prevalent in Italy (0.6%) than in Greece (5.2%). OXA-48-like carbapenemases ]. NDM-producing P. aeruginosa isolates were only apparent in Serbia, representing 6/10 samples.In countries providing ≥8 isolates, the majority of carbapenemase-producing MDR A. baumannii isolates from countries with ≥8 available isolates established that the majority produced OXA-23-like carbapenemases,Analysis of MDR 90 values were ≤4 mg/L for all carbapenemases across all strains where ≥10 isolates were available for testing  and NDM  MBL producers. Isolates producing OXA-48-like (n = 85), OXA-23-like (n = 135) and OXA-24/40-like (n = 88) carbapenemases had cefiderocol MIC90 values of ≤4 mg/L.Cefiderocol MICing Table. Across 90 of KPC-producing isolates was 4 mg/L, with 83.6% being cefiderocol susceptible at the EUCAST breakpoint of ≤2 mg/L, but with 98.3% being cefiderocol susceptible at the CLSI breakpoint of ≤4 mg/L. Ceftazidime/avibactam demonstrated potent activity against 97.9% of KPC-producing Enterobacterales by both EUCAST and CLSI breakpoints. Similarly, against OXA-48-like-producing Enterobacterales, only cefiderocol and ceftazidime/avibactam (both 88.2% susceptible) demonstrated efficacy in >75% isolates. Against VIM-producing Enterobacterales, only cefiderocol (79.0% susceptible) and colistin (93.5% susceptible) demonstrated notable activity. The proportion of susceptible NDM-producing Enterobacterales isolates was higher for cefiderocol (51.4%) and colistin (78.4%) versus all other comparators (<3%).The activity of cefiderocol and comparators by carbapenemase is summarized in TableP. aeruginosa, cefiderocol and colistin demonstrated potent activity, with 100% of isolates being susceptible to both agents. Cefiderocol and colistin were also active against non-carbapenemase-producing MDR P. aeruginosa, with 96.6% of isolates being cefiderocol susceptible and 100% being colistin susceptible.Against both VIM- and GES-producing A. baumannii producing OXA-23-like  and OXA-24/40-like  carbapenemases.Cefiderocol and colistin also demonstrated potency against P. aeruginosa and A. baumannii isolates is apparent in FigureThe wide variety of carbapenemases produced by European Enterobacterales, n = 39), followed by NDM (n = 19), VIM (n = 13), OXA-48-like (n = 10), OXA-23-like (n = 6) and OXA-24-like (n = 5). Twelve isolates carried either OSBLs/ESBLs or no known β-lactamase, with eight isolates harbouring the PER-type β-lactamase.A range of β-lactamases were identified across 99 isolates with cefiderocol MIC values of >2 mg/L, including isolates with co-carriage of multiple β-lactamases. The most common carbapenemase was KPC  and carbapenemase-producing K. pneumoniae (n = 107) were cefiderocol susceptible.Similar isolate studies involving cefiderocol have used provisional CLSI breakpoints. In a study including 1086 CR isolates from the USA (737 KPC producers), MICs were higher for isolates of Enterobacterales with β-lactamases compared with those without, but no clear association was found between the type of β-lactamase and the MIC.A key limitation of the SIDERO-CR study was the geographical spread and number of collection sites within countries. Isolates were selected by a limited number of sites per country based on the MDR/CR phenotype, therefore the relative frequency observed may not reflect the national prevalence. Additionally, the number of sites and isolates per country were not proportionate to population. Consequently, the isolate collection is not necessarily representative, as there may be considerable heterogeneity in the numbers and types of pathogens and mechanisms of resistance provided by different locales. However, from SIDERO-CR-2014–2016 isolate characterization, it is apparent that carbapenem resistance across Europe is associated with a diverse range of β-lactamases. Of the CR isolates included, 83.8% (729/870) produced at least one carbapenemase, with 65.2% (475/729) of carbapenemase-producing isolates also carrying ESBLs and/or OSBLs.K. pneumoniae with genes encoding for OXA-48 and NDM-1 has recently been reported in the Mecklenburg-Western Pomerania state of Germany.K. pneumoniae was investigated across eight Greek hospitals between 2013 and 2016.K. pneumoniae isolates were of a clonal type (ST11) similar to those identified in Bulgaria in 2015–16, indicating that the Balkan region is also at risk of increasing prevalence of MBL-producing Enterobacterales.K. pneumoniae infection has led to a shift in the carbapenemase landscape in Greece, with the incidence of MBLs increasing between 2015–17 (12.0%) and 2018 (51.1%), mainly due to VIM-producing K. pneumoniae becoming more prevalent.,,P. aeruginosa isolates resistant to ceftazidime/avibactam and 77.4% of all MDR P. aeruginosa isolates resistant to ceftolozane/tazobactam , recent reports have described an increasing proportion of non-KPC MBL-mediated resistance. A Rapid Risk Assessment report from the ECDC described an increase in NDM-containing carbapenemase-producing Enterobacterales in Italy,tam Table.A. baumannii.,P. aeruginosa and A. baumannii, but particularly in K. pneumoniae , mediated, to some extent, by transmissible mcr-1 resistance.K. pneumoniae) and 6.4% of A. baumannii, with 100% of P. aeruginosa isolates being colistin susceptible. These results do not necessarily reflect the incidence of colistin resistance in specific geographical locations or from specific infection sources; for example, the incidence of colistin-resistant A. baumannii was reported to be 47.7% in a set of A. baumannii RTI isolates from Greece, Italy and Spain (n = 65).In vitro assessment of CR-GN isolates from 18 hospitals in Greece demonstrated that ∼40% of K. pneumoniae and A. baumannii isolates were resistant to colistin; 100% of these isolates had cefiderocol MICs of ≤4 mg/L.Colistin, often considered a treatment of last resort, has a broad spectrum of GN activity and is frequently used for CR- and MDR-GN infections, particularly for MDR P. aeruginosa. Cefiderocol retained activity against the majority of isolates harbouring MBLs, with only 3.0% (3/99) of MBL-positive CarbNS Enterobacterales isolates having MIC values >4 mg/L. The in vitro activity (MIC ≤2 mg/L) of cefiderocol against a range of serine-β-lactamases varied from 100% in OXA-58-producing isolates to 83.6% in KPC producers. In addition, 91.5% of isolates with no carbapenemase and 97.5% with no β-lactamase had a cefiderocol MIC ≤2 mg/L, demonstrating potent cefiderocol activity where the mechanism of resistance was not clear.In this study, the proportions of isolates susceptible to cefiderocol are similar to comparators in Enterobacterales and generally similar to colistin but greater than other comparators in P. aeruginosa isolates (98.3% cefiderocol susceptible), yet half carried no carbapenemase. The low prevalence of carbapenemases in SIDERO-CR-2014–2016 P. aeruginosa isolates aligns with recently reported data describing the prevalence of XDR P. aeruginosa; of 1445 P. aeruginosa isolates collected from 51 Spanish hospitals, 252 (17.3%) were classified as XDR and only 3.1% carried either carbapenemases or ESBLs.P. aeruginosa isolates in SIDERO-CR-2014–2016 is likely to be due to porin- and efflux pump-mediated mechanisms.in vitro activity against strains with alterations in outer membrane porins or overexpressed efflux pumps.,Cefiderocol demonstrated potency against the MDR in vitro activity against a broad range of CarbNS and MDR pathogens. The isolates tested included β-lactamase-producing strains from all Ambler classes, with cefiderocol demonstrating activity against the key MBLs VIM and NDM, as well as against clinically important serine-β-lactamases KPC, GES and OXA, and isolates co-carrying ESBLs and OSBLs. Consequently, cefiderocol represents a key addition to the limited armamentarium available for the treatment of infections caused by CR- and MDR-GN organisms and could be a particularly valuable and timely treatment option for when resistance is apparent but the mechanism is unknown.In conclusion, the SIDERO-CR European dataset, coupled with the rise of resistance to existing agents and the shift towards MBL- from serine-β-lactamase-producers, demonstrates the diverse and dynamic nature of the European carbapenemase landscape. In SIDERO-CR-2014–2016, cefiderocol exhibited potent dlaa060_Supplementary_DataClick here for additional data file.


EGFR) T790M mutation on osimertinib efficacy remains unclear.Seventy-eight patients were studied with EGFR-mutated NSCLC and LM. Case data were collected and EGFR mutation status of circulating cell-free DNA from paired CSF, and plasma of 23 patients with LM was detected using droplet digital PCR. The median overall survival (mOS) was 8.08 months (95% CI: 6.07–10.09) in the study. Forty-four osimertinib-treated patients had an improved mOS of 13.15 (95% CI: 5.74–20.57) and a median progression-free survival (PFS) of 9.50 months (95% CI: 6.77–12.23) when compared with patients treated with first- or second-generation EGFR-TKI (mOS = 3.00 months (95% CI: 1.32–4.68) and median PFS = 1.50 months (95% CI: 0.00–3.14)). In the osimertinib group, mOS values for CSF with and without T790M mutation were 22.15 months (95% CI: 9.44–34.87) and 13.39 months (95% CI: 7.01–19.76), respectively, with no statistical differences. Regardless of the CSF T790M mutation status, osimertinib demonstrated significant efficacy against LM associated with NSCLC.Osimertinib has demonstrated promising efficacy against leptomeningeal metastasis (LM) associated with T790M-positive non-small-cell lung cancer (NSCLC). However, the effect of cerebrospinal fluid's (CSF's) epidermal growth factor receptor ( EGFR) is the most important driver gene in NSCLC. EGFR mutations occur in 10–20% of Caucasian patients with NSCLC but in 40–60% of Asian patients [EGFR mutations [EGFR tyrosine kinase inhibitor (EGFR-TKI) treatments [Leptomeningeal metastasis (LM) is defined as the spread of malignant cells within the leptomeninges and subarachnoid space, resulting in a devastating prognosis with limited treatment options . Non-smautations , 4. ThisEGFR mutation status is a critical prognostic factor in the treatment of NSCLC with LM. A secondary EGFR test is often required to determine the status of T790M drug-resistant mutation. However, it is difficult to obtain a second tumor tissue from most patients. Circulating cell-free DNAs (cfDNAs) are extracellular nucleic acids released by tumor cells that can be useful biomarkers for early diagnosis and prognosis [rognosis . Howeverrognosis , 7.EGFR-TKIs remains dismal [EGFR-TKIs, such as afatinib, could partially penetrate the blood–brain barrier, they exhibit no obvious advantage as a treatment for LM [LM is often associated with an extremely poor prognosis, with a median OS (mOS) of 3–10 months after diagnosis . Currents dismal –13. AlthEGFR-TKI that selectively inhibits EGFR and EGFR T790M mutations. It is highly effective in both untreated and previously treated patients with EGFR-mutated NSCLC by prolonging OS and progression-free survival (PFS) [EGFR-mutated positive NSCLC [EGFR T790M mutation [Osimertinib is an irreversible third-generation al (PFS) –16. Accove NSCLC . In contEGFR T790M mutation status and osimertinib efficacy in patients with NSCLC [EGFR-TKIs and evaluate the factors influencing osimertinib efficacy.Few studies have focused on the relationship between CSF th NSCLC , 18. In EGFR-mutated NSCLC and LM at the People's Hospital of Zhengzhou University, recruited from January 1, 2014, to December 31, 2020. Inclusion criteria were: (1) space-occupying lesions of the lungs detected using computed tomography (CT) or whole-body positron emission tomography/computed tomography (PET/CT) and confirmed as NSCLC using histological biopsy, (2) symptoms and signs of the central nervous system metastases, (3) typical leptomeningeal enhancement on magnetic resonance imaging (MRI), and (4) cytologic identification of malignant cells within the CSF. All patients were scored using the Eastern Cooperative Oncology Group performance status (ECOG PS), CT, or PET/CT. In addition, the patient's histology, metastasis site, imaging, CSF parameters, and molecular profiling were collected. And no patients had complicated organ failure, infection in the central nervous system, and diseases of the autoimmune system or hematological system.This retrospective study involved patients with EGFR-mutated NSCLC and LM, 34 patients were treated with first- or second-generation EGFR-TKIs treatment, and 44 patients  were treated with osimertinib  analysis or next-generation sequencing.EGFR kit . All protocols of analysis were carried out in strict accordance with the manufacturer's instructions.Secondary T790M mutation analysis was performed using one of the following samples: biopsied tumor tissues, plasma, or CSF samples. Approximately 10 mL of whole blood and 10 mL of CSF (EDTA as the anticoagulant) were collected for the purpose of cfDNA extraction. The retrieved CSF was used for analysis within 4 h of the lumbar puncture procedure. Plasma and CSF were centrifuged at 1,600 × g and 10,000 × g, respectively, for 10 min at room temperature. CfDNA was extracted using the QiAamp circulating nuclear acid kit . Droplet digital PCR (ddPCR) was used to detect cfDNA using the Sysmex OncoBeam EGFR-TKI treatment to tumor progression. The cutoff date was December 31, 2020.OS for LM was defined as the time from LM diagnosis to death. PFS was calculated from the onset of p values were used for survival analysis at 95% confidence intervals (95% CI). Cox regression analysis was performed to estimate hazard ratios (HRs) and 95% CIs for OS. Statistical significance was set at p < 0.05.Statistical analyses were performed using SPSS software . Chi-square univariate analysis was used for continuous variables, and a non-parametric test was used for non-normal distribution variables. Kaplan–Meier estimation and log rank EGFR-mutated NSCLC and LM were included. The median age was 61 years (range: 28–78), and 74.4% (58/78) were non-smokers. The ECOG PS scores at the time of LM diagnosis were: <1 in 23 (29.5%), 2–3 in 43 (55.1%), and 4 in 12 patients (15.4%). Twenty-five patients with NSCLC also presented with LM at initial diagnosis. The median time from NSCLC to LM diagnosis of the remaining 53 patients was 21 months (95% CI: 12.0–24.5). Statistically significant differences were not observed in age, sex, smoking status, histology, initial ECOG PS scores, and EGFR mutations.As shown in EGFR T790M mutation analysis. Of these samples, EGFR T790M mutation was identified in 28 cases.Among the 78 patients, 66 (84.6%) of them underwent secondary EGFR gene of cfDNA in 23 patients simultaneously, and T790M mutation was detected in 8 patients (Supplementary EGFR mutations in plasma and CSF samples were 56.5% (13/23) and 60.9% (14/23), respectively, but without any statistically significant difference (p=0.765).Plasma and CSF were used to detect the EGFR-TKI: 24 (45.3%) of them received gefitinib, 11/53 (20.8%) received erlotinib, 7/53 (13.2%) received icotinib, 5/53 (9.4%) received afatinib and gefitinib, and 6/53 (11.3%) received osimertinib. Moreover, 35/53 (66.0%) patients received cytotoxic chemotherapy, and 11/53 (20.8%) had undergone whole-brain radiation therapy (WBRT) for brain metastasis prior to LM  were treated with osimertinib at the time of LM diagnosis. The median duration of osimertinib treatment was 7.0 months (95% CI: 4.0–9.0). Thirty-four patients received first- or second-generation TKI treatment without osimertinib. Eighteen (52.9%) of them received gefitinib; 10 (29.4%) received erlotinib; 3 (8.8%) received icotinib; and 1 (2.9%) received afatinib and gefitinib. Twelve patients (35.7%) received cytotoxic chemotherapy; 4 (12.9%) received WBRT; 2 (6.5%) underwent IT chemotherapy; and 3 (9.7%) had VPS insertion .At the end of follow-up, 58 patients died from the disease, 15 were still alive, and 5 were lost to follow-up (6.4%). The 1-year OS rate in the study was 32.7% (19/73), with a mOS of 8.08 months , with a 1-year OS rate of 12.9%, which was shorter than that of patients treated with osimertinib (HR: 0.58 (95% CI: 0.44–0.77) and p ≤ 0.001; Among the 44 patients who received osimertinib treatment, 39 (88.6%) had positive clinical responses. Twenty-eight patients died, and 14 were still alive at the time of follow-up, with a mOS of 13.15 months (95% CI: 5.74–20.57). The 1-year OS rate was 53%. Among the patients who were treated with first- or second-generation Compared to the group treated with osimertinib, PFS of the group treated without osimertinib also was significantly shorter (PFS: 1.50 months (95% CI: 0.00–3.14) and HR: 0.57 month (95% CI: 0.44–0.75); p=0.564). Regardless of T790M status, the survival benefit of the osimertinib treatment group was better than that of the first- or second-generation EGFR-TKI treatment group (mOS = 3.00 months (95% CI: 1.32–4.68) and HR: 0.56 (95% CI: 0.41–0.78); Furthermore, we further divided the group treated with osimertinib into two subgroups according to T790M mutational status. The mOS for patients with and without T790M mutational status was 15.92 months (95% CI: 7.70–24.14) and 9.00 months (95% CI: 5.50–12.50), respectively, but without any significant difference (EGFR mutation in CSF cfDNA was detected in 23 patients of the osimertinib treatment group. Seven patients with T790M mutation in CSF had a mOS of 22.15 months (95% CI: 9.44–34.87), whereas 16 patients without T790M mutation had a mOS of 13.39 months (95% CI: 7.01–19.76). No statistical difference was found according to Kaplan–Meier survival analysis  and 13.39 months (95% CI: 3.59–23.18), respectively. However, survival analysis revealed no significant differences between the two groups p=0.416; .p=0.001 and multivariate HR: 3.22 (95% CI: 1.74–5.95), p=0.021). Osimertinib treatment was identified as a significant independent favorable prognostic factor in patients with NSCLC and LM (univariate HR: 0.59 (95% CI: 0.45–0.78), p=0.000 and multivariate HR: 0.65 (95% CI: 0.49–0.87), p=0.004). Univariate analysis revealed a statistically significant correlation between WBRT and prognosis (0.45 (95% CI: 0.23–0.96), p=0.038). However, no significant associations were observed with concurrent brain metastases, initial EGFR and EGFR T790M mutations, CSF pressure, protein and glucose levels, and systemic and intrathecal cytotoxic chemotherapy , otherapy .EGFR-TKI drugs have resulted in significant survival benefits and improved quality of life for patients with EGFR mutations. Third-generation EGFR-TKIs, such as osimertinib, have also been found to extend the survival of patients with T790M mutation by more than 10 months [The treatment of lung cancer has gradually evolved from traditional therapies to gene-oriented personalized treatments. The first- and second-generation 0 months .EGFR + NSCLC, regardless of T790M mutation that confers drug resistance, with a median LM PFS of 9.50 months and a mOS of 13.15 months. Patients treated with only first- or second-generation EGFR-TKI after LM diagnosis had a median PFS of 1.50 months and a mOS of 3.00 months. Compared to first- or second-generation EGFR-TKIs, osimertinib showed a markedly improved survival benefit.Our study found that osimertinib is a clinically effective standard regimen in patients with LM associated with EGFR-TKIs for LM is limited because of the blood–brain barrier. The first- and second-generation EGFR-TKIs have low CSF penetration, with an average CSF penetration of 1–3% for gefitinib and 3–6% for erlotinib compared to only 1% for afatinib [As mentioned, the efficacy of first-generation afatinib –13. Highafatinib . Althougafatinib .EGFR mutation NSCLC [EGFR-TKI therapy failure [Preclinical studies of osimertinib using mouse models of brain metastasis with 19 gene deletions (PC9) have demonstrated that a dose-dependent tumor reduction can be achieved . The AURon NSCLC . Several failure , 23. SimEGFR T790M screening failure in the BLOOM study might be a missed opportunity in revealing osimertinib activity in the CNS [EGFR-TKI-resistant NSCLC and without the T790M mutation should be further investigated.The excluded patients based on  the CNS . A recen the CNS . SimilarEGFR mutations detected using CSF cfDNA tended to be higher compared to plasma cfDNA [EGFR mutation rate based on CSF samples [in vivo, in vitro, and/or in silico along with other math tools and models, including meta-analysis [Due to the blood–brain barrier, plasma cfDNA may not accurately reflect the actual state of mutations in intracranial tumors, whereas the rapid CSF circulation within the cerebral ventricle and spinal cord cavity indicates that CSF cfDNA may be a reliable biomarker for intracranial tumors. In patients with LM associated with metastasized lung carcinoma, the proportion of ma cfDNA . A simil samples , which i samples . Howeveranalysis , 26, resanalysis , 28, netanalysis , and molanalysis , 31.This study had several limitations. Firstly, this was a retrospective study involving a small number of patients. All the patients included in our study were confirmed using cytological findings, whereas patients diagnosed via clinical examination and imaging were not included, which may have contributed to selection bias. Secondly, the ddPCR method for detecting plasma and CSF cfDNA had low sensitivity, which could have potentially resulted in a lower detection rate. Therefore, more sensitive cfDNA detection assays, such as NGS, will be employed for further verification analysis.EGFR mutation status of circulating cell-free DNA from paired CSF, and plasma of 23 patients with LM was detected using droplet digital PCR. The median overall survival (mOS) was 8.08 months (95% CI: 6.07–10.09) in the study. Forty-four osimertinib-treated patients had an improved mOS of 13.15 months (95% CI: 5.74–20.57) and a median progression-free survival (PFS) of 9.50 months (95% CI: 6.77–12.23) when compared with patients treated with first- or second-generation EGFR-TKI (mOS = 3.00 months (95% CI: 1.32–4.68) and median PFS = 1.50 months (95% CI: 0.00–3.14)). In the osimertinib group, mOS values for CSF with and without T790M mutation were 22.15 months (95% CI: 9.44–34.87) and 13.39 months (95% CI: 7.01–19.76), respectively, with no statistical differences. Regardless of the CSF T790M mutation status, osimertinib demonstrated significant efficacy against LM associated with NSCLC.Case data were collected and


Kappaphycus alvarezii, Eucheuma denticulatum, Halymenia durvillaei (Rhodophyta), Caulerpa lentillifera, Caulerpa racemosa (Chlorophyta), Dictyota dichotoma and Sargassum polycystum (Ochrophyta). This review aims to highlight the therapeutic potential of North Bornean seaweeds and their nutraceutical profiling. North Bornean seaweeds have demonstrated anti-inflammatory, antioxidant, antimicrobial, anticancer, cardiovascular protective, neuroprotective, renal protective and hepatic protective potentials. The protective roles of the seaweeds might be due to the presence of a wide variety of nutraceuticals, including phthalic anhydride, 3,4-ethylenedioxythiophene, 2-pentylthiophene, furoic acid , eicosapentaenoic acid, palmitoleic acid, fucoxanthin, β-carotene (E. denticulatum), eucalyptol, oleic acid, dodecanal, pentadecane (H. durvillaei), canthaxanthin, oleic acid, pentadecanoic acid, eicosane (C. lentillifera), pseudoephedrine, palmitic acid, monocaprin (C. racemosa), dictyohydroperoxide, squalene, fucosterol, saringosterol (D. dichotoma), and lutein, neophytadiene, cholest-4-en-3-one and cis-vaccenic acid (S. polycystum). Extensive studies on the seaweed isolates are highly recommended to understand their bioactivity and mechanisms of action, while highlighting their commercialization potential.Malaysia has a long coastline surrounded by various islands, including North Borneo, that provide a suitable environment for the growth of diverse species of seaweeds. Some of the important North Bornean seaweed species are  Marine organisms have been widely used as sources of functional bioactive compounds over the years . Among mKappaphycus alvarezii (Doty) Doty, Eucheuma denticulatum (Burman) Collins et Harvey, Halymenia durvillaei Bory de Saint-Vincent, Caulerpa lentillifera J. Agardh, Caulerpa racemosa (Forsskål) J. Agardh, Dictyota dichotoma (Huds.) Lamouroux, and Sargassum polycystum C. Agardh [Malaysia is part of the Coral Triangle, a geographical area in Southeast Asia and the Pacific that includes the oceans close to Indonesia, Malaysia, Philippines, Papua New Guinea, Timor-Leste and Solomon Islands. The temperatures of Malaysia’s coastal waters make it ideal for the development and growth of a wide variety of seaweed species. The North Borneo region of Malaysia is one of the main seaweeds growing areas; it has a suitable environment for cultivating a diverse variety of seaweeds and is the only region of Malaysia where seaweeds are farmed commercially . In Nort. Agardh . Brief dSeaweeds are also known as sea vegetables and have been used for the treatment of various disorders ,14,15,16Inflammation is a recognized defensive mechanism evolved in high-level organisms in response to stressors that disrupt bodily homeostasis. Microbial infections, tissue stress and some traumas are examples of hazards that cause inflammation with common symptoms of fever, redness, swelling and pain ,24. OverK. alvarezii has been reported to have anti-inflammatory potential in asthma-induced rats. The extract changed circulating white blood cell levels, reduced mucin synthesis, and downregulated the expression of TNF-α, IL-4, nuclear factor kappa beta (NF-κB), epidermal growth factor receptor (EGFR) and matrix metalloproteinase (MMP-9). The consumption of seaweed may be useful for asthma patients. The extract decreases bronchiole smooth muscle thickness for airflow facilitation and decreases asthmatic inflammation, lung eosinophil infiltration, and mucin production [oduction .C. lentillifera polysaccharides has been reported to have an inhibitory impact on lipopolysaccharide (LPS)-induced HT29 colorectal carcinoma cells, lowering the overproduction of TNF-α and IL-1β, SIgA and mucin2 (related proteins), as well as decreasing TNF-α and IL-1β expression [C. racemosa polysaccharides have been reported to have anti-inflammatory potential and activate the hemoxigenase-1 (HO-1) pathway to sustain the production of hemoxigenase-1 enzyme, crucial for the prevention of inflammation [The anti-inflammatory activity of pression . C. raceammation . D. dichotoma extract at a concentration of 25 ug/mL inhibited the production of NO and PGE2, followed by a reduction in the expression of inducible nitric oxide synthase (iNOS) and COX-2 proteins, and iNOS and COX-2 mRNA in a dose-dependent pattern. COX-2 and iNOS are implicated in a variety of pathological processes, including inflammation. The solvent fractions of D. dichotoma extracts also decrease the mRNA expression of other cytokines including TNF-α, IL-1, and IL-6 in the murine macrophage cell line [In murine macrophage RAW 264.7 cells, the dichloromethane fraction of ell line .S. polycystum has been reported using a mouse model with the paw edema method, where the mouse paw was inflamed and the hexane fraction of seaweed extract at a concentration of 70 mg/kg b.w. was applied. The anti-inflammatory effect was measured by the decreased percentage of edema size. The hexane fractions of S. polycystum extract significantly reduced the edema size compared to untreated mice [Anti-inflammatory and analgesic activity from brown algae ted mice .Antioxidant phytochemical compounds can scavenge the reactive oxygen species (ROS) and reactive nitrogen species (RNS) in the human body, and slow down or prevent the onset of oxidative stress-related diseases including cardiovascular diseases (CVDs), neurological diseases , Parkinson’s disease, multiple sclerosis, amyotrophic lateral sclerosis (ALS), and depression), cancer , respiratory disease (chronic obstructive pulmonary disease (COPD) and asthma), rheumatoid arthritis, delayed sexual maturation, and kidney and liver diseases ,31,32,33K. alvarezii 1.63 and 225.00 TEAC and FRAP mM.mg/dry extract and 22.50 TPC mg PGE/g dry extract, E. denticulatum 1.54 and 153.97 TEAC and FRAP mM.mg/dry extract and 15.82 TP mg PGE/g dry extract, H. durvillaei 1.67 and 182.29 TEAC and FRAP mM.mg/dry extract and 18.90 TP mg PGE/g dry extract, C. lentillifera 2.16 and 362.11 TEAC and FRAP mM.mg/dry extract and 42.85 TP mg PGE/g dry extract, C. racemosa 2.01 and 355.36 TEAC and FRAP mM.mg/dry extract and 40.36 TP mg PGE/g dry extract, D. dichotoma 1.66 and 268.86 TEAC and FRAP mM.mg/dry extract and 35.23 TP mg PGE/g dry extract, S. polycystum 1.86 and 366.69 TEAC and FRAP mM.mg/dry extract and 45.16 TP mg PGE/g dry extract compared to butylated hydroxytoluene (standard) (3.84 and 615.71 TEAC and FRAP mM.mg/dry extract) [C. lentillifera has high antioxidation activities followed by C. racemosa, S. polycystum, H. durvillaei, D. dichotoma, K. alvarezii and E. denticulatum; while in terms of TP, S. polycystum has indicated high values followed by C. lentillifera, C. racemosa, D. dichotoma, K. alvarezii, H. durvillaei and E. denticulatum [Marine seaweeds from North Borneo are a good source of antioxidants ,11,34,35Pathogenic microbes including bacteria, fungi, viruses and parasites are responsible for the development of various diseases that arise in the community . SeaweedK. alvarezii against Staphylococcus aureus (S. aureus)  has been examined. Administration of the extract at a concentration of 200 mg/mL resulted in an inhibition zone of 10.03 mm [K. alvarezii was also effective against S. aureus, Staphylococcus epidermidis (S. epidermidis) , Pseudomonas aeruginosa (P. aeruginosa)  and Bacillussubtilis (B. subtilis)  bacterial strain at a concentration of 30–80% (w/v) with an inhibition zone of 11.93–14.85 mm, while ethyl acetate fractions of the ethanol extract of K. alvarezii at a concentration of 50% (w/v) inhibited the growth of S. aureus, S. epidermidis, P. aeruginosa and B. subtilis with inhibition zones of 7.14, 19.70, 0.73 and 18.30 mm, respectively. No antifungal activity of K. alvarezii extract and fractions have been published against Candida albicans  (C.P. Robin) Berkhout and Aspergillus niger (A. niger) (Tiegh) [K. alvarezii extract have been reported against Lagenidium spp and Haliphthoros fungal strains [Lagenidium thermophilum (L. thermophilum)  and Haliphthoros sabahensis (H. sabahensis)  are pathogenic to the eggs and larval stages of Scyllaserrata  and Scyllatranquebarica  (mangrove crabs) [K. alvarezii inhibited the hyphal growths of L. thermophilum IPMB 1401  and H. sabahensis IPMB 1402 [K. alvarezii  exhibited strong anti-influenza activity against a wide spectrum of influenza virus strains, including the newly evolving swine-origin H1N1-2009 influenza strain. The mechanism involved the direct binding of ECA-2 to the viral envelope protein hemagglutinin (HA) and inhibited influenza virus propagation [The antibacterial potential of the aqueous extraction of 10.03 mm . The eth (Tiegh) . On the e crabs) ,40. The pagation .E. denticulatum inhibited the growth of S. aureus with inhibition zones of 6.0–16.3 mm. Furthermore, the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values for E. denticulatum extract against S. aureus were 10% and 15%, respectively. At 10%, minimum bacterial growth was observed, the number of bacteria greatly decreased from 3.0 × 107 to 1.5 × 102 CFU/plate, and turbidity levels also decreased; while at 15% of the extract, no bacterial growth was noticed [E. denticulatum extract has been reported against Aspergillus flavus (A. flavus) (Link) [E. denticulatum have been tested for in vitro antiviral activity against human herpes virus type 1 (HHV-1). Carrageenans indicated an antiviral impact by the inhibition of virus attachment and interference in a subsequent stage of the virus replicative cycle. HHV-1 viral DNA synthesis was reduced by 3 folds in cultures treated with sulphated polydigalactosides from E. denticulatum (0.75 mg/mL) [The ethanol extract of  noticed . No anti) (Link) . Sulphat5 mg/mL) .H. durvillaei has been reported to have antimicrobial effects. The presence of antimicrobial activities of H. durvillaei extract against pathogenic bacteria was determined using the disc diffusion method. The solvent extract of H. durvillaei inhibited the growth of P. aeruginosa (11.89 mm), S. aureus (12.22 mm), and Streptococcus pyogenes (S. pyogenes) Rosenbach, 1884 (10.67 mm), respectively. However, no fungicidal activity of the solvent extract of H. durvillaei has been reported against C. albicans [The extracts of albicans .C. lentillifera were tested against Methicillin-resistant S. aureus (MRSA) and neuropathogenic Escherichia coli K1 (E. coli K1). Moderate antibacterial activity of 62.17% against MRSA and poor antibacterial impact against E. coli K1 of 12.42% were demonstrated by C. lentillifera extract at a concentration of 250 μg/mL [C.lentillifera (0–128 μg/mL) was recorded against the shrimp pathogenic bacteria Vibrio vulnificus  Farmer, 1980, V. alginolyticus  Sakazaki, 1968, V. parahaemolyticus  Sakazaki et al., 1963 or V. harveyi, , as compared with positive  and negative controls [C. lentillifera extract was tested against White Spot Syndrome Virus (WSSV)  [C. lentillifera extract yielded very good outcomes. Shrimps injected with WSSV and C. lentillifera (1–10 mg/mL) preincubated solutions exhibited significantly lower mortality of 0.0–6.7%, compared with the positive control (100%) (only WSSV-injected). This inhibitory effect was further confirmed by the reduction in viral loads of WSSV, and C. lentillifera (1–10 mg/mL) expressed significantly lower viral loads  than the positive control  [C. lentillifera against L. thermophilum and H. sabahensis have been reported as well. An ethanol extract of C. lentillifera inhibited hyphal growths of L. thermophilum IPMB 1401, L. thermophilum IPMB 1601 and H. sabahensis IPMB 1603 [The chloroform extracts of 50 μg/mL . Howevercontrols . The ant genome) ,47. WSSV genome) . The admtal DNA) . FungiciPMB 1603 .C. racemosa demonstrated antibacterial activity against MRSA and E. coli K1. The extract of C. racemosa, at a concentration of 250 μg/mL, displayed a high antibacterial effect of 97.7% against MRSA, but a weak effect of 19.90% against E. coli K1. A methanol extract of C. racemosa (250 μg/mL) also showed antibacterial activity of 61.54% and 42.91% against MRSA and E. coli K1 [C. racemosa has been reported to show antifungal activity against A. flavus. An ethanol extract of the seaweed demonstrated the strongest inhibitory power with a 30 mm diameter inhibition zone against A. flavus [C. racemosa was demonstrated against the Chikungunya virus (CHIKV) [Aedes aegypti and Aedes albopictus mosquitoes, which cause high fever, joint pain, back pain, vomiting, headache, kidney, liver, heart disease, etc. [C. racemosa was determined based on inhibition of the cytopathic effect caused by CHIKV on African monkey kidney epithelial (Vero) cells. Chloroform, ethyl acetate, ethanol, and methanol extracts (5 to 640 μg/mL) of C. racemosa showed a significant inhibition effect [A chloroform extract of  coli K1 , respect. flavus . The ant (CHIKV) . The virn effect .D. dichotoma at a concentration of 1.5 mg/disc were investigated for in vitro antibacterial and antifungal activities. The results indicated that the methanol extract inhibited the growth of B. subtilis (6.5 mm) and S. aureus (7.5 mm). The dichloromethane extract inhibited the growth of B. subtilis (7.0 mm), Enterobacter aerogenes (E. aerogenes)  (6.5 mm), E. coli (6.5 mm), Proteus vulgaris (P. vulgaris) (11.0 mm) and Salmonella typhimurium (S. typhimurium)  (7.0 mm), whereas the hexane extract inhibited the growth of B. subtilis (9.0 mm) and S. aureus (7.5 mm) only [D. dichotoma has been shown against Salmonella typhi (S. typhi), Klebsiella pneumoniae (K. pneumoniae)  Trevisan, 1887 and Shigella boydii (S. boydii)  [D. dichotoma has been reported against Mucor sp. and A. flavus [D. dichotoma (1.5 mg/disc) has been observed against C. albicans [D. dichotoma extract has been tested against herpes simplex virus (HSV) and coxsackievirus B3 (CVB3) [Methanol, dichloromethane and hexane extracts of mm) only . The antg, 1949) . The ant. flavus , while nalbicans . The ant3 (CVB3) . HSV bel3 (CVB3) ,55. The 3 (CVB3) .S. polycystum was tested against human pathogenic bacteria. The methanol extract of the seaweed resulted in the inhibition of P. aeruginosa (15 mm), K. pneumoniae (16 mm), E. coli (19 mm), and S. aureus (20 mm) [S. polycystum indicated no inhibition against B. subtilis or S. enteritidis. Similarly, no antifungal activity has been observed against A. niger [A solvent extract of  (20 mm) . HoweverA. niger .Cancer is one of the main causes of mortality in the world, and many research facilities are now focusing on the development of new anticancer medicines that could improve chemotherapy treatment and reduce mortality rates . The proK. alvarezii has been reported with anti-breast and anti-colorectal cancer potential. The anticancer activities were expressed with inhibitory concentration (IC50) value (µg/mL). An IC50 value of less than 100 is considered to indicate an active compound with anticancer properties. An ethanolic extract of K. alvarezii exhibited anticancer activity against human breast adenocarcinoma cell line MCF-7 with an IC50 of 75.7 µg/mL, while ethyl acetate and hexane extracts showed anti-colorectal cancer activity against human colorectal carcinoma cell line HCT-116 with IC50 values of 21.4 and 43.0 µg/mL, respectively [A solvent extract of ectively .E. denticulatum against Ehrlich carcinoma and Meth-A fibrosarcoma has been reported. Oral administration of the extract (1600 mg/kg b.w.) for 28 days resulted in the inhibition of Ehrlich carcinoma by 25% in tumor-implanted mice. Similarly, intraperitoneal administration of E. denticulatum extract (50 mg/kg b.w.) for 7 days resulted in the inhibition of Meth-A fibrosarcoma by 17% [H. durvelaei was investigated against four cancer cell lines . The results indicated that administration of H. durvelaei extracts reduced the growth of AGS and HT-29 cell lines by 27.17% and 1.47%, respectively [The antitumor activity of a by 17% . The antectively .C. lentillifera have been reported to show antitumor properties against human breast adenocarcinoma cell line MCF-7. Exposure to C. lentillifera oligosaccharides inhibited the growth of MCF-7 cells in a dose-dependent manner and induced apoptosis (triggered chromatin condensation and poly ADP-ribose polymerase degradation) [Oligosaccharides  obtained from adation) .C. racemosa have been reported to show antitumor activity in tumor-inoculated mice (H22 tumor). The results indicated that administration of C. racemosa polysaccharide fractions of different doses could significantly inhibit the H22 tumor. After 14 days of transplantation, the weight of tumors in mice without polysaccharide treatment increased to 1.02 g while the tumor weight in mice exposed to the polysaccharide at a dose of 100 mg/kg b.w./day by routine oral passage decreased to 0.47 g, and the tumor inhibition rate reached 53.9% [Polysaccharide fractions (coded as CRP) obtained from ed 53.9% .D. dichotoma were tested against seven different cancer cell lines including HCT-116, MCF-7, HepG2, A-549, PC-3, HeLa, and CACO in 96-well plates. The extract and fractions were applied at concentrations ranging from 0.86 to 100 μg/mL. The results demonstrated that D. dichotoma extract and fractions displayed significant anticancer effects against several cancer cell lines in a dose-dependent manner but were generally more selective against MCF-7 and PC-3 cell lines. The chloroform fraction was the most effective in MCF-7, PC3, and CACO cells  followed by the petroleum ether fraction against MCF-7 and PC-3  and the ethyl acetate fraction against HepG2 and CACO  [A methanol extract and fractions  of ctively) .S. polycystum at a concentration ranging from 25–150 μg/mL has been reported against human breast adenocarcinoma cell line MCF-7 via cell viability assay. Treatment with the sulphated polysaccharides indicated the highest percentage of inhibition (90.4%) against the MCF-7 cell line at 150 μg/mL with an estimated IC50 of 50 μg/mL [S. polycystum polysaccharide induced apoptosis in colorectal cancer cell lines (HCT-15 cell) [The anticancer activity of sulphated polysaccharides (fucoidan) from 50 μg/mL . In anot15 cell) .The hypocholesterolemic effects of various marine algae and algal polysaccharides have been reported. The administration of extract significantly reduced serum total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and triglycerides (TG) ,20.K. alvarezii to the HCF diet dramatically decreased body weight (29.1%), LDL-C (49.3%), plasma TC (11.4%), TG (36.1%), plasma MDA level (10.7%), GSH-Px (13.49%), SOD (9.4%) and CAT (24.48%), and significantly increased HDL-C levels (55%), compared to rats fed the HCF diet only [After 16 weeks on high-cholesterol/high-fat (HCF) diets, male Sprague-Dawley rats weighing 260–300 g had significantly higher body weight (b.w.), lipid peroxidation , end-product of lipid peroxidation), plasma low-density lipoprotein cholesterol (LDL-C), plasma total cholesterol (TC), plasma triglycerides (TG), erythrocyte glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and catalase (CAT) levels. The addition of 5% iet only . E. denticulatum played a vital role in the reduction of fat absorption by the body via inhibition of pancreatic lipase [E. denticulatum extract at a concentration of 3.8 mg/mL showed pancreatic lipase activity inhibition with an 83% reduction [c lipase . Inhibitc lipase ,66. An Eeduction . C. lentillifera was reported to show anti-obesity and cardiovascular protection activity. Supplementation with 5% C. lentillifera extract for 16 weeks in HCF-diet rats reduced body weight by 39.5%, significantly increased HDL-C levels by 48.7%, reduced plasma TC by 18.4%, LDL-C by 34.6% and TG by 33.7%, and lowered plasma MDA level by 9%, GSH-Px by 31.8% and CAT by 3.14%, compared to the corresponding levels in high-cholesterol-diet rats [Treatment of HCF diet rats with iet rats . S. polycystum significantly reduced body weight gain, plasma antioxidant enzymes and plasma lipid peroxidation to levels closer to those of healthy rats. Supplementation with 5% S. polycystum in rats fed a high-fat diet reduced body weight by 42.6%, significantly increased HDL-C levels by 16.2%, reduced plasma TC by 11.4%, LDL-C by 22% and TG by 7.69%, and decreased the plasma MDA level by 6.8%, GSH-Px by 43.4% and CAT by 15.7%, as compared to the corresponding levels in hypercholesterolemia and hypertriglyceridemia rats [The exposure of induced-hypercholesterolemia and -hypertriglyceridemia rats to mia rats .Millions of people die each year as a result of hepatic diseases across the world ,68. The K. alvarezii ethanolic extract administered for 25 days against lead acetate-induced hepatic injury in mice has been investigated. The extract at a concentration of 800 mg/kg b.w. reduced AST, ALT and ALP levels by 15.79%, 18.52% and 16.11%, respectively, compared to lead acetate-treated mice . Mice administered with ethanol extract of K. alvarezii (800 mg/kg b.w.) also demonstrated a significant (p < 0.05) elevation in SOD and GPx levels by 45.94% and 18.78%, respectively, and a significant (p < 0.05) reduction in MDA level by 22.83%, compared with lead acetate-treated mice. Histological observations of mouse hepatic tissues treated with K. alvarezii ethanolic extract indicated improved hepatic cell structure, blood congestion, and fatty degeneration compared to lead acetate-treated mice [The hepatoprotective activity of ted mice .C. lentillifera demonstrated hepatoprotection against acetaminophen  induced hepatic damage in juvenile zebrafish (aged 1–3 months). The administration of APAP to the control group at a concentration of 10 μM caused fish mortality; while the introduction of the methanol extract of C. lentillifera at concentrations of 10, 20 and 30 μg/l to tanks holding 10 μM APAP-treated groups reduced fish mortality. Histological observation by hematoxylin and eosin staining of zebrafish hepatic tissues exposed to 10 μM APAP and concurrently administered C. lentillifera extract indicated a reduction in hepatic necrosis, hepatocyte swelling, hepatocyte vacuolization and leukocyte infiltration in a dose-dependent manner, as compared to the control group treated solely with 10 μM APAP [A methanol extract of  μM APAP .C. racemosa at a concentration of 200 mg/kg b.w. was administered for 30 days on a daily basis to 40% carbon tetrachloride (CCl4) induced hepatic fibrosis rats . Intoxicated rats treated with water extracts of C. racemosa showed significant (p < 0.05) decreases in their high levels of AST (46.7%), ALT (82.2%), ALP (41.3%), LDH (25.8%) and total bilirubin (69.6%), as compared to CCl4 treated control rats [An aqueous extract of rol rats . S. polycystum was examined in acetaminophen  induced hepatic oxidative injured rats. The oral administration of S. polycystum extract in intoxicated rats at a concentration of 200 mg/kg b.w./day for 15 days reduced elevated levels of ALT (27.64%), AST (56.43%), LDH (43.38%), ALP (72.53%) and MDA (31.50%), compared to the levels in an APAP-administered control group [The protective effect of a solvent extract of ol group .Neuroprotection is a strategy for halting the progression of neurodegeneration . NeurodeK. alvarezii might be beneficial as a food supplement or medication for those who are prone to neurological disorders. In primary cultures of hippocampal neurons, the effects of K. alvarezii extracts on the development and complexity of neuronal cytoarchitecture were reported. A solvent extract of K. alvarezii with an optimal concentration of 1 μg/mL was added to primary cultures of fetal rat hippocampal neurons. The extract significantly elevated axonal length, number of secondary axonal collateral branches, length of primary dendrites and number of secondary dendritic branches by 58%, 8 folds, 68% and 2.6 folds, respectively, as compared to control [It was reported that an extract of  control .D. dichotoma has been reported. A methanol extract of D. dichotoma at a concentration of 1.3–6.5 mg/mL showed significant (p < 0.05) inhibition of cholinesterase enzyme (54.42%), compared to standard donepezil (cholinesterase inhibitor) (57.57%) at a concentration of 0.40–4.15 mg/mL [Alzheimer’s disease is a common neurologic disorder, responsible for brain shrinkage and cell death. It is the most prevalent type of dementia and one of the top four causes of mortality in developed countries . So far,15 mg/mL . C. racemosa and S. polycystum at various concentrations (0.0125–0.2 mg/mL) have been determined. Solvent extracts of C. racemosa and S. polycystum indicated anti-acetylcholinesterase activities with IC50 values ranging from 0.086–0.115 mg/mL, while C. racemosa extracts indicated anti-butyrylcholinesterase activity with an IC50 value of 0.156 mg/mL [The acetylcholinesterase and butyrylcholinesterase inhibitory activities of 56 mg/mL .A summary regarding the protective nature of the above-mentioned North Bornean seaweeds is shown in Seaweed nutraceutical bioactive compounds have great potential in biomedical and pharmaceutical applications ,90,91,92The information was retrieved from multiple internet databases such as ScienceDirect, PubMed, Wiley, ACS publications, etc., and registers including theses and proceedings. Records were searched with keywords related to seaweed, North Borneo, distribution, taxonomy, bioactivity, secondary metabolites, and diseases. Around 250 records approximately from the year 2000 to 2021 were retrieved and screened. Among these, about 100 records were excluded due to being out of the scope of the review. Eventually, a total of 149 records were adopted for the current review paper, and data from organizations such as the World Health Organization were included as well.K. alvarezii), eicosapentaenoic acid, palmitoleic acid, fucoxanthin, β-carotene (E. denticulatum), eucalyptol, oleic acid, dodecanal, pentadecane, (H. durvillaei), canthaxanthin, pentadecanoic acid, eicosane (C. lentillifera), pseudoephedrine, palmitic acid, monocaprin (C. racemosa), dictyohydroperoxide, squalene, fucosterol, saringosterol (D. dichotoma) and lutein, neophytadiene, cholest-4-en-3-one, and cis-vaccenic acid (S. polycystum). In the current review, the protective effects of North Bornean seaweeds in terms of anti-inflammatory, antioxidant, antimicrobial, anticancer, anti-obesity and cardiovascular protection, neuroprotection, and renal protection as well as hepatic protection were described and followed by nutraceutical profiling. The protective roles of the seaweeds might be due to the presence of a wide variety of nutraceuticals including phthalic anhydride, 3,4-ethylenedioxythiophene, 2-pentylthiophene, furoic acid (K. alvarezii and E. denticulatum are widely cultivated, developed as a functional food source, and used for carrageenans production. Locally, C. lentillifera and C. racemosa are also consumed as a nutrition source. For future perspectives, studies on functional food development and cultivation techniques are highly recommended. Furthermore, extensive studies on the seaweed isolates are needed to understand their bioactivity and mechanisms of action, while highlighting their commercialization potential.Despite their excellent pharmacological characteristics, only


Increasing evidence emphasizes the implications of dysregulated apoptosis and autophagy cellular processes in coronary artery disease (CAD). Herein, we aimed to explore apoptosis- and autophagy-related long noncoding RNAs (lncRNAs) in peripheral blood of CAD patients. p value < 0.05, differentially expressed apoptosis- and autophagy-related mRNAs were screened between CAD and healthy blood samples. Also, differentially expressed lncRNAs were identified for CAD. Using the psych package, apoptosis- and autophagy-related lncRNAs were defined with Spearson's correlation analysis. Receiver operating characteristic (ROC) curves were conducted for the assessment of the diagnosed efficacy of these apoptosis- and autophagy-related lncRNAs.  The mRNA and lncRNA expression profiles were retrieved from the Gene Expression Omnibus (GEO) database. With ∣fold change | >1.5 and adjusted  Our results showed that 24 apoptosis- and autophagy-related mRNAs were abnormally expressed in CAD than normal controls. 12 circulating upregulated and 1 downregulated apoptosis- and autophagy-related lncRNAs were identified for CAD. The ROCs confirmed that AC004485.3 (AUC = 0.899), AC004920.3 (AUC = 0.93), AJ006998.2 (AUC = 0.776), H19 (AUC = 0.943), RP5-902P8.10 (AUC = 0.956), RP5-1114G22.2 (AUC = 0.883), RP11-247A12.1 (AUC = 0.885), RP11-288L9.4 (AUC = 0.928), RP11-344B5.2 (AUC = 0.858), RP11-452C8.1 (AUC = 0.929), RP11-565A3.1 (AUC = 0.893), and XXbac-B33L19.4 (AUC = 0.932) exhibited good performance in differentiating CAD from healthy controls.  Collectively, our findings proposed that circulating apoptosis- and autophagy-related lncRNAs could become underlying diagnostic markers for CAD in clinical practice. Coronary artery disease (CAD), as a commonly diagnosed heart disease, contributes to the dominant cause of cardiovascular-related deaths . This diApoptosis and autophagy, as two types of programmed cellular deaths, are both involved in the development of CAD . Undue ahttps://www.ncbi.nlm.nih.gov/geo/query/acc.cgi) according to the following criteria: organism—Homo sapiens; experiment type—noncoding RNA profiling by array; and disease—CAD. As a result, two datasets including GSE113079 and GSE69587 datasets were obtained for this study. The GSE113079 dataset included 93 CAD and 48 healthy blood samples based on the GPL20115 platform [The mRNA and lncRNA expression profiles of CAD patients and healthy controls were searched from the Gene Expression Omnibus  | >1.5 and adjusted heatmaps .https://www.kegg.jp/) [Genes in autophagy (entry: map04140) and apoptosis (entry: map04210) were obtained from the Kyoto Encyclopedia of Genes and Genomes database  . They wehttp://string-db.org/) [Physical or functional interactions between specified proteins were analyzed via the STRING online tool (db.org/) . Requiredb.org/) . Connectdb.org/) .p value < 0.05 with at least 50% of differentially expressed autophagy- and apoptosis-related mRNAs were considered as differentially expressed autophagy- and apoptosis-related lncRNAs.Spearson's correlation analysis between differentially expressed lncRNAs and differentially expressed autophagy- and apoptosis-related mRNAs was presented via the psych package in R. lncRNAs with correlation The expression of differentially expressed autophagy- and apoptosis-related lncRNAs was externally verified in blood samples from 5 CAD patients and 5 healthy controls in the GSE169256 dataset. Moreover, associations between their expression and clinical features (age) were analyzed by Spearson's correlation tests. Their expression was also compared between male and female patients.Based on the expression profiles of the differentially expressed autophagy- and apoptosis-related lncRNAs, relative operating characteristic curves (ROCs) were conducted via the pROC package in R in the GSE113079 dataset .To explore CAD-related mRNAs, we screened abnormally expressed mRNAs between 93 CAD and 48 healthy blood samples in the GSE113079 dataset. Firstly, we normalized the microarray data via the limma package Figures . 988 up-To find autophagy- and apoptosis-related mRNAs in CAD, we overlapped the abnormally expressed mRNAs and autophagy- and apoptosis-related mRNAs. As a result, 24 mRNAs were identified for CAD , as follCirculating lncRNAs have been considered as diagnosed biomarkers for CAD . Herein,Since the GSE69587 dataset has been standardized, this study no longer standardized the dataset. In total, 430 circulating lncRNAs were upregulated and 305 circulating lncRNAs were downregulated in CAD compared to healthy samples Figures . The topp < 0.05; p < 0.05). Meanwhile, JUN and ITPR3 had positive correlations with downregulated LOC338758 (both p < 0.05). Thus, these lncRNAs could be distinctly related to autophagy and apoptosis in CAD.We analyzed the correlation between 13 abnormally expressed circulating lncRNAs and autophagy- and apoptosis-related mRNAs. Herein, we found that PRKACA, PIK3R2, and NGF were positively related to the 12 upregulated lncRNAs , AJ006998.2 , H19 , RP11-247A12.1 , RP11-288L9.4 , RP11-344B5.2 , RP11-452C8.1 , RP11-565A3.1 , RP5-1114G22.2 , RP5-902P8.10 , and XXbac-B33L19.4  are as follows: AC004485.3 , and XXbac-B33L19.4 (AUC = 0.932) exhibited good performance to differentiate CAD from healthy controls. The above findings concerning circulating lncRNAs might possess effective diagnostic value on CAD, thereby reducing mortality. Among them, circulating H19 is correlated to risk of CAD among a Chinese cohort [On account of the shortcomings of current diagnostic markers on CAD, circulating lncRNAs appear to have attracted close attention. After verification, our data demonstrated that AC004485.3 (AUC = 0.899), AC004920.3 (AUC = 0.93), AJ006998.2 (AUC = 0.776), H19 (AUC = 0.943), RP5-902P8.10 (AUC = 0.956), RP5-1114G22.2 (AUC = 0.883), RP11-247A12.1 (AUC = 0.885), RP11-288L9.4 (AUC = 0.928), RP11-344B5.2 (AUC = 0.858), RP11-452C8.1 (AUC = 0.929), RP11-565A3.1  were distinctly upregulated in CAD compared to healthy controls. More importantly, they had good performance in distinguishing CAD from healthy individuals. Thus, these circulating lncRNAs could be promising diagnostic markers for CAD.


Flavobacterium strains isolated from a remote Antarctic island, James Ross Island, were studied using a polyphasic taxonomic approach to determine their taxonomic position. Phylogenetic analyses based on the 16S rRNA gene and 92 core genes clearly showed that these strains formed two distinct phylogenetic clusters comprising three and five strains, with average nucleotide identities significantly below 90% between both proposed species as well as between their closest phylogenetic relatives. Phenotyping revealed a unique pattern of biochemical and physiological characteristics enabling differentiation from the closest phylogenetically related Flavobacterium spp. Chemotaxonomic analyses showed that type strains P4023T and P7388T were characterized by the major polyamine sym-homospermidine and a quinone system containing predominantly menaquinone MK-6. In the polar lipid profile phosphatidylethanolamine, an ornithine lipid and two unidentified lipids lacking a functional group were detected as major lipids. These characteristics along with fatty acid profiles confirmed that these species belong to the genus Flavobacterium. Thorough genomic analysis revealed the presence of numerous cold-inducible or cold-adaptation associated genes, such as cold-shock proteins, proteorhodopsin, carotenoid biosynthetic genes or oxidative-stress response genes. Genomes of type strains surprisingly harbored multiple prophages, with many of them predicted to be active. Genome-mining identified biosynthetic gene clusters in type strain genomes with a majority not matching any known clusters which supports further exploratory research possibilities involving these psychrotrophic bacteria. Antibiotic susceptibility testing revealed a pattern of multidrug-resistant phenotypes that were correlated with in silico antibiotic resistance prediction. Interestingly, while typical resistance finder tools failed to detect genes responsible for antibiotic resistance, genomic prediction confirmed a multidrug-resistant profile and suggested even broader resistance than tested. Results of this study confirmed and thoroughly characterized two novel psychrotrophic Flavobacterium species, for which the names Flavobacterium flabelliforme sp. nov. and Flavobacterium geliluteum sp. nov. are proposed.Despite unfavorable Antarctic conditions, such as cold temperatures, freeze-thaw cycles, high ultraviolet radiation, dryness and lack of nutrients, microorganisms were able to adapt and surprisingly thrive in this environment. In this study, eight cold-adapted  For a long time, Antarctica was considered an inhospitable environment with low biodiversity. Although it is a part of Earth’s cryosphere which covers about 20% of the Earth’s surface , it has Bacteroidetes   . Its mem05.2021) . In addic losses .Flavobacterium spp. in Antarctica is supported by their adaptation and coping mechanisms required for survival in harsh conditions including low temperatures, lack of nutrients or intense UV exposure, and freeze-thaw cycles that exert a strong evolutionary pressure on microbial cells . Microbiological sampling is conducted yearly on the James Ross Island (near the north-eastern extremity of the Antarctic Peninsula), Antarctica. Sampling is performed during the summer period when the majority of the island becomes an ice-free area. Antarctic summer season allows sampling of upper soil layers, permafrost, lakes and proglacial streams, which are all areas of intense microbial activities  sampled during the years 2010–2019  for parallel testing and comparison purposes. All strains were routinely cultivated on R2A agar plates at 20°C for 48 hrs.The present study describes a taxonomic investigation of eight  strains  isolatedtivities . Flavoba010–2019 . Water s3. The 16S rRNA sequences were submitted to the EzBioCloud server  software (v7.0) (Genomic DNA was extracted and purified using High Pure PCR Template Preparation Kit (Roche Diagnostics) according to the manufacturer’s recommendations. The 16S rRNA genes were amplified and sequenced using universal bacterial primers pA (5′-AGAGTTTGATCCTGGCTCAG-3′) and pH (5′-AAGGAGGTGATCCAGCCGCA-3′) . PCR prod server  for inite (v7.0) . The maxde novo genome assembly. The MismatchCorrector was run to reduce short indels and the number of mismatches. The read coverage cut-off value was automatically computed by the software resulting in contigs and scaffolds. Quality assessment was performed by the QUAST tool  with 7 min of enzymatic fragmentation and final fragments length within the range of 550–650 bp. Sequencing was performed on Illumina MiSeq platform with MiSeq Reagent Kit v2 (500-cycles) . The St. Petersburg genome assembler (SPAdes v3.11.1)  was usedAST tool  and BowtAST tool .Flavobacterium spp. The WGS data of type strains P4023T and P7388T were submitted to the Type Strain Genome server (TYGS)  values were calculated using the ANI calculator on the Kostas Laboratory website5 using reciprocal best hits (two-way ANI) (6 (The whole genome sequence (WGS) data were analyzed for further confirmation of the taxonomic status of the analyzed strains and for comparison with genomes of related r (TYGS)  and to tr (TYGS) . To furtpipeline  with defway ANI) . Proteinway ANI)  and subj ANI)  8 with CARD v3.1.1 database   and Operdatabase , NCBI Badatabase , MEGAResdatabase  and ARG-database . Furtherdatabase . The enzdatabase .T and P7388T, was also examined by transmission electron microscopy using a Morgagni 268D Philips (FEI Company) electron microscope.The colony morphology was determined on R2A agar after 48 h cultivation at 20°C. The cellular morphology of all strains was observed by light microscopy after Gram staining. The cellular morphology of the type strains, P40232PO4/0.1 M NaOH; pH 9.0–10.0, 0.1 M NaHCO3/0.1 M Na2CO3. The pH value of the TSB was confirmed after autoclaving.The growth at different temperatures  and the salt tolerance  were determined on R2A agar plates. The pH tolerance was assessed in Trypticase Soya Broth (TSB) inoculated with two drops of cell suspension of concentration as McFarland 2.0 with pH 5–10 at intervals of 1.0 pH unit at 20°C adjusted with the following buffer systems: pH 5.0–8.0, 0.1 M KHF. hercynium CCM 9054T, F. branchiicola CCM 9061T, F. chilense CCM 7940T, F. araucananum CCM 7939T, F. psychroterrae CCM 8827T and F. saccharophilum CCM 8770T were phenotypically characterized by the most relevant tests recommended for description of new taxa within the family Flavobacteriaceae (2S on Triple Sugar Iron Agar (HiMedia), hydrolysis of aesculin, starch   , phosphoe (ONPG) , nitratee (ONPG) , utilizae (ONPG)  and sodie (ONPG) . The pree (ONPG) .2, both on R2A agar plates. All above listed tests were inoculated using cells of analyzed strains grown on R2A agar at 20°C for 48 hrs. All tests were read daily for up to 7 days with exception of the tyrosine hydrolysis test (up to 10 days). The utilization of carbon sources, enzyme activities and other additional phenotypic characteristics were further tested using identification test kits GEN III MicroPlateTN (Biolog) with the protocol A, API 20 NE and API ZYM (bioMérieux) according to the manufacturer’s instructions.Capability of growth on different media was tested on R2A agar (Oxoid), Nutrient agar (Oxoid), Plate Count agar (Oxoid), Tryptone Soya agar (Oxoid), marine agar , MacConkIn vitro antimicrobial susceptibility was assessed using the disk diffusion method. Cell suspensions of concentration as 0.5 McFarland were prepared from cultures cultivated on R2A agar. Suspension containing each strain (100 μl) was spread on the Mueller-Hinton agar plates. Tested antibiotic disks  were as follows: ampicillin (10 μg), aztreonam (30 μg), carbenicillin (100 μg), cefixime (5 μg), ceftazidime (10 μg), cephalotin (30 μg), ciprofloxacin (5 μg), gentamicin (10 μg), chloramphenicol (30 μg), imipenem (10 μg), kanamycin (30 μg), co-trimoxazole (25 μg), piperacillin (30 μg), polymyxin B (300 U), streptomycin (10 μg) and tetracycline (30 μg). CLSI  and EUCAST  standards were strictly followed for cultivation and inhibition zone diameter reading , F. hercynium DSM 18292T (98.54%) and F. chilense LMG 26360T (98.39%). A proposed type strain for the second cluster, P7388T, showed the highest sequence similarities to the F. branchiicola 59B-3-09T (99.12%), F. araucananum DSM 24704T (99.08%) and F. psychroterrae CCM 8827T (99.04%). The 16S rRNA gene sequence similarities of strain P4023T fell below the cut-off value 98.65%, whereas strain P7388T reached similarities above this threshold. Nevertheless, several studies have already proved that the standard 16S rRNA gene sequence cut-off value is not sufficient for some genera, such as Streptomyces, Streptococcus or Bacillus  could not be assigned to a specific class of orthologous genes clusters (COGs) or with specific function  . Howeverhnsoniae .T contained 112 scaffolds with a draft genome size of 4,387,206 bp and the genomic G + C content 34.5 mol% . Prediction of PULs found seven putative loci involved in degradation of carrageenan, dextran, hemicellulose, chitin, pectin, starch and xylan, which implies the ability of strain P7388T to degrade various polysaccharide substrates.Functional annotation revealed that 485 genes (12.97%) could not be assigned to any COGs or with specific function  . The remT and P7388T. The analysis indicated that these strains harbor four (P4023T) and seven (P7388T) gene clusters potentially related to secondary metabolite production. Out of these, six clusters did not match any known BGCs deposited in the MIBiG database -non-ribosomal peptide synthetase (NRPS) BGC for strain P7388T that did not match any known BGCs   , 3. ThisF. hercynium DSM 18292T, F. saccharophilum DSM 1811T and F. pectinovorum DSM 6368T. Genome-mining focused on cold-adaptation confirmed that Flavobacterium spp. are well adjusted to environmental stress and harbor a significant number of genes associated with cold-adaptation regardless of their thermotype , transcription termination/antitermination factor (nusA), translation initiation factors IF1 and IF2 , polynucleotide nucleotidyltransferase (pnp), heat-shock cognate proteins (hsc), recombinase A (recA) and many others , a member of family Flavobacteriaceae, was found to substantially increase tolerance to freezing and thus considered a primary response to ensure freeze-tolerance of its hosts , chromosomal replication initiation ATPase (dnaA), DNA topoisomerases IV , transcription antitermination factor (nusA), translation factors IF-1, 2, 3  along with other cold-inducible genes were present in all genomes in one copy, regardless of the thermotype (rpoD) and sigma factor 24 (rpoE) were present in genomes only in one and two copies, respectively, although psychrotrophic Flavobacterium spp. were previously found to harbor multiple copies of these genes , superoxide-dismutase and peroxidase , or thioredoxin and peroxiredoxin reductases . Although all analyzed strains harbored peroxiredoxin genes in multiple copies , strain P4023T harbored multiple copies of superoxide-dismutase (sodA) and thioredoxin (trxA), which may be explained by its isolation from surface rather than deeper soil layers and therefore exposed to higher oxidative stress and stronger UV radiation. Similarly versatile microorganisms to flavobacteria are pseudomonads that colonize diverse Antarctic areas including soils, fresh or marine waters  by Pseudomonas extremaustralis 14-3T (Cold-adaptation is inseparably associated with oxidative stress response as a result of higher oxygen solubility at lower temperatures leading to increased levels of reactive oxygen species (ROS). Notably, e waters . Considee waters . Howeveris 14-3T . None ofT harbored a crtZ gene required for zeaxanthin biosynthesis  is a membrane light-driven protein acting as an outward Hon pumps . Both th retinal . PRs wer retinal  and inte retinal . Neverth. MED134 , which mFlavobacterium strains, since the type II CRISPR/Cas-system was found to be an adaptive immune system in other cold-adapted flavobacteria  were predicted in the P4023T genome likely to provide resistance against 16 different drug classes (T genome (in vitro antibiotic susceptibility testing. Correlation between resistome predictions and in vitro testing was found for resistance to aminoglycosides which is presumably encoded by aadS gene (MBP4138141.1) in the genome of P7388T, a widely spread bacterial adenylyltransferase conferring resistance to aminoglycoside antibiotics (JOHN-1 present in P4023T (MBP4142760.1) and P7388T (MBP4137930.1) genomes. This gene provides resistance to a broad spectrum of β-lactam antibiotics in Flavobacterium johnsoniae (in vitro susceptibility results. Additive effects on resistance to β-lactam antibiotics may result from the presence of putative OXA beta-lactamase OXA-29 (MBP4140625.1) in P4023T, known to hydrolyze penams and cephalosporines but is not effective against carbapenems , ciprofloxacin (5 μg) and chloramphenicol (30 μg). Disagreement between the phenotypic susceptibility pattern and ARGs computational prediction is a well-known controversy, especially in clinical bacteriology (in vivo vs. in vitro), testing of single strain culture vs. biofilm formation, presence of specific metabolites and growth factors, presence of specific phages or inadequate time/concentration of tested antibiotics that induce expression of resistance genes. Any of above-mentioned factors or even their combination may be also responsible for discrepancies observed among analyzed Antarctic Flavobacterium isolates, in particular considering extreme environment they colonize and presence of phages in their genomes.Although routine screening did not find any resistance genes, RGI prediction of the resistome from draft genomes of P4023 classes . InactivT genome , they maibiotics . All Antibiotics . This wihnsoniae , howeverbapenems . Both steriology . There aT additionally harbors a putative rosA gene (MBP4140897.1) encoding an efflux pump/potassium antiporter system. This efflux transporter has been detected in marine Flavobacterium spp. (Bacteroidetes (rosA gene is related to polymyxin B resistance and represents a part of a two component efflux antiporter system (RosAB) found in Yersinia enterocolitica and confers resistance to cationic antimicrobial peptides including polymyxin B , which is a well-known environment with resistant bacteria . AlthougT and P7388T were aerobic, Gram-negative rods with rounded ends, with average cell size 0.4–0.6 μm × 1.2–2.4 μm and 0.3–0.4 μm × 1.5–3.0 μm, respectively , Summed Feature 3 (C16:1 ω7c/C16:1 ω6c) (9.9%), anteiso-C15:0 (9.8%), iso-C15:0 (9.3%) and iso-C16:0 3OH (9.3%). The major fatty acid of the group comprising strain P7388T were iso-C15:0 (23.4%), Summed Feature 3 (C16:1 ω7c/C16:1 ω6c) (9.9%), iso-C17:0 3OH (8.5%) and iso-C15:0 3OH (8.4%). The proposed species differ not only qualitatively and quantitatively in the profiles of major fatty acids, but also quantitatively in minor fatty acids, especially iso-C14:0, iso-C16:1 H and Summed Feature 9 (C16:0 10-methyl/iso-C17:1). The overall fatty acids profile of both groups is in agreement with the genus description ] and minor amounts of putrescine [0.7 μmol (g dry weight–1)] and spermidine [0.2 μmol (g dry weight–1)]. The polyamine pattern of strain P7388T was similar with the major polyamine sym-homospermidine [20.5 μmol (g dry weight–1)] and minor amounts of putrescine [0.2 μmol (g dry weight–1)] and spermidine [0.3 μmol (g dry weight–1)]. Quinone systems, polar lipid profile and polyamine pattern are well in agreement with the genus description .L-tyrosine agar. Negative for hydrolysis of Tween 80, aesculin, ONPG, starch, DNA, CMC and agar. Does not produce lecithinase. Does not produce H2S. Positive for arginine dihydrolase, and negative for ornithine and lysine decarboxylases. Does produce acid from glucose and maltose in aerobic conditions. Negative for production of acid from fructose and xylose in aerobic conditions. Positive for utilization of glucose and maltose by API 20 NE. Negative for utilization of arabinose, mannose, N-acetylglucosamine, gluconic acid, capric acid, adipic acid, malic acid, citric acid and phenylacetic acid by API 20 NE. Positive for alkaline phosphatase, leucine arylamidase, valine arylamidase and acid phosphatase by API ZYM. Negative for esterase (C 4), esterase lipase (C 8), lipase (C 14), cystine arylamidase, trypsin, α-chymotrypsin, naphtol-AS-BI-phosphohydrolase, α-galactosidase, β-galactosidase, β-glucuronidase, α-glucosidase, β-glucosidase, N-acetyl-β-glucosaminidase, α-mannosidase, and α-fucosidase by API ZYM. Carbon source utilization ability via respiration determined by Biolog GEN III MicroPlate test panels is positive for D-maltose, α-D-glucose, D-glucose-6-PO4, gelatine, L-arginine, L-aspartic acid, L-glutamic acid, acetoacetic acid and acetic acid. Negative for D-trehalose, D-cellobiose, gentiobiose, sucrose, stachyose, D-raffinose, α-D-lactose, D-melibiose, β-methyl-D-glucoside, D-salicin, N-acetyl-β-D-mannosamine, N-acetyl neuraminic acid, D-mannose, D-fructose, D-galactose, 3-methyl glucose, D-fucose, L-fucose, L-rhamnose, inosine, D-sorbitol, D-mannitol, D-arabitol, myo-inositol, glycerol, D-fructose-6-PO4, D-aspartic acid, D-serine, L-alanine, L-histidine, L-pyroglutamic acid, L-serine, D-galacturonic acid, D-galactonic acid lactone, D-gluconic acid, D-glucuronic acid, glucuronamide, mucic acid, quinic acid, D-saccharic acid, p-hydroxy phenylacetic acid, methyl pyruvate, D-lactic acid methyl ester, L-lactic acid, citric acid, α-keto glutaric acid, D-malic acid, L-malic acid, bromo-succinic acid, Tween 40, γ-amino-butyric acid, α-hydroxy-butyric acid, β-hydroxy-D,L-butyric acid, α-keto butyric acid, propionic acid and formic acid.Cells are Gram-stain-negative rods with rounded ends, cell size in range 0.4–0.6 μm × 1.2–2.4 μm, cells occurring in irregular clusters, occasionally singly or in pairs. Endospores are not formed. Does not produce a capsule. Negative for presence of flexirubin-type pigments. Does not adhere to agar. Yellowish colonies. Motile with gliding-activity. Catalase and oxidase positive. Growth occurs on R2A, PCA, TSA, marine agar NA, blood agar with 5% sheep blood, BHI, Mueller-Hinton and Endo agar. Does not grow on MacConkey agar. Grows in pH range 6–9 and temperature range 1–30°C with optimum growth at 20°C and pH around 7. Cells grow well in presence of 1% NaCl, and 2% NaCl inhibits growth. Does grow in microaerophilic conditions but growth in anoxic conditions is limited. Does not produce fluorescein on King B medium. No utilization of Simmon’s citrate, malonate and acetamide. Negative for reduction of nitrates and nitrites. Negative for production of urease and indole. Positive for hydrolysis of gelatine, casein, and tyrosine. Does not produce brown diffusible pigment on 15:1 ω6c, Summed Feature 3 (C16:1 ω7c/ C16:1 ω6c), anteiso-C15:0, iso-C15:0 and iso-C16:0 3OH. Major respiratory quinone is MK-6 and major polyamine is sym-homospermidine. Polar lipid profile contains major amounts of phosphatidylethanolamine, an ornithine lipid, and two unidentified lipids  lacking a functional group, moderate amounts of unidentified lipid L1, unidentified glycolipid GL, minor amounts of an unidentified aminophospholipid (APL), and two unidentified lipids . The DNA G + C content of the type strain is 31.2 mol%.The major fatty acids are iso-CT (= CCM 9062T = LMG 31963T) was isolated in 2011 from the organic material of an abandoned bird nest located at the Lachman Cape . All characteristics listed in the species description are shared by all strains, except for the following strain-dependent test results: motility, production of acid from mannitol, and variable results observed by Biolog GEN III MicroPlate were utilization of dextrin, D-turanose, N-acetyl-D-glucosamine, glycyl-L-proline and pectin. A formal proposal of the species “Flavobacterium flabelliforme sp. nov.” is given in Type strain P4023Flavobacterium flabelliforme sp. nov. P4023T are MW691162 and JAGFBU000000000, respectively.The GenBank/EMBL/DDBJ accession numbers for the near full length 16S rRNA gene sequences and whole genome sequences for Flavobacterium geliluteum .L-tyrosine agar. Negative for hydrolysis of DNA and agar. Does not produce lecithinase. Does not produce H2S. Positive for arginine dihydrolase and negative for ornithine and lysine decarboxylases. Does produce acid from glucose, maltose and xylose in aerobic conditions. Negative for production of acid from mannitol in aerobic conditions. Positive for utilization of glucose, arabinose, mannose, N-acetyl-glucosamine, maltose, and hydrolysis of aesculin by API 20 NE. Negative for utilization of gluconic acid, capric acid, adipic acid, malic acid, citric acid and phenylacetic acid by API 20 NE. Positive for alkaline phosphatase, leucine arylamidase, acid phosphatase, naphtol-AS-BI-phosphohydrolase and β-glucosidase by API ZYM. Negative for esterase (C 4), esterase lipase (C 8), lipase (C 14), valine arylamidase, cystine arylamidase, trypsin, α-chymotrypsin, α-galactosidase, β-galactosidase, β-glucuronidase, N-acetyl-β-glucosaminidase, α-mannosidase and α-fucosidase by API ZYM. Carbon source utilization ability via respiration determined by Biolog GEN III MicroPlate test panels is positive for D-trehalose, D-cellobiose, gentiobiose, N-acetyl-D-glucosamine, N-acetyl-D-galactosamine, α-D-glucose, D-mannose, D-glucose-6-PO4, glycyl-L-proline, L-aspartic acid, L-glutamic acid, D-galacturonic acid, acetoacetic acid and acetic acid. Negative for sucrose, stachyose, D-raffinose, α-D-lactose, D-melibiose, N-acetyl-β-D-mannosamine, N-acetyl neuraminic acid, 3-methyl glucose, D-fucose, L-fucose, L-rhamnose, inosine, D-sorbitol, D-mannitol, D-arabitol, myo-inositol, glycerol, D-aspartic acid, D-serine, L-alanine, D-gluconic acid, D-glucuronic acid, glucuronamide, D-saccharic acid, quinic acid, p-hydroxy phenylacetic acid, L-lactic acid, α-keto glutaric acid, D-malic acid, L-malic acid, bromo-succinic acid, γ-amino-butyric acid, α-hydroxy-butyric acid, β-hydroxy-D,L-butyric acid, α-keto butyric acid, propionic acid, and formic acid.Cells are Gram-stain-negative rods with rounded ends, cell size in range 0.3–0.4 μm × 1.5–3.0 μm, occurring singly and in irregular clusters, occasionally in pairs. Endospores are not formed. Does not produce a capsule. Dark yellow to orange pigmented colonies. Flexirubin-type pigment present. Does not adhere to agar. Motile with gliding-activity. Catalase positive. Oxidase negative. Growth occurs on R2A, PCA, TSA, NA, blood agar with 5% sheep blood, BHI and Mueller-Hinton agar. Does not grow on marine and MacConkey agar. Grows in pH range 6–8 and temperature range 15–30°C with optimum growth at 20°C and pH around 7. Cells grow in presence of 0.5% NaCl, and 1% NaCl inhibits growth. Does grow in microaerophilic conditions but growth in anoxic conditions is limited. Does not produce fluorescein on King B medium. No utilization of Simmon’s citrate, malonate and acetamide. Negative for reduction of nitrates and nitrites. Negative for production of urease and indole. Positive for hydrolysis of gelatine, aesculin, ONPG, starch, casein, tyrosine and CMC. Does not produce brown diffusible pigment on 15:0, Summed Feature 3 (C16:1 ω7c/C16:1 ω6c), iso-C17:0 3OH and iso-C15:0 3OH. Major respiratory quinone is menaquinone MK-6 and major polyamine is sym-homospermidine. Polar lipid profile contains major lipids phosphatidylethanolamine, an ornithine lipid, two unidentified lipids lacking a functional group , moderate amounts of unidentified lipid L1, minor amounts of lipids L2 and L6 and an unidentified glycolipid (GL). The DNA G + C content of the type strain is 34.5 mol%.The major fatty acids are iso-CT (= CCM 9064T = LMG 31962T) was isolated from water samples taken in 2016 from a small temporary lake . All characteristics listed in the species description are shared by all strains, except for the following strain-dependent test results: growth at 5 and 10°C, growth on Endo agar, hydrolysis of Tween 80, production of acid from fructose, presence of α-glucosidase in API ZYM and variable results observed by Biolog GEN III MicroPlate were utilization of dextrin, D-maltose, pectin, β-methyl-D-glucoside, D-salicin, D-fructose, D-galactose, D-fructose-6-PO4, gelatine, L-arginine, D-galactonic acid lactone, mucic acid, methyl pyruvate, D-lactic acid methyl ester, citric acid and Tween 40. A formal proposal of the species “Flavobacterium geliluteum sp. nov.” is given in Type strain P7388Flavobacterium geliluteum sp. nov. P7388T are MW691150 and JAGFBV000000000, respectively.The GenBank/EMBL/DDBJ accession numbers for the near full length 16S rRNA gene sequences and whole genome sequences for https://www.ncbi.nlm.nih.gov/genbank/, MW691162; https://www.ncbi.nlm.nih.gov/genbank/, JAGFBU000000000; https://www.ncbi.nlm.nih.gov/genbank/, MW691150; https://www.ncbi.nlm.nih.gov/genbank/, JAGFBV000000000.The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found below: SK performed phylogenetic, phylogenomic, genomic analysis, analysis of fatty acid-methyl esters, and drafted and finalized the manuscript. H-JB performed chemotaxonomic analyses of polar lipids, menaquinones, and polyamines. MB was responsible for whole-genome sequencing. MS-P was involved in genomic analysis of biosynthetic potential. MN was involved in genome assembly and quality of WGS data. DK performed electron microscopy. ES and IS performed morphological, physiological, and biochemical characterization including antibiotic susceptibility. SK drafted the manuscript with inputs of H-JB, MB, MS-P, and IS. All authors edited the draft manuscript and agreed to the final manuscript version for submission.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.