The current study was designed to evaluate the role of semi-quantitative EUS- elastography (strain ratio) in staging malignant pancreatic lesions.Pancreatic cancer is considered one of the most lethal malignancies with a survival rate of only 5% worldwide. Pancreatic lesions include a wide range of diagnoses from benign to malignant forms. Biopsy and pathological study are the gold standard for the differentiation of malignant lesions and staging of tumors. Recently, endoscopic ultrasound sonography (EUS) elastography has been noticed as a non-invasive diagnosis modality. Nevertheless, no evidence of its potential to determine different stages of malignant tumors is available.This prospective study included 81 adult patients with a confirmed diagnosis of malignant pancreatic lesion in different clarified stages. All diagnoses were confirmed after endoscopic ultrasound sonography via pathological investigation of surgical specimens or needle biopsies. The results of EUS-elastography based on tumor size (T staging), involved lymph nodes (N staging), and metastasis (M staging) were compared with the gold standard. p=0.57, p=0.92, p=0.11, respectively). Moreover, the Spearman rank correlation coefficients for the correlation between T staging, N staging, M staging and SR elastography were not significant . The mean age of patients was 60.11±13.57 years. The mean SR elastography value was 52.78±48.97. Elastography could not significantly discriminate T stage, N stage, or M stage of tumors (The non-invasive modality EUS-elastography cannot replace the gold standard in staging tumors; however, EUS-elastography seemed to differentiate benign lesions from malignant ones.  Presurgical diagnosis of solid pancreatic lesions (SPLs) remains a major clinical challenge and controversial subject for gastroenterologists. With a survival rate of less than 5% for 5 years, malignant pancreatic tumors make the most important group of SPLs . PancreaSince 2006, EUS elastography has been commonly used to approach solid pancreatic masses. This new ultrasound technique enables measuring the firmness of the target lesion non-invasively , 10. EUSThis prospective study was conducted to evaluate the benefit of using EUS elastography to non-invasively assess pancreatic tumor staging. As the next step, the relationship between tumor size and elastography finding was also considered to determine the accuracy of this technique in the improvement of tumor classification. PatientsThis prospective single-center study was organized in Taleghani Hospital , a referral center for pancreatic disorders in Tehran, Iran. The population of this study included 81 patients who were diagnosed with malignant pancreatic tumors and underwent EUS-elastography in our center between October 2017 and 0ctober 2019. Patients younger than 18 years of age, had other types of pancreatic lesions, and those with liquid and cystic components were excluded from the study. CT-SCAN, EUS FNA biopsy, surgical specimen, and pathological findings were used to diagnose pancreatic cancer. Written informed constant was obtained from all participating patients. This investigation was approved by the Ethics Committee of Shahid Beheshti University of Medical Sciences. The experiments were conducted according to the World Medical Association’s Declaration of Helsinki for human-involved studies.EUS procedure and methodsThe entire EUS procedure was performed by an expert endosonographist using Olympus EUME2  coupled with an electronic ultrasound probe Olympus GF-UE 180. To perform the FNA biopsy, a Cook needle 22G was applied during the EUS procedure . Pancreatic masses were assessed using elastography. All endosonographic images and elastography videos were recorded and evaluated by an endosonographist who was blind to the final pathology diagnosis but informed about clinical and EUS findings. Moreover, to enhance the accuracy of the investigation, all videos were rechecked by a second expert endosonographist who had no previous knowledge about either pathology findings or clinical and EUS findings. A quantitative score, which is indeed a semi-quantitative elastography score, was used to show strain ratio. In this method, two endosonographic fields were chosen. The strain ratio was determined by dividing the normal region into the region of interest. The strain ratio means were determined and utilized as the ultimate result of each patient.Patients were diagnosed according to a surgical specimen histology core needle biopsy or EUS-oriented fine needle aspiration (EUS-FNA). Contrast-improved multidetector computed tomography (CT) was performed, and the TMN staging score was recorded for all patients. Finally, the elastography findings, including quantitative scores, were compared with malignant mass sizes and cancer staging. For cancer staging, TMN staging was used and this score was calculated based on surgical, pathology, EUS, and CT-scan findings.Statistical Analysisp-value of less than 0.05  was considered statistically significant. Statistical analyses were conducted using R software version 3.6.3.EUS-elastography variables including tumor size, involved lymph nodes, and metastasis were described as frequency rates, percentages, and means and standard deviations. Tumor size means were compared using analysis of variance (ANOVA). Involved lymph nodes and metastasis were analyzed using an independent sample t-test. The correlation coefficients between factors were obtained by Spearman correlation test. A p=0.57) (p=0.40) (p=0.92), nor did it correlate with the elastography ratio (p=0.94) (p=0.11) (p=0.39) , most patients were observed to be in the T4 stage (60.5%), and only 3.7% of the participants were categorized in the T1 stage .. Accordp=0.57) . Further(p=0.40) . The fin(p=0.40) . Moreove(p=0.94) ; Table 3(p=0.94) . However(p=0.11) . NeverthThis prospective study was designed to evaluate the proficiency of elastography in discriminating different stages of solid pancreatic tumors. Malignant pancreatic lesions were studied according to stiffness score (strain ratio). It was observed that elastography could not be considered an effective modality for distinguishing between pancreatic tumor stages. Pancreatic lesions are usually detected incidentally or in routine radiographic evaluations, including ultrasound or CT-scan. The findings of these modalities include pancreatic masses, tumors, cysts, or benign nodules. Although most of these lesions are benign, their malignant forms should be surgically resected. Therefore, pathological investigations are vital in pancreatic lesions . SeveralThis study had several limitations, including a small population size, a single setting, and no comparison between different types of pancreatic lesions, including benign or malignant. Thus, we believe that this study could be used as a cornerstone for future analyses. This study investigated the role of EUS-elastography as a non-invasive diagnosis modality in distinguishing between different stages of pancreatic tumors in terms of tumor size, number of involved lymph nodes, and far metastasis. However, it appears that the studied modality had no significant effects in differentiating tumor stages in malignant lesions. Furthermore, the staging was not correlated with the SR of elastography.The authors declare that they have no conflict of interest.


There are no widely accepted pharmacologic treatments for frailty prevention. Since frailty is associated with inflammation, aspirin has the potential to reduce frailty. We investigated whether low-dose aspirin reduces incident frailty in participants of the ASPirin in Reducing Events in the Elderly (ASPREE) trial. In the U.S and Australia, 19,114 healthy community-dwelling individuals aged ≥70 years (U.S. minorities ≥65 years) were enrolled in ASPREE, a double-blind, placebo-controlled trial of 100mg daily low-dose aspirin vs. placebo. Frailty was defined according to a modified Fried frailty definition, and a frailty index which used a deficit accumulation model. Competing risk Cox proportional hazards models were used to compare time to incident frailty for aspirin vs. placebo. At baseline, 2.2% and 8.1% met criteria for frailty by Fried and frailty index criteria, respectively. Over a median of 4.7 years of follow-up, 2252 participants developed incident frailty according to Fried classification, and 4376 according to the frailty deficit accumulation index. There was no difference in the risk of incident frailty between individuals randomized to aspirin versus placebo according to either criteria . Change in frailty over time was not different between the aspirin and placebo treatment arms. The results were consistent across a series of sub-groups, including baseline frailty status. Based on these results, aspirin use in healthy older adults does not reduce incident frailty.


Bacillus amyloliquefaciens (OGBa) are all Gram-positive, endospore-forming, and rod-shaped. Taxonomically, the OGBa belongs to the Bacillus subtilis species complex, family Bacillaceae, class Bacilli, and phylum Firmicutes. To date, the OGBa comprises four bacterial species: Bacillus amyloliquefaciens, Bacillus siamensis, Bacillus velezensis and Bacillus nakamurai. They are widely distributed in various niches including soil, plants, food, and water. A resurgence in genome mining has caused an increased focus on the biotechnological applications of bacterial species belonging to the OGBa. The members of OGBa are known as plant growth-promoting bacteria (PGPB) due to their abilities to fix nitrogen, solubilize phosphate, and produce siderophore and phytohormones, as well as antimicrobial compounds. Moreover, they are also reported to produce various enzymes including α-amylase, protease, lipase, cellulase, xylanase, pectinase, aminotransferase, barnase, peroxidase, and laccase. Antimicrobial compounds that able to inhibit the growth of pathogens including non-ribosomal peptides and polyketides are also produced by these bacteria. Within the OGBa, various B. velezensis strains are promising for use as probiotics for animals and fishes. Genome mining has revealed the potential applications of members of OGBa for removing organophosphorus (OPs) pesticides. Thus, this review focused on the applicability of members of OGBa as plant growth promoters, biocontrol agents, probiotics, bioremediation agents, as well as producers of commercial enzymes and antibiotics. Here, the bioformulations and commercial products available based on these bacteria are also highlighted. This review will better facilitate understandings of members of OGBa and their biotechnological applications.Bacteria under the operational group  Bacillus amyloliquefaciens from the soil. The species is named after its unique character because it produced (faciens) a liquefying (lique) α-amylase (amylo) [B. amyloliquefaciens was combined with the closely related Bacillus subtilis and Bacillus licheniformis into the B. subtilis species complex, based on phylogenetic and phenetic evidence [B. subtilis species complex, it can be further sub-grouped into the operational group B. amyloliquefaciens (OGBa) that comprises four bacterial species; the soil-borne B. amyloliquefaciens, the plant-associated Bacillus siamensis and Bacillus velezensis, and a black-pigment-producing strain Bacillus nakamurai [In 1943, a Japanese scientist, Juichiro Fukumoto, first isolated  (amylo) . Later, Ba, namely B. amyloliquefaciens subsp. plantarum, Bacillus methylotrophicus and Bacillus oryzicola, were reclassified as strains of B. velezensis [B. velezensis belongs to a conspecific group consisting of B. velezensis, B. amyloliquefaciens subsp. plantarum FZB42 (reclassified as B. velezensis FZB42) and B. methylotrophicus. However, B. velezensis is distinct from the closely related species of B. amyloliquefaciens and B. siamensis [Ba have been deposited into the National Center Biological Information (NCBI) database  content, fatty acid methyl esters (FAME) and DNA–DNA hybridization (DDH) [Ba is constantly causing confusion to researchers, especially for non-professional taxonomy researchers.Bacterial species from the OGon (DDH) . TherefoBa within the B. subtilis species complex [gyrB), signal transduction histidine kinase CheA (cheA) and RNA polymerase β-subunit (rpoB) were used for the study of speciation within the B. subtilis species complex before the advent of multilocus sequence analysis (MLSA) [Ba has been solved by genome-based [Ba comprised four species: (i) B. amyloliquefaciens; (ii) B. siamensis; (iii) B. velenzensis; and (iv) B. nakamurai, as confirmed by cladistic analysis  ,13. The me-based  and geneme-based  phylogenanalysis ; Table 1Ba had been directly isolated from rare dormant volcanic soils [The ability to produce endospores when facing harsh conditions allowed the members of the operational group to survive in various niches including soil, animal faeces, plants, food, bee products, drugs, air, and the aquatic environments . Evidentic soils , mango oic soils  and animic soils ,22. Theyic soils  and applic soils ), roots ic soils  and peanic soils ) and leaic soils  and campic soils ).Moreover, traditional fermented foods including bibimbap , douchi Ba are cultivated routinely in Luria–Bertani (LB) medium at 30–37 °C aerobically [Ba such as B. nakamurai grew well on nutrient agar (NA), trypticase soy agar (TSA), Reasoner’s 2A agar (R2A) and tryptone glucose yeast extract agar (TGY) at 30 °C for two days [B. velezensis and B. siamensis were also reported to grow well on TSA at 37 °C and 32 °C, respectively [Generally, the members of OGobically ,16,17. Stwo days . MoreoveBa [B. velezensis 157, B. velezensis DKU_NT_04, and B. velezensis NJAU-Z9 , and B. velezensis LB002 (which contained three plasmids). Interestingly, some studies have focused on the functionality of the genes carried by the plasmid. For instance, the B. velezensis S499 plasmid, pS499, was reported as containing a rap-phr cassette. This cassette encoded for the regulator aspartate phosphatase (rap) and the Rap regulatory peptide (phr) with a role in governing protease secretion, growth and motility, biofilm formation and production of surfactin [B. amyloliquefaciens LL3 plasmid, pMC1, has a 6.8 kbp plasmid that includes a rap which is not homologous to the pS499 [rap and the origin of replication of the pMC1 plasmid were cloned into the pKSV7, vector which brought about the production of plasmid-cured strains. The plasmid-cured strains have increases in glutamate-independent poly-γ-glutamic acid production by 6% as compared to the B. amyloliquefaciens LL3 [In 2019, 254 bacterial strain genomes which had been deposited in the NCBI database were reported as belonging to the OGBa . Some ofurfactin . Meanwhihe pS499 . The hypiens LL3 .Ba. The genomic and metabolic features of the members of the group were similar; however, species-specific features including secondary metabolite biosynthesis-related and energy metabolism-related genes were also identified [B. velezensis, whereas energy metabolism-related genes are enriched in B. amyloliquefaciens. In the core-genome, B. velezensis harbors more genes involved in the biosynthesis of antimicrobial compounds as well as genes involved in D-galacturonate and D-fructuronate metabolisms compared to B. amyloliquefaciens and B. siamensis. Moreover, a xanthine oxidase gene cluster that is involved in metabolizing xanthine and uric acid to glycine and oxalureate was found in the core-genome of all the members of the group. Pan-genome analysis revealed the abilities of members of OGBa to metabolize diverse carbon sources aerobically or anaerobically. Their abilities to produce various metabolites such as lactate, ethanol, xylitol, diacetyl, acetoin, and 2,3-butanediol were also identified [Ba to produce antimicrobial compounds [Genome analysis allowed for further biological studies on the members of OGentified ,44. Secoentified . Genome ompounds .Ba as biocontrol agents were determined based on their abilities to improve plant growth and health [B. velezensis S141 with Bradyrhizobium diazoefficiens USDA110 into soybean resulted in enhanced nodulation and nitrogen fixation efficiency by producing larger nodules [Ba were able to solubilize phosphate, and produce IAA, ACC deaminase and siderophores [In the agricultural sector, the biocontrol strategy has received great attention because it provides safe, environmentally friendly, long-lasting, and inexpensive alternatives . The chad health . These a nodules . In anotrophores ,50,51.Ba produced antimicrobial compounds such as hydrogen cyanide (HCN) and cyclic lipopeptides such as surfactin used to inhibit the growth of pathogenic microbes [Meanwhile, the indirect mechanism is mainly due to their biocontrol activities attributed to the production of antimicrobial compounds in response to biotic stress . The memmicrobes ,54. The microbes ,55.Ba were proven to provide advantages to the agricultural sector by contributing to plant pathogen disease suppression. In plant disease management, the members of OGBa acted as plant growth-promoting bacteria (PGPB) that aid in the development of plants and reduce the proliferation of plant pathogens  [The members of OGathogens . The secnfection . Similarus (TMV) .Ba are used in bioformulations. For instance, the bacterial strain B. velezensis FZB42 had been established as a model strain for plant growth promotion and as a biocontrol agent [B. velezensis FZB42 showed great inhibitory effects against Fusarium solani infections under in vitro conditions. Increments in germination percentage and germination rate as compared with the control were also reported [Bacterial species from the OGol agent . In 2019reported .Ba available on the market  which contains B. velezensis QST 713 (previously B. subtilis QST 713) and Double Nickel 55TM  which contains B. velezensis D747 (previously B. amyloliquefaciens D747) [® together with Fracture fungicide , which contains BLAD polypeptide, had shown notable success in controlling Botrytis blossom blight disease infection in blueberries [TM was found to be effective in controlling white mold in snap beans caused by Sclerotinia sclerotiorum. Double Nickel 55TM, a biofungicide, was approved for organic vegetable production by the National Organic Program and Organic Materials Review Institute [To date, there are a few bioformulations containing bacterial species from the OGe market , such aseberries . Applicanstitute .Ba have also been applied as biocontrol agents against parasitic nematodes and protist pathogens. In 2008, B. velezensis FZB42 was reported to reduce nematode eggs in roots, juvenile worms in soil and plant galls on tomato [B. velezensis FZB42 encoded a diverse spectrum of different antimicrobial compounds able to suppress harmful nematodes living within the plant rhizosphere [B. velezensis HB-26 (previously B. amyloliquefaciens HB-26) showed promising capability for controlling Plasmodiophora brassicae, a root-infecting protist that causes clubroot disease in brassica species. Many antimicrobial compounds showing specific activities against P. brassicae were found in the genome of B. velezensis HB-26 [Ba to inhibit nematodes and protists beyond in silico genomic studies. Understanding such attributes will help to shed light on the functionalities as well as the biological roles of antimicrobial compounds from OGBa not only for improved plant growth but as biocontrol agents to minimize the proliferation of plant pathogens including viruses, bacteria, fungus, nematodes, and protists.Apart from the aforementioned uses, the members of OGn tomato . Genomiczosphere . In contis HB-26 . Overallα-amylase, protease, and lipase have been used in various biotechnological applications including textile applications, feed industry, food industry, and organic synthesis [α-amylase and protease originating from B. subtilis are Generally Recognized as Safe (GRAS) for use as direct food ingredients [B. subtilis species complex, OGBa bacteria are a potent bacterial group due to their abilities to produce various types of enzymes including α-amylase, protease, lipase, cellulase, xylanase, pectinase, aminotransferase, barnase, peroxidase, and laccase  and fungi  pathogens [, in silico genomic study of B. siamensis JFL15 had gene clusters involved in the biosynthesis of antimicrobial compounds. The LC–MS/MS analysis confirmed the presence of iturin A and bacillomycin F. Both compounds showed strong antifungal activities against Magnapothe grisea, R. solani and Colletotrichum gloeosporioides, as analyzed under in vitro conditions [B. velezensis OSY-S3 that showed inhibition activities against Listeria innocua, Escherichia coli, Penicillium sp., Cladosporium sp., and Staphylococcus aureus [Non-ribosomal peptides produced by bacteria and fungi contain two or more moieties derived from amino acids . The modathogens . In anotnditions . Moreoves aureus .B. velezensis OSY-GA1 [B. velezensis YJ11-1-4 isolated from doenjang exhibited good antimicrobial activities against bacterial  and fungal (Aspergillus flavus subsp. flavus) foodborne pathogens. Genomic analysis reveals the presence of antibiotic biosynthesis operons including bacillaene, macrolactin and difficidin in the genome of B. velezensis OSY-GA1 [B. velezensis AH159-1. These compounds inhibited S. aureus peptide deformylase and also showed antibacterial activities against E. coli and S. aureus [Polyketides are biopolymers of acetate and other short carboxylates that are biosynthesized by polyketide synthases, a natural metabolite produced by microorganisms and plants which possess various antifungal and antibacterial activities ,112. Sin OSY-GA1 . Moreove OSY-GA1 . Additio. aureus .Ba, some B. velezensis strains are reported to display probiotic potential and have been applied as probiotics for animals [B. velezensis H57 (previously B. amyloliquefaciens H57) isolated from lucerne was first investigated in the research to prevent fungal spoilage of hay [B. velezensis H57 was commercialized as a spoilage control agent under the product name HayRite™ . Interestingly, sheep and cattle fed on HayRite™ showed improvements in digestibility and nitrogen retention leading to increased weight gain [B. velezensis H57 to synthesize antimicrobial compounds including surfactin (srfABCD), iturin (ituABCD), bacillomycin D (bmyABC), fengycin (fenABCDE), macrolactin (mlnABCDEFGHI), difficidin (dfnABCDEFGHIJ) and bacillaene (baeEDLMNJRS) were suggested to facilitate the probiotic effects of B. velezensis H57 [B. velezensis FTC01 manifested itself as a probiotic [B. velezensis FTC01. Additionally, peptidylprolyl isomerase (prsA) gene  was also found. Moreover, in silico genome analysis of B. velezensis FTC01 proved the presence of gene clusters involved in the synthesis of antimicrobial peptides. Similarly, gene clusters involved in the synthesis of antimicrobial peptides were also found in the genome of B. velezensis JT3-1, a probiotic strain isolated from faeces of the domestic yak [B. velezensis JT3-1 was confirmed using an antimicrobial assay. Strain JT3-1 manifested strong antagonistic activities against various intestinal pathogenic flora including L. monocytogenes, S. aureus, E. coli, Salmonella typhimurium, Mannheimia haemolytica, Staphylococcus hominis, Clostridium perfringens and Mycoplasma bovis.Probiotics are live microbial feed supplements that benefit the host animal by improving the microbial balance. Probiotics have become increasingly popular due to continuously expanding scientific evidence pointing to their beneficial effects on both humans and animals . Within  animals . For inse of hay . Becauseght gain . Genomicnsis H57 . In anotrobiotic . Genes cstic yak . The antB. velezensis B-1895 (previously B. amyloliquefaciens B-1895) has been commercially used as a probiotic in the fish industry, particularly for Alburnus leobergi [Streptococcus intermedius and Porphyromonas gingivalis). Moreover, the endospores of B. velezensis B-1895 were found tolerant to 0.3% (w/v) bile salts and survived incubation for 4 h in MRS broth at pH 2.0–3.0. Overall, the results suggested the potential of B. velezensis B-1895 as a fish probiotic [B. velezensis JW also manifested itself as a fish probiotic [Aeromonas hydrophila, Aeromonas salmonicida, Lactococcus garvieae, Streptococcus agalactiae and Vibrio parahemolyticus). Dietary administration of B. velezensis JW induced an immune response in Carassius auratus. The immune-related genes in C. auratus such as interferon gamma gene (IFN- γ), tumor necrosis factor-α (TNF-α), interleukin-1 (IL-1), interleukin-4 (IL-4) and interleukin-10 (IL-10) were found to be upregulated by B. velezensis JW-supplemented diets. It is noteworthy that C. auratus fed with B. velezensis JW-supplemented diets showed improvements in survival rate after A. hydrophila infection. This was supported genomically by the presence of antimicrobial gene clusters in the genome of B. velezensis JW [B. velezensis V4 on the growth performance of Oncorhynchus mykiss had also been investigated [B. velezensis V4 with anti-A. salmonicida was shown to contain antimicrobial compounds including iturin, macrolactin and difficidin. The mortality rate of O. mykiss was reduced by 27% and the weight gain ratio was increased by 71% through the 1% (v/w) addition of B. velezensis V4. Overall, the findings demonstrated that B. velezensis V4 was an effective probiotic in O. mykiss.leobergi ,121. Itsrobiotic . In anotrobiotic . Strain ensis JW . Moreovestigated . Cell-frB. amyloliquefaciens as a probiotic in aquaculture is not as common compared to its agricultural applications  were applied to the culture water for 42 days. At the end of the trial, there was no significant improvement in the water quality. However, it showed notable changes in the immune system of the shrimp. As compared to the control treatment, there was an increase in the total protein concentration and granular hemocytes, and a decrease in the cell number with apoptosis in the hemolymph in all treatments. Therefore, other than being mixed with feed, B. amyloliquefaciens in the commercial probiotic Ecobiol® Soluble Plus could also be applied directly to the culture system; this research proved it provided better resistance to shrimps against the outbreak of pathogens in shrimp biofloc systems [The commercialization of ications . Ecobiol systems .Ba as potential probiotics for commercialization purposes in the aquaculture industry. Most of the studies have emphasized probiotic feed formulations, feeding trials on a small scale before moving to field trials. For instance, dietary inclusion of B. amyloliquefaciens at 106 CFU/g fed to zebra fish improved the expression levels of metabolism-related genes, enzyme activities and oxidative stress-related genes in the fish liver as well as enhanced their immune resistance against pathogenic A. hydrophila and S. agalactiae. In addition, the strain of B. amyloliquefaciens used in this study was able to express recombinant xylanase, an important enzyme that aided in better feed digestibility and efficiency [B. amyloliquefaciens (1 × 109 CFU/g), together with Spirulina platensis in formulated diet for tilapia, improved growth performance and feed utilization after a 60 day feeding trial. The mRNA level of the TNF-α gene and the transcription of SOD were considerably higher in tilapia fed with dietary B. amyloliquefaciens and S. platensis compared to the control group [B. amyloliquefaciens at a concentration of 106 CFU/mL provided significant protection to juvenile blue swimming crabs, Portunus pelagicus, when challenged with Vibrio harveyi in in vivo trials [Ba for aquaculture use.There is much ongoing research on the development and formulations of bacterial strains belonging to the OGficiency . In anotol group . Moreoveo trials . NeverthIn vivo and field trials are critical in probiotic development. Occasionally, there were negative outcomes in in vivo studies which were carried out based upon the positive results acquired from the preliminary in vitro assays, which indicated the possibility of negative correlations between trials in vitro and in vivo. Hence, it is crucial to understand and to optimize various conditions in in vivo studies or field trials including the probiotic formulation which may affect the survival, colonization, proliferation, and interaction of the probiotic with the host in a certain environment .Ba is as described above. Interestingly, in 2019, B. amyloliquefaciens YP6 was reported to exhibit both plant growth-promoting activity and broad-spectrum organophosphorus pesticide (OP) removal [B. amyloliquefaciens YP6 found it to contain a variety of promising genes, including phosphorus solubilizing and OP-degrading related genes , indole-3-acetic acid synthesis related genes , and siderophores synthesis related genes . The results hinted at the potential application of B. amyloliquefaciens YP6 in agricultural and environmental remediations. Overall, much more focus is still needed to understand the OP-degrading related genes beyond in silico genome analysis. Therefore, it is necessary to conduct further studies to determine the in vitro functional genomics and the OP-degrading enzyme activities of the members of OGBa. Understanding such attributes will help to shed light on the applicability of the OGBa in OPs degradation and in the bioremediation processes as a whole.The use of microorganisms as bioremediation agents has become a burgeoning trend . To date removal . In siliBa is remarkable. The bacteria are important not only industrially, but also environmentally. A plethora of studies have addressed the abilities of the members of OGBa as plant growth-promoters, biocontrol agents, probiotics, bioremediation agents as well as producers of commercial enzymes and antibiotics. Moreover, the use of the bacteria in optimized bioformulations as well as the demonstration of the great success of the commercialized products give us hope towards more sustainable agricultural and aquacultural industries. Owing to the listed biotechnological applications and potentials, more research should be done focusing on the integration of system biology data derived from genomics, phenomics, proteomics, metabolomics and fluxomic analyses in order to expand our basic understanding on the versatility of the members of OGBa. Enabling the prediction of cellular functions and metabolites produced by the members of this operational group could provide fundamental knowledge towards the enhancement of the applications of their potentials in biotechnology and bioprocessing for the benefit of all.In conclusion, the progress of the research on the biotechnological applications of bacterial species that belong to OG


As the use of single-cell technologies has grown, so has the need for tools to explore these large, complicated datasets. The UCSC Cell Browser is a tool that allows scientists to visualize gene expression and metadata annotation distribution throughout a single-cell dataset or multiple datasets.https://cells.ucsc.edu.We provide the UCSC Cell Browser as a free website where scientists can explore a growing collection of single-cell datasets and a freely available python package for scientists to create stable, self-contained visualizations for their own single-cell datasets. Learn more at Bioinformatics online. Single-cell RNA-seq assays allow for the exploration of gene expression at unprecedented detail, for surveying cellular diversity in organs . This grx, y coordinates by dimensionality reduction algorithms like tSNE  normalization, (ii) dimensionality reduction, (iii) clustering, (iv) marker gene identification and cluster labeling and (v) visualization and sharing. Expression matrices are often normalized, trimmed or batch-corrected. They are mapped from many dimensions to just two as ike tSNE  and UMAPx, y coordinates, scientists can switch between different layouts. Scientists can pan and zoom along the plane, similar to navigating within geographic map viewers. Cells can be colored based on provided annotations  or by gene expression using several built-in color palettes. The display can be split into two panes offering a side-by-side view allowing comparison between metadata attributes or genes .The UCSC Cell Browser can be used to display any high-dimensional data, such as single-cell ATAC-seq data  or bulk Data from single-cell experiments can be visualized using a number of tools in addition to the visualization options available through the analysis packages Scanpy (http://loompy.org/) and other files. Both Seurat and Scanpy provide functions to export data and build a Cell Browser instance. Installation instructions and other documentation are available at https://cellbrowser.readthedocs.io/.A cell browser can be built from plain text files , but we provide utilities to import data from Seurat, Scanpy, Cellranger  to M.H., W.J.K., M.L.S.]; California Institute for Regenerative Medicine ; University of California Office of the President Emergency COVID-19 Research Seed Funding [R00RG2456 to M.H.]; Chan Zuckerberg Initiative Foundation ; Simons Foundation [SFARI 491371 to T.J.N.]; Brain and Behavior Research Foundation [NARSAD Young Investigator Grant to T.J.N]; Gifts from Schmidt Futures and the William K. Bowes Jr Foundation to T.J.N.Conflict of Interest: none declared.No new data were generated or analysed in support of this research.btab503_Supplementary_DataClick here for additional data file.


Helicobacter pylori (H. pylori) and nutritional status in children are conflicting. We designed a large-sampled prospective community-based study to examine the differences in average body indices among Polish teenagers depending on their H. pylori status.  Data on an association between 13C urea breath test for current H. pylori infection was performed and data on anthropometric measurements and sociodemographic characteristics were collected. Z scores of height for age (HAZ), weight for age (WAZ), and BMI for age (BMIZ) were calculated.  From September 2008 to June 2015, 3067 second junior high school students aged between 13 and 17 years (mean age: 14.5) from 11 randomly selected schools from Grudziadz, Poland, were recruited. For the cohort, H. pylori colonisation rate was 23.6% with no gender difference. Compared to noninfected, H. pylori infected had significantly lower mean WAZ (0.0085) and BMIZ scores (p = 0.0246). Univariate linear regression models showed that living in the old town district and consumption of tap water were negative predictors of HAZ, living in the old town district, using collective catering facilities, and H. pylori infection were negative predictors of WAZ, and using collective catering facilities and H. pylori infection were negative predictors of BMIZ. In the multiple regression analyses, living in the old town district (p = 0.0039), using collective catering facilities (p < 0.0001), and H. pylori infection (p = 0.0269) were confirmed to be independently associated with lower WAZ, whereas using collective catering facilities (p < 0.0001) and H. pylori infection (p = 0.0265) were confirmed to be independently associated also with lower BMIZ.  The H. pylori infection on nutritional status in Polish teenagers. Our finding confirms the evidence on independent negative influence of  Helicobacter pylori (H. pylori) is the most common chronic bacterial infection as it affects approximately one-third of children worldwide. Most infections are commonly acquired in childhood. Furthermore, in most, if not all, infected individuals, H. pylori causes chronic gastritis that in children seems to be mainly antrum-predominant. Unlike adults, most infected children remain asymptomatic. Only a minority develop duodenal and gastric ulcer [H. pylori infection plays a role in nutritional status of affected children in numerous research studies has been examined, but results were inconsistent. Given these controversial data, we designed a large-sampled prospective community-based study to examine the differences in average body indices among Polish teenagers depending on their H. pylori status.ic ulcer . The queThe study was conducted in Grudziadz, an industrial city in north-central part of Poland with a population about 100,000 residents. A selection of the city for the research was associated with the source of financial coverage (the City Council of Grudziadz) and the city-level socioeconomic variation. The city is divided into 7 new districts and 6 old districts. Living in the old district reflects a combination of lower household income, higher unemployment rate, and lower educational level of parents, as well as poorer housing condition and lower level of hygiene and sanitation when compared to the new city district.H. pylori status. Moreover, data on sociodemographic and hygiene practices with use of a self-completed questionnaire were collected. The questionnaire was divided into three main sections as follows: (a) sociodemographic variables , (b) clinical symptoms, and (c) hygiene practices . Of these 3241 subjects, 1892 (58.4%) were girls and 1349 (41.6%) were boys. Of these 3241 subjects, 13C urea breath test results were obtained from 3067 participants, who were eligible for the final analysis.In this prospective cross-sectional study conducted between September 1, 2008, and June 30, 2015, 3375 second junior high school students from eleven randomly selected secondary schools from Grudziadz, Poland, were recruited. A total of 3241 participants aged between 13 and 17 years (mean age: 14.5), who signed an informed consent, were investigated for body height, weight, and In the group, 67 of 3067 (2.2%) were older than 16 years and 1382 (45.1%) lived in old city districts. Collective catering facilities were used by 734 (27.5%) questionnaire responders. Consumption of raw vegetables, raw meat, and tap water was self-reported by 1735 (56.6%), 365 (11.9%), and 835 (27.2%) participants, respectively. Hand washing after coming back home, before meal, after using the toilet, and after contact with animals was not practiced by 842 (27.5%), 590 (19.2%), 170 (5.5%), and 1053 (34.4%) participants, respectively. Owing a cat, a dog, or both was reported by 336 (11%), 1070 (34.9%), and 259 (8.4%) questionnaire responders, respectively.13C urea breath test (UBT) for current H. pylori infection was performed using HeliFAN plus analyzer . The protocol of the test was consistent with the manufacturer's specifications. A validation of the test was conducted once a year with the use of samples of known gas concentration. The accepted variation in the test results between tested and standardized samples was below 10% as recommended. Breath samples before and 30 minutes after the intake of 75 mg of 13C-labeled urea were collected. The final result was expressed as the difference between the two scores, delta over baseline (DOB). The cut-off point was 4.0. A result equal to or higher than 4.0 DOB was considered positive for H. pylori infection. All participants tested positive for UBT were offered upper gastrointestinal endoscopy with multiple gastric biopsies. All children with histologically confirmed H. pylori-related lesions were scheduled for the empiric eradication therapy.For the cohort, 2). Height-for-age, weight-for-age, and BMI-for-age were expressed as Z scores. Z scores of height for age (HAZ), weight for age (WAZ), and BMI for age (BMIZ) were calculated using the formula: Z score = /standard deviation value of reference population. The calculations were based on the 2010 Polish growth reference charts.Anthropometric measurements were performed by trained registered nurses. Body weight was measured using a digital scale , and body height was measured by a stadiometer. The calculation of BMI (body mass index) was performed on the basis of height and weight measurements  were excluded. Nonlinear relations between outcome and continuous predictors were considered by identifying, at each iterative step of the stepwise process, the best fitting fractional polynomial terms. This model development process led to a final model for the prediction of HAZ, WAZ, and BMIZ based on the selected predictors along with their corresponding estimated β coefficients and the associated intercept term. Statistical analyses were performed using TIBCO Statistica® 13.3.0 and R, version 3.3.2 . All tests were considered statistically significant at p value less than 0.05.The mean and the standard deviation of HAZ, WAZ, and BMIZ scores for The study was approved by Bioethics Committee of Nicolaus Copernicus University in Toruń (KB 490/2008). The informed consent was obtained from each study participant and signed by a parent/caregiver for children below 16 years and by both a parent and a child aged 16 years and older.13C UBT results (Hp (+)) and 2344 subjects (76.4%) had a negative 13C UBT results (Hp (-)) with no statistically significant difference in 13C-UBT positivity between males and females.Seven hundred and twenty-three participants (23.6%) had a positive M) and standard deviation (SD) of HAZ, WAZ, and BMIZ scores in children tested positive and negative for H. pylori are presented in H. pylori infected had lower mean HAZ, WAZ, and BMIZ scores with WAZ and BMIZ scores being statistically significant.The mean .Human growth is complex and depends on factors that may also be associated with affected –5 or onlaffected , 7 or onaffected , 9 beingn growth  or even n growth . The incH. pylori infection than their noninfected counterparts. Furthermore, this negative association between H. pylori infection and both WAZ and BMIZ scores was independent of sociodemographic variables and different hygienic practices. These results support a negative relation between weight and BMI and H. pylori infection and suggest that except for improvements in socioeconomic and hygienic conditions, in children with growth disturbances, H. pylori testing should be offered. This is in agreement with a Peruvian [H. pylori infection on weight among children from low socioeconomic background. According to the systematic review by Lender et al. [H. pylori infection and overweight and obesity in developed countries, which also confirms the negative effect of H. pylori on body weight. In contrast with these findings, in the largest pediatric study on the subject to date, differences with regard to both body weight and body mass index between infected and noninfected children were not significant. However, in this study, boys with H. pylori infection had significantly lower weight than those noninfected [In this large-sampled prospective community-based study, we showed that WAZ and BMIZ scores were significantly lower in 13-17-year-old Polish teenagers tested positive for Peruvian  and Coloinfected .H. pylori infection had lower HAZ score, but the difference was not statistically significant. This finding may suggest that weight gain is influenced by H. pylori infection in an earlier time period, compared to height. It can be hypothesized that with exposure time elongation, the effect of H. pylori infection on height would become more prominent. This hypothesis is consistent with a prior cohort study of Kocaoglu et al. [H. pylori infection may negatively influence growth [H. pylori infection [In this study, children tested positive for u et al.  who conce growth –6 and cae growth . In contnfection , 16.H. pylori might negatively affect physical growth, likely after the establishment of persistent infection. It is no clear by which mechanism H. pylori may impair physical growth; nonetheless, there are multiple plausible explanations. It has been shown that H. pylori-related inflammation can affect the levels of neuroendocrine gastric hormones such as ghrelin and leptin. They play a crucial role in regulation of eating behaviour and energy expenditure. Ghrelin is a peptide hormone well known for its growth hormone-releasing and appetite-stimulating effect. In a systematic review published in 2011, the significantly lower circulating ghrelin concentration in H. pylori-positive subjects compared to those not infected has been conclusively confirmed [H. pylori infection on plasma ghrelin level has been demonstrated mainly in adults with the fundus-predominant gastritis. In children, in whom H. pylori-related gastritis is mainly of antrum-predominant type, ghrelin secretion impairment is not expected [Our study and other well-designed studies support the notion that onfirmed . Howeverexpected . Some stexpected .H. pylori may induce a decrease in iron stores. Children with iron deficiency may have reduced appetite that may affect their nutritional intake and their overall physical status. In addition, some studies have shown an association between chronic H. pylori infection and disturbed absorption of many nutrients and vitamins, which in turn impairs childhood growth [H. pylori-infected individuals compared to uninfected group. The association was stronger in children, for whom a 2-fold higher prevalence of iron deficiency anemia was observed [H. pylori positivity and the dietary intake [Another explanation is that d growth . The curobserved . Howevery intake .H. pylori infection and the largest conducted in Poland. Furthermore, for the diagnosis of H. pylori infection, we used the 13C urea breath test that remains a most reliable noninvasive test with the highest diagnostic odds ratio when compared to 14C urea breath test, serology, and stool antigen [This study has both limitations and strengths. The strengths of our study include its prospective design and a very large sample of the population. It is one of the largest studies concerning height and weight in relation to  antigen .H. pylori acquisition.The main limitation of the study is the lack of detailed socioeconomic data. We analyzed the patients' place of residence as a surrogate marker for socioeconomic status. It can be misleading, as living in old city district might not be inseparably associated with poor hygienic practices and other factors playing a role in H. pylori infection on the nutritional indices in individuals from different socioeconomic conditions.Secondly, the study was conducted among teenagers from small region in Poland; therefore, conclusions drawn based on its result may not be universally applicable. However, we recruited a cohort of individuals from fairly heterogeneous socioeconomic background, which enables us to assess the effect of H. pylori infection.Finally, the present study provides no insight into the pathogenesis of the inverse correlation between anthropometric indicators of nutritional status and Helicobacter pylori detection, our finding adds to the evidence on independent negative influence of H. pylori infection on nutritional status in Polish teenagers.In conclusion, by employing the most accurate noninvasive diagnostic method for


Listeria monocytogenes, Staphylococcus aureus, Salmonella enterica and Escherichia coli. In addition, biofilm formation by Pseudomonas aeruginosa is discussed because it aids the persistence of many foodborne pathogens forming polymicrobial biofilms on food contact surfaces, thus significantly elevating food safety and public health concerns. Furthermore, in-depth analyses of several bacterial molecules with dual functions in biofilm formation and pathogenicity are highlighted.Biofilm formation is an integral part of the microbial life cycle in nature. In food processing environments, bacterial transmissions occur primarily through raw or undercooked foods and by cross-contamination during unsanitary food preparation practices. Foodborne pathogens form biofilms as a survival strategy in various unfavorable environments, which also become a frequent source of recurrent contamination and outbreaks of foodborne illness. Instead of focusing on bacterial biofilm formation and their pathogenicity individually, this review discusses on a molecular level how these two physiological processes are connected in several common foodborne pathogens such as  Listeria monocytogenes found on cantaloupe skin caused a multistate outbreak in 2011 [Most microbes found in nature exist in biofilms, a well-structured, dynamic, diverse, synergistic and protective microbial community ,2. Biofi in 2011 ,9.Staphylococcus saprophyticus were more resistant to several antibiotics in their biofilm status compared to their planktonic form. Likewise, biofilms of a commonly used model food spoilage bacterium Lactobacillus plantarum were more resistant to several biocides, including organic acids, ethanol, and sodium hypochlorite, than in its planktonic state [Microbial attachment and biofilm formation on solid surfaces provide the advantages of living in a protective scaffold against desiccation, antibiotics, or biocides (sanitizers), ultraviolet radiation, metallic cations, and physical impact from washing and cleaning. For instance, Martins et al.  recentlyic state . Bacteriic state ,13. In aic state .Listeria monocytogenes, Micrococcus spp., Staphylococcus spp., Clostridium spp., Bacillus spp., Lactobacillus spp., Brochothrix thermosphacta, Salmonella enterica, Escherichia coli, Serratia spp., Campylobacter spp. and Pseudomonas spp. [Common biofilm-forming microorganisms vary in different food processing environments, which may include nas spp. ,16. The nas spp. . FurtherPathogen transmission through food results in approximately 2 billion cases and over one million annual deaths globally . In the L. monocytogenes in nature gives it numerous routes to be introduced in a food processing environment with various fresh produce or raw materials [L. monocytogenes finds a niche in a food processing facility, it can attach to several abiotic surfaces, such as stainless steel, PVC, and polystyrene, and start to form biofilms, which can be resistant to sanitation and may lead to recurrent food contamination [L. monocytogenes strains can persist for a few months and up to 12 years [L. monocytogenes isolates in the food processing environment may also be due to the same strains that were consistently introduced by raw material, or because of ineffective sanitation practices [Pathogen transmissions occur primarily through raw uncooked or undercooked foods and by cross-contamination during unsanitary food preparation practices. Pathogens find a harborage site or niche in food production facilities or product surfaces by forming biofilms . These naterials ,24. Oncemination ,26. Repe12 years . The perractices ,29.Listeria monocytogenes, Staphylococcus aureus, Escherichia coli, and Salmonella enterica  attachment, (ii) microcolony formation, (iii) maturation with cellular differentiation, and (iv) detachment or dispersion . In biofL. monocytogenes infection process and biofilm formation. L. monocytogenes is a rod-shaped, Gram-positive, facultative intracellular foodborne pathogen. The gastrointestinal phase of infection is central to it causing localized inflammatory disease (gastroenteritis) or systemic invasive disease [L. monocytogenes uses three major invasive pathways: (a) Listeria adhesion protein (LAP), (b) Internalin A (InlA), and (c) M-cell-mediated translocation [Pathogen interaction with host cells is a prerequisite for initiating the infection process. Several adhesion proteins play a key role in the  disease ,46. As alocation   In the LAP-dependent pathway, LAP  interacts with the host cell receptor and heat shock protein 60 (Hsp60) ,49. It a barrier ,51.L. monocytogenes in the lumen when cell extrusion or mucus exocytosis creates a transient opening [L. monocytogenes into a vacuole followed by cytoplasmic inhabitation for cell-to-cell movement [(b) In the InlA-mediated pathway, surface-expressed InlA anchored to the peptidoglycan interacts with host receptor E-cadherin and promotes transcytosis . Althoug opening ,53. The movement . Though movement .L. monocytogenes are passively transported by microfold cells (or M cells) in the Peyer’s patch during sampling of luminal antigens [(c) In the M-cell pathway, antigens . Internaantigens  while itantigens ,58.L. monocytogenes systemically disseminate to extraintestinal organs, including the spleen, liver, mesenteric lymph node, gall bladder, brain, and placenta in pregnant women [L. monocytogenes expresses a pore-forming toxin, Listeriolysin O (LLO) (encoded by hly), and bacterial phospholipases, PlcA and PlcB to escape from the vacuole. LLO oligomerizes into “arc- or slit-shaped” assemblies in the membrane to lyse the vacuole and facilitate L. monocytogenes to escape by the inactivation of phagocyte NOX2 (NADPH oxidase), which produces reactive oxygen species as an antimicrobial strategy [L. monocytogenes in the endocytic vacuole to facilitate escape from the phagosome. The bacterium can also survive in the vacuole for an extended period, causing latent infection [After crossing the intestinal barrier, nt women . The spestrategy ,60. Besinfection .L. monocytogenes also suppresses the cellular proinflammatory response using internalin C (InlC) and moves from cell-to-cell by polymerizing host actin protein (ActA) [In addition, n (ActA) ,63. Thisn (ActA) . ActA ton (ActA) ,66 by stn (ActA) .Listeria pathogenicity island-1 (LIPI-1), including LLO, ActA, PlcA, PlcB, Mpl , and PrfA, and three additional chromosomal loci, which are the inlAB operon and inlC and hpt monocistron. In addition, the expression of as many as 145 other L. monocytogenes genes may indirectly be regulated by PrfA [L. monocytogenes to convert from an environmental saprotroph to a pathogen by taking on many environmental cues, such as temperature, oxidative stress, carbon sources, and low pH to modulate its virulence [prfA are defective in surface-adhered biofilm formation [The activation of these virulence genes is regulated under positive control by the master regulator-PrfA (positive regulatory factor). More specifically, the PrfA regulon directly regulates genes in the  by PrfA . The selirulence ,70. PrfAormation  possiblyL. monocytogenes  consisting of alternating phosphate and ribitol  or glycerol  groups, which are replaced with D-alanine and N-acetylglucosamine . LTA or ytogenes . Inhibititecture . Furtherytogenes .Lactobacillus casei expressing LAP from L. monocytogenes or L. innocua on the bacterial surface showed aggregation and increased biofilm formation on a microtiter plate [Lactobacillus casei , these cells showed upregulated expression of virulence genes in mouse intestinal lumen and eventually caused similar systemic dissemination as planktonic counterparts 48 h post-infection [L. monocytogenes murinized InlA (InlAm) strain (this strain has a high affinity for mouse E-cadherin) had a similar tissue distribution as the non-murinized (WT) strain in mice at 12–24 h post-infection confirming InlA involvement is limited to early stages of infection. These findings suggest that L. monocytogenes cells in biofilms have similar virulence potential as planktonic cells, which highlights the importance of the management of Listeria biofilms in food processing environments. In addition, biofilm-isolated cells showed significantly reduced virulence in in vitro cultured mammalian cell models compared to planktonic siblings and provided an inaccurate assessment of biofilm-forming pathogen infectivity. Therefore, animal models are necessary to accurately determine the virulence of biofilm-forming pathogens.surfaces ,16. Thern et al.  showed trch team  showed tnfection . Even anStaphylococcus aureus is a Gram-positive opportunistic pathogen and is commonly found in natural waters, soils, animal mucus membranes, and on human skin [S. aureus [S. aureus has a remarkably high salt tolerance, a feature that allows it to be a commensal skin bacterium on approximately 25% of the population, making humans a common vehicle for this pathogen’s transmission to foods.man skin ,80. The . aureus . S. aureS. aureus produce multiple heat-tolerant enterotoxins , toxic shock syndrome toxins, superantigens, exfoliative toxin, and many more virulence factors that can cause food poisoning with symptoms of nausea, vomiting, and stomach cramps [S. aureus contamination [S. aureus [S. aureus do not usually infect the gastrointestinal tract. However, antibiotic treatment for skin infection has become a great challenge because of biofilm formation and widespread antibiotic resistance.As a foodborne pathogen, h cramps . These smination . In the . aureus . Since sS. aureus contamination of food is a global challenge affecting multiple types of products. Between January 2000 and March 2012, approximately 14 S. aureus outbreaks were recorded in Australia, which affected 429 people including 25 hospitalizations and one death [S. aureus intoxication, a broad screening of dairy products showed that 102 out of 971 samples were positive for S. aureus [S. aureus cultures contained at least one enterotoxin coding gene.ne death . Approxi. aureus . ApproxiS. aureus form strong biofilms, which is critical for its persistence not only on food contact surfaces [S. aureus express a large number of surface molecules, which are collectively called microbial surface components recognizing adhesive matrix molecules (MSCRAMMs) that are involved in bacterial adhesion and biofilm formation [S. aureus surface protein G (SasG), S. aureus protein A (SpA), cell wall-anchored clumping factor (ClfA), polysaccharide intercellular adhesin (PIA) and teichoic acids (TA), and many have a redundant function [surfaces  but alsosurfaces ,87,88. Sormation . More spfunction   locus, including icaADB and icaC. Products from icaA and icaD are responsible for PIA synthesis using UDP-N-acetylglucosamine [S. aureus share 59–78% identity with those proteins from S. epidermidis [ica locus in S. aureus reduced its ability to form multilayer thick biofilms in microtiter plates, suggesting that PIA of S. aureus has the same function for cell-to-cell adhesion. The chemical nature of S. aureus PIA was identified to be poly-N-succinyl-β-1,6-glucosamine, which is homologous to PIA of S. epidermidis [S. epidermidis with a deficiency in the ica operon did not induce subcutaneous abscess and was possibly eradicated by the immune system. However, a study reported by Francois et al. [ica-negative S. aureus or S. epidermidis strains. This suggests that the function of PIA in a pathogen during infection may vary based on different infection methods or models used.PIA was initially discovered in ormation . Chemicacosamine . Ica prodermidis . Deletiodermidis . PIA expdermidis . It is ndermidis  reporteds et al.  contradiS. aureus can also form a biofilm independent of PIA under certain conditions. A two-component system, encoded by arlRS, is a repressor of S. aureus biofilm formation in Hussain–Hastings–White Modified Medium (HHWm) [arlRS mutant strain when compared to the parental strain [ica operon in the arlRS mutant strain did not affect S. aureus biofilm formation, suggesting that a PIA-independent pathway is responsible for biofilm formation under specific conditions.m (HHWm) . Crystall strain . Furtherspa gene is reported to have a similar function for cell-to-cell adhesin in biofilm development [S. aureus. Consequently, the spa mutant strain exhibited significantly weakened biofilm formation. Furthermore, in an in vitro aggregation experiment, S. aureus cells lacking protein A precipitated much slower than a strain expressing protein A, suggesting that protein A may be responsible for S. aureus cell-to-cell aggregation [spa mutant strain [Protein A coded by elopment . Mass spregation . Surprist strain .Protein A also serves as an important virulence factor and helps the bacterium to evade the immune system during infection. Protein A is covalently bound to peptidoglycan through an LPXTG motif. It also binds to the Fc region of antibodies, thus forcing antibody binding with the Fab portion facing outward from the cell, reducing the chance of opsonization and phagocytosis . Proteinbap gene that encodes Bap, anchored to the cell wall via LPXTG motif. Bap expression is regulated by a global regulator, SarA [S. aureus without Bap caused less persistent infection than wild-type S. aureus, suggesting that Bap is a virulence factor responsible for persistent infection [S. aureus biofilms [Another proteinaceous molecule that works as a cell-to-cell adhesin in biofilm formation is the biofilm-associated protein (Bap) . Cucarelor, SarA . An in vor, SarA . A mousenfection . Bap hasbiofilms .S. aureus biofilms are more susceptible to immune clearance by macrophages [The fibronectin-binding proteins A and B (FnBPA and FnBPB) are cell-to-surface-anchored (via LPXTG) proteins that bind to host cell fibrinogen and fibronectin and also form biofilm on medical implants and devices ,106. FnBrophages . The FnBrophages .S. aureus surface protein G (SasG) has been shown to participate in biofilm formation and adhesion to epithelial cells [S. aureus biofilm formation happens independently of PIA [al cells . Zinc acal cells ,110. Sasy of PIA .S. aureus [S. aureus mutant strain affected LTA biosynthesis, and biofilm formation [S. aureus [S. aureus adhesion to epithelial cells and reduced NF-κB activation and inflammatory response, suggesting that LTA is an important virulence factor promoting biofilm formation and pathogenesis in this bacterium [Teichoic acid is a highly charged cell wall polymer that also contributes to biofilm formation in . aureus ,77. Ablaormation . Likewisacterium .S. aureus. The role of eDNA in biofilm formation was first studied in P. aeruginosa [P. aeruginosa could effectively reduce biofilm formation without affecting bacterial growth. Interestingly, nicotine (a major component of tobacco smoke) promotes eDNA release and biofilm formation in S. aureus; however, it reduces its invasion to alveolar epithelial cells (A549) [Extracellular DNA (eDNA) is another important component that is involved in biofilm formation independent of PIA in ruginosa . The ress (A549) .cidA gene), a protein produced by S. aureus, is indirectly involved in biofilm formation [cidA gene affected eDNA release and biofilm formation [S. aureus biofilm formation. In addition, the high density of bacteria in biofilms also increases the rate of gene exchange [CidA , enteropathogenic E. coli (EPEC), enteroinvasive E. coli (EIEC), enteroaggregative E. coli (EAEC), diffusely adherent E. coli (DAEC), adherent invasive E. coli (AIEC), and Shiga toxin-producing E. coli (STEC) that includes enterohemorrhagic E. coli (EHEC) [E. coli are reported to form biofilms in the gastrointestinal tract and is orchestrated by regulatory networks, adhesion molecules, and extracellular matrix [i (EHEC) . Epitheli (EHEC) . Both par matrix .E. coli outbreaks are not only associated with raw meat or meat products [E. coli, which caused 3816 cases of infection including 800 cases of HUS and 53 fatalities [E. coli O157:H7 on lettuce [5 CFU/mL waterborne E. coli can survive on lettuce for 77 days possibly by forming biofilms on the produce surface. These findings further support the importance of biofilm control in the food processing environment to prevent pathogen contamination.products  but alsoproducts ,122. Fortalities . Patient lettuce , it was E. coli is a model bacterium that forms biofilm after well-programmed production of various extracellular molecules [As one of the pathogens causing the most gastroenteritis cases around the world, olecules . Curli aolecules .Curli fimbriae are proteinaceous extracellular fibers and are responsible for cell-to-cell and cell-to-surface attachment ,128. TheE. coli are composed of self-assembled CsgA nucleated by CsgB that also anchors curli to the bacterial surface [csgBAC and csgDEFG, which are responsible for secretion (CsgC and CsgG), transcription regulation (CsgD), and processing (CsgE and CsgF) [70 and σS in response to a variety of environmental signals. Under laboratory conditions, the expression of curli is optimal during the stationary phase in an environment with low nutrients and low medium osmolarity at a temperature ≤30 °C [Curli fimbriae of  surface . Includind CsgF) . The asse ≤30 °C . Curli fe ≤30 °C . The Csge ≤30 °C , which me ≤30 °C .E. coli with a mutation in csgA and csgD genes showed similar adhesion rates on both HeLa and HT-29 cell lines as the parental strain [E. coli adhesion to epithelial cells. On the other hand, when CsgD, the curli biosynthesis regulator, was overexpressed, the adhesion of E. coli was increased by approximately five-fold, suggesting that curli may still be a redundant adhesion factor. Recently, Elpers and Hensel [E. coli O157:H7, Sakai strain) and a subset of six gene clusters were responsible for adhesion to epithelial cell lines  and biofilm formation.In the intestinal tract, curli may not play an essential role in adhesion to the epithelial cells. l strain , suggestd Hensel  reportedE. coli in biofilms is cellulose [E. coli was closely connected with several genes that are homologous to bacterial cellulose synthesis (bcs) genes from Acetobacter xylinus [E. coli as cellulose. Interestingly, the regulator of the bcs operon is AgfD that is homologous to the curli regulator, CsgD [The major exopolysaccharide secreted by ellulose . Cellulo xylinus . Enzymator, CsgD . Celluloor, CsgD .E. coli strains form biofilm independent of curli or cellulose. Enteroaggregative E. coli (EAEC) can form thick aggregating biofilm on the intestinal mucosal surface and cause diarrhea [Some diarrhea . EAEC stdiarrhea . In the diarrhea . In addidiarrhea . Moreovediarrhea .Salmonella enterica is a Gram-negative rod-shaped bacterium and is closely related to E. coli. Based on genetic analysis, Salmonella diverged from E. coli approximately 100 million years ago [Salmonella enterica also cause outbreaks via contaminated fresh produce, poultry, eggs, nuts, spices, flours, milk, meat and drinking water [Salmonella enterica, referred to as non-typhoidal Salmonella (NTS), are responsible for 1.3 billion cases and 3 million deaths worldwide [Salmonella enterica serovars, Enteritidis, Typhimurium, Newport, Heidelberg, and Javiana represent the most commonly reported outbreak causing serovars [ears ago . Salmoneng water ,36,152. orldwide . In the orldwide . Among tserovars ,155.Salmonella occurred. Since the beginning of July 2020, the CDC has reported 1127 cases of Salmonella Newport infection including 167 hospitalizations from 48 states [S. Typhimurium can persist on parsley for at least four weeks when the concentration of the pathogen was above 108 CFU/mL. Biofilm formation and virulence properties of Salmonella isolates from ready to eat shrimps showed that S. Typhimurium was the strongest biofilm former among the isolates tested [While the whole world was focusing on addressing the COVID-19 pandemic in 2020, an outbreak of foodborne 8 states . This ou8 states  found ths tested .E. coli, secreted curli from Salmonella forms red dry and rough (rdar) colonies on Congo red-supplemented plates [Salmonella also form biofilms on food, the environment, and human and animal intestine [Salmonella has been proposed to exert anti-virulence properties [S. Typhimurium cells exhibit higher virulence gene expression than the other cells which aggregate, precipitate and show typical biofilm-related gene expression [csgD and cellulose-related genes [Similar to d plates . Salmonentestine . Severalntestine ,162,163 operties ,164. Plapression . Anothered genes .Salmonella biofilm formation. QS molecules such as autoinducers (AI)-1, AI-2 and AI-3 are responsible for S. Enteritidis growth, motility, adhesion and biofilm formation [Salmonella infection [Quorum sensing also plays an important role in ormation . The appnfection ,42.E. coli, curli fimbriae are also a major proteinaceous extracellular fiber expressed by Salmonella for cell-to-cell and cell-to-surface binding [Salmonella strain are bacterial aggregation and fibronectin-binding [csgBAC operons which are responsible for the synthesis of curli fimbriae [Salmonella Enteritidis can be classified into four types, SEF14, SEF17, SEF18, and SEF21, based on their molecular weights [sefA, agfA, sefD, and fimA, respectively. Austin et al. [S. Enteritidis on epithelial cells. Fuller et al. [Salmonella Typhimurium [Like  binding ,167. Two-binding . CsgD isfimbriae . The adh weights . Specifin et al.  showed tn et al.  found thr et al.  showed thimurium .S. Enteritidis to aid in biofilm formation [bapA resulted in weaker biofilm and the strain did not participate in biofilm formation when co-cultured with the parental strain. BapA expression is also coordinated with curli synthesis by csgD [Salmonella pathogenesis was shown using a bapA mutant strain that had reduced colonization and persistence in the intestine, invasion to liver and spleen, and lethality in mice [bcsA-encoded cellulose synthase is another critical component in Salmonella biofilms. Knockout of csgD and bcsA in a S. Enteritidis strain impaired biofilm formation and reduced lethality in specific pathogen-free chicken [csgD and ΔbcsA) strains [BapA is reported in ormation . Bap sec by csgD ,161. Bap in mice . In addi chicken . Vertica strains , suggestPseudomonas aeruginosa is a Gram-negative, non-spore-forming, and aerobic rod-shaped bacterium. It is an opportunistic pathogen affecting mostly immunocompromised individuals who are also suffering from other illnesses. Its metabolic activity is broad, and growth can occur in either nutrient-rich or nutrient-poor conditions. The ability to utilize a wide range of substrates as carbon and nitrogen sources supports the ability of P. aeruginosa to colonize in a variety of natural niches, such as water supply pipes and containers, and soil. Under laboratory conditions, the bacterium can grow well in a medium containing acetate and ammonium sulfate as carbon and nitrogen sources, respectively [ectively . The bacectively .P. aeruginosa produce a robust biofilm, which enhances its survival on different surfaces and protects the cells from other harsh conditions and treatments [P. aeruginosa is its resistance to multiple classes of antibiotics. In 2017, multidrug-resistant P. aeruginosa caused an estimated 32,600 infections among hospitalized patients and 2700 deaths in the US [P. aeruginosa makes it harder for antibiotics to access cells embedded in biofilms, which significantly reduces the effectiveness of antibiotics to treat infections.eatments . Anothern the US . In addiP. aeruginosa can be easily spread to humans, especially in health care settings. It can cause serious acute and chronic infections in immunocompromised people. For example, it is the most important bacterial pathogen causing progressive lung infection in cystic fibrosis patients leading to high fever, respiratory failure and death. P. aeruginosa can cause chronic urinary tract infections and ventilator-associated pneumonia in patients with permanent bladder catheters and intubation, respectively [P. aeruginosa is challenging because it can form infective biofilms after infection, which functions like a barrier protecting bacteria from complement-mediated immunity and phagocytosis, and significantly decreases the accessibility of antibiotics [P. aeruginosa biofilm formation is important for preventing and treating resistant infections.Due to its ubiquity, persistence, and drug resistance, ectively . Treatmeibiotics . TherefoP. aeruginosa rarely cause foodborne infection, it is still included in this review because it produces widespread biofilms in food processing plants [Pseudomonas has been shown to support the persistence and survival of L. monocytogenes on conveyor belts in a salmon processing plant, which was further verified under simulated conditions [Pseudomonas provide a shelter for foodborne pathogens for persistence and product contamination, and indirectly jeopardizes the safety of food products. P. aeruginosa is also one of the most well-studied model bacteria for bacterial biofilm research. Understanding Pseudomonas biofilm formation could help elucidate steps in the control and destruction of biofilms formed by other pathogens. Besides, the physiological regulation of biofilm formation and pathogenicity of P. aeruginosa is also similar to other bacteria. In P. aeruginosa biofilms, the highest cell density is arranged closest to the surface, and cells occupy only a minor fraction (approximately 2–28%) of biofilm volume, while EPS occupies the rest. Biofilm architecture is formed by EPS composed of exopolysaccharides, eDNA, and polypeptides [Pseudomonas biofilms have an open and porous structure that may be designed for the transportation of nutrients and waste. Affected by the different rheological characteristics of its environment, P. aeruginosa can form mushroom- or pillar-like matured biofilms [Though g plants  and contg plants . In polynditions . These fpeptides . Scanninbiofilms .P. aeruginosa provides swimming motility, chemotactic response, and a social movement referred to as swarming [P. aeruginosa produce weakened biofilm [P. aeruginosa were less invasive in the mouse burn wound model and colonized the murine intestine less [P. aeruginosa without fliC caused no mortality, whereas the mortality rate of the wild-type strain was approximately 30% [The polar flagellum on swarming . Both flswarming ,178. In  biofilm . The flaine less . In a netely 30% .P. aeruginosa belong to the type IV pilus family, which is assembled by the polymerization of monomeric major pilin and minor pilin proteins encoded by pilAEWV [Pseudomonas pili have a helix structure made up of a single subunit pilin of 18 kDa and each pilin monomer spontaneously assembles into filaments of 10–200 nm in length with a diameter of 1.2–5 nm [P. aeruginosa strains indicate there is great heterogeneity in pilin amino acid sequences [P. aeruginosa pilin is an intrachain disulfide loop of 12–17 amino acids located at the far end of the C-terminal, which is suggested to by the site for specific binding [Pseudomonas adhesion to and invasion of epithelial cells, as demonstrated in A549 cell (lung cells) [Various types of filamentous appendages on the bacterial surface have been studied and classified as pili . Pili ex pilAEWV . Besides pilAEWV . Pili-me pilAEWV ,183. Pse1.2–5 nm . The pilequences . However binding . Pili arg cells) . Glycospg cells) .P. aeruginosa are involved in the formation of cell monolayers and the typical mushroom-like structures. By expressing fluorescent proteins of different colors in the wild-type and pili-deficient P. aeruginosa strains, researchers visualized the distribution of these two types of bacteria and found that only the wild-type strain was located on the cap of the mushroom-like structure, suggesting that functional pili-mediated motility is necessary for forming this type of biofilm [P. aeruginosa: the Las, Rhl, PQS, and IQS systems [Both flagella and pili of  biofilm . Quorum  biofilm . QS is c biofilm . Current systems . Each sy systems . The Las systems .12-HSL), which is regulated by the AI synthases LasI, and sensed by the transcription factor LasR. The Rhl system uses AI synthase RhlI to produce an autoinducer, N-butanoyl-L-homoserine lactone (C4-HSL), which uses RhlR as its cognate receptor. Both Las and RhI are essential regulating systems for the maturation of Pseudomonas biofilm and their inhibition using synthetic molecules prevented pathogenesis in animal models [The Las system involves the production of an autoinducer N-(3-oxododecanoyl)-L-homoserine lactone  system is also involved in the release of eDNA and biosurfactants, which are essential for the development of mature biofilms during late exponential phase [pqsA mutant strain contains less eDNA than biofilms formed by their wild-type counterpart [pqsA expression by binding to the pqsA promoter [The al phase . The pqsnterpart . PqsR ispromoter . The binpromoter . Productpromoter .pqsA mutant strains was approximately 50% higher than infection with the parental strain [P. aeruginosa is also an opportunistic plant pathogen. A mutant strain with dysfunctional PQS production grew dramatically less than the wild-type strain in Arabidopsis, suggesting a critical role of PQS for the overall pathogenicity of P. aeruginosa [The PQS system is important in virulence factors generation during biofilm development. PQS mutants showed reduced biofilm development and less production of virulence factors, such as pyocyanin, elastase, lectin, and rhamnolipids. The correlation between the PQS system and infectivity has been tested using several in vivo models. In burn-wound mouse models, the survival rate of mice infected with l strain . P. aeruruginosa .ambBCDE is responsible for IQS synthesis. This disruption led to a decrease in PQS and BHL production, along with other virulence factors such as pyocyanin, rhamnolipids, and elastase [amb gene cluster is not responsible for aeruginaldehyde production, since its production is also found in Pseudomonas strains lacking this cluster. The important contribution of IQS to virulence of P. aeruginosa has been shown in animal models. For instance, mice infected by P. aeruginosa without ambB and lasI genes had a higher survival rate than either the wild-type or a lasI knockout mutant strain [The integrated quorum sensing (IQS) system is less studied compared to the other three systems. It produces 2-(2-hydroxyphenyl)-thiazole-4-carbaldehydte  as its cognate AI, while the receptor has not been found. A non-ribosomal peptide synthase gene cluster elastase . Howeverelastase  recentlyt strain . IQS prot strain . Under iP. aeruginosa and S. aureus, the presence of the latter organism can also increase exotoxin A expression [In addition, in mixed-culture biofilms of pression ,210, indL. monocytogenes, ActA rearranges actin in the host cell cytosol to propel cell-to-cell movement and also initiates biofilm formation by precipitating bacteria. Likewise, teichoic acids responsible for maintaining Gram-positive bacterial cell architecture also induce inflammatory response during infection and contribute to biofilm formation in both L. monocytogenes and S. aureus. Protein A of S. aureus not only helps the pathogen to evade the immune system but also facilitates cell-to-cell adhesion in biofilm development. Other proteins, including FnBP, SasG, and Bap, are also responsible for biofilm formation and pathogenesis in S. aureus. Curli is critical for biofilm formation and pathogenesis in E. coli. Similarly, curli and Bap are important in biofilm formation, intestinal colonization, and pathogenesis in gastroenteritis-causing non-typhoidal Salmonella. In Pseudomonas, PqsR plays a key role in the Pseudomonas quinolone signaling system and also regulates the production of virulence factors promoting bacterial biofilm formation and attachment to host epithelium. Other factors including flagella and type IV fimbriae are important in biofilm formation and colonization on epithelial cells. Many of the virulence factors that are involved in biofilm formation and host cell colonization have redundant functions, suggesting that even in the absence of one factor, bacteria can still form biofilms that are food safety and public health concerns.In summary, multifunctional molecules involved in both bacterial pathogenesis and biofilm formation demonstrate a close connection between the two aspects. In L. monocytogenes as a model foodborne pathogen to investigate the virulence of the bacteria in biofilms. Our data indicate that the virulence of biofilm-isolated L. monocytogenes was upregulated after 48 h bacterial adaption to the intestinal environment. These findings enhanced our understanding of bacterial pathogenesis of biofilm-isolated bacteria, and these data should be beneficial for the accurate evaluation of biofilm risks in food processing environments. Similarly, the assessment of the pathogenicity of other foodborne pathogens, such as E. coli and Salmonella, isolated from biofilms could also be further investigated using animal models. Using bacteria isolated from biofilms could also be a good model for studying bacteria switching from a saprophytic lifestyle to pathogenic status in animal hosts.Although the pathogenesis of multiple foodborne pathogens has been comprehensively studied, most of the results were generated using planktonic cultures under laboratory conditions. The actual risk of consuming pathogens from biofilms should be further characterized using animal models instead of only in vitro cultured mammalian cell models or virulence factor expression analyses. Recently, we used Although there are many studies of biofilm formation on plastic, stainless steel, or glass surfaces, more in-depth studies are needed of foodborne pathogen biofilms formed directly on food surfaces, for example, cantaloupe skin or eggshell. Bacteria isolated from these biofilms should represent a more realistic model to assess the risk of consuming foodborne pathogens found on food surfaces.


Primary central nervous system lymphoma (PCNSL) is characterized by a lack of specificity and poor prognosis. Further understanding of the tumor heterogeneity and molecular phenotype of PCNSL is of great significance for improving the diagnosis and treatment of this disease.To explore the distinct phenotypic states of PCNSL, transcriptome-wide single-cell RNA sequencing was performed on 34,851 PCNSL cells from patients. The cell types, heterogeneity, and gene subset enrichment of PCNSL were identified. A comparison of the PCNSL cells with 21,250 normal human fetal brain (nHFB) cells was further analyzed to reveal the differences between PCNSL and normal sample.Six cell populations were mainly identified in the PCNSL tissue, including four types of immune cells—B cell, T cell, macrophage and dendritic cell—and two types of stromal cells: oligodendrocyte and meningeal cell. There are significant cellular interactions between B cells and several other cells. Three subpopulations of B cells indicating diffident functions were identified, as well as a small number of plasma cells. Different subtypes of T cells and dendritic cells also showed significant heterogeneity. It should be noted that, compared with normal, the gene expression and immune function of macrophages in PCNSL were significantly downregulated, which may be another important feature of PCNSL in addition to B cell lesions and may be a potential target for PCNSL therapy. Primary central nervous system (CNS) lymphoma (PCNSL), defined as diffuse large B cell lymphoma (DLBCL) confined to the CNS, can occur in the setting of immunosuppression  or in immunocompetent individuals , 2. The The molecular subtype of PCNSL has been studied by different methodological approaches with conflicting conclusions. On the basis of several IHC studies, an ABC-like immunophenotype is typical , 9, but Standard chemotherapeutic regimens for systemic DLBCL show little efficacy in PCNSL, likely because of inefficient drug delivery across the blood–brain barrier . High-doThe rarity of the disease and the difficulty of obtaining intracranial specimens have hindered understanding of the pathophysiology of PCNSL. The observed overexpression of BCL6 and aberrant somatic hypermutation (aSHM) of many genes, together with expression of immunoglobulin M at the cell surface, have suggested that PCNSL cells may be arrested at the stage of terminal B-cell differentiation . The genGenome-wide gene expression in PCNSL compared with non-CNS DLBCL suggests that PCNSL has specific signatures to be distinguished from non-CNS DLBCL and greater molecular heterogeneity , 25. Of Some vague evidences suggest that CNS lymphoma and peripheral lymphoma are heterogeneous, which may be related to different immune environments and different origins of lymphoma cells. At present, there is no clinical significance to reveal the molecular characteristics of CNS lymphoma.Here, we performed an analysis of scRNA-seq data of CNS tissue sequenced by 10x Genomics to identify the cell types of CNS cells; we next performed differential expression (DE) analysis on two PCNSL-associated cells  and gene set enrichment analysis (GSEA) of their DE genes on the cell type-specific marker genes to examine the cell-specific functionalities in PCNSL development.Human PCNSL tumor samples were obtained surgically from clinical patients confirmed by pathology from our hospital. After sample collection processing and suspensions, we performed scRNA-seq following the manufacturer’s protocol. Briefly, the cells were washed with PBS and resuspended in 500 μl PBS. scRNA-seq libraries were prepared using a Chromium Single cell 3′ Reagent kit, version 2. Amplified cDNA and final libraries were evaluated using a High Sensitivity DNA Kit (Agilent Technologies). Sequencing was performed on NovaSeq 6000 (Illumina) at a depth of approximately 400M reads.http://support.10Xgenomics.com/single-cell-gene-expression/software/overview/welcome) provided by 10x Genomics was used to demultiplex cellular barcodes. Unique molecular identifier (UMI) counts were obtained by mapping reads to the human reference genome (GRCH38 3.1.0) genome and align transcriptomes using the STAR aligner and down-sample reads as required to generate normalized aggregate data across samples. In the end, a matrix of gene counts by cells was produced. We processed the UMI count matrix using the Seurat R package (version 4.0.2), resulting in 34,851 cells with 36,601 genes for PCNSL samples.The Cell Ranger software pipeline  the number of expressed genes was <200 or >2,000; (ii) the number of total counts was > 20,000; and (iii) the percentage of mitochondrial counts > 10%. We only kept the genes detected in at least 20 cells. After applying these quality control criteria, we obtained 20,307 cells with 12,229 genes in total, which were used for downstream analysis.https://ega-archive.org/studies/EGAS00001004422). For accessibility reasons, we directly used the cellranger matrices from HFA567_total.filtered_gene_matrices, HFA570_total.filtered_gene_matrices, and HFA571_total.filtered_gene_matrices (https://datahub-262-c54.p.genap.ca/GBM_paper_data/GBM_cellranger_matrix.tar.gz) . The raw data are from the European Genome-Phenome Archive: EGAS00001004422 (http://bio-bigdata.hrbmu.edu.cn/CellMarker/) database. (ii) By comparing scRNA-seq with HumanPrimaryCellAtlasData and BlueprintEncodeData data, we utilized both the SingleR R package (version 1.4.1) and manual work to assign cell types to cluster automatic annotation. Finally, we compare the annotation results and determine the cell types.scRNA-seq data were normalized using log transforms and scaled so that the mean expression across cells is 0 and the variance across cells is 1. Then, we performed dimension reduction and visualization on the single-cell data through the Seurat R package (version 4.0.2) . We usedhttps://david.ncifcrf.gov/summary.jsp). It is an essential tool for systematically extracting biological information from a set of genes. Adj. p-value < 0.05 was considered statistically significant.We then performed gene ontology (GO) enrichment analysis using the database for annotation, visualization, and integrated discovery database  to conduct interactive analysis on cell subpopulations. CellPhoneDB is a publicly available repository of selected receptors, ligands, and their interactions . Here, wAfter quality filtering, 40,322 cells with 20,452 genes were obtained from the PCNSL and nHFB samples . All cells were classified into 13 subpopulations by combining the automatic and manual annotation methods. The cell types were identified as B cell, T cell, macrophage, dendritic cell, oligodendrocyte, meningeal cell, excitatory neuron (EN), interneuron (IN), radial glia (RG), inhibitory neuronal progenitor cell (INP), excitatory neuronal progenitor (ENP), astrocyte, and oligodendrocyte progenitor cell (OPC) Table 1.CD79A served as a typical marker gene of the B cell population in total PCNSL cells  of the classical B cells. The B cell-2 cluster was significantly marked by, e.g., NCL, LTB, NEAT1, and enriched functional pathways of antigen processing and presentation of endogenous peptide antigen via MHC class I and positive regulation of T cell-mediated cytotoxicity, suggesting that these B cells have strong antigen–antibody immunity and synergistic disturbance with T cells. The B cell-3 cluster  indicated functions of cell–cell adhesion regulation, neuroinflammation response, and hydrogen peroxide biosynthetic process. Plasma cells  played a significant role in complement activation.Malignant proliferation and abnormal differentiation of B cells (toward plasma cells) are the main pathological characteristics of PCNSL. Molecular targeted therapy targeting B cells has also been considered as a promising way to treat PCNSL in recent years. Nevertheless, the heterogeneity of B cells in PCNSL remains unclear. Therefore, we first analyzed the heterogeneity of B-cell subtypes and genetic characteristics in PCNSL from single-cell resolution. Cell–cell communication is a fundamental process that shapes biological tissue. In particular, ligand–receptor pairs can infer intercellular communication from the coordinated expression of their homologous genes . Here, wLTB serves as a marker gene of the T cell population . We identified that B cells can be divided into three subgroups with completely different gene expression and function, including a small number of terminally differentiated plasma cells. In addition to the subsets of normal B cells (B cell-1 overexpressed CD79A and CD79B), B cell-2 showed an important role in antigen processing and regulating the cytotoxicity of T cells, and its function was similar to that of dendritic cells, which may be related to the abnormal differentiation of B cells in PCNSL. This also indicated from the molecular phenotype that B cells in PCNSL were in the pathological overactivation state.Cell–cell communication between B cells and other immune cells is also an important mechanism for the progression of PCNSL disease , 15, 33.In addition, we also demonstrated that immune cells would be affected by the tumor microenvironment. Heterogeneities were also observed in these cell types. We identified T cell, T helper cell, NKT cell, and MPC subtypes within T cell populations that possessed different ontologies. Moreover, we also identified three subtypes within the dendritic cells, including conventional dendritic cell, myeloid dendritic cell, and plasmacytoid dendritic cell. They also enriched into NF-κB signaling, antigen processing and presentation, and neutrophil degranulation pathways, respectively. Further studies on the immune regulation mechanism of these subtypes of T cells and dendritic cells might provide a new strategy for the treatment of PCNSL.In conclusion, our single-cell analysis enabled us to study the cellular heterogeneity and gene expression characteristics of PCNSL at a single-cell resolution and whole-transcriptome scale. Our study is the first to analyze the cellular and molecular pathological map at single-cell resolution. Despite the small size of single-cell samples in our study, it reveals novel potential for targeted immunotherapy of PCNSL.https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE181304).The raw and processed data supporting the findings in this study are available from the corresponding author, and has been submitted to the Gene Expression Omnibus (GEO) database under the accession code GSE181304  for the publication of any potentially identifiable images or data included in this article.Conceptualization, B.Y.W. and Z.L.; Methodology, B.Y.W. and Z.L.; Validation, Y.F. and G.C.; Formal Analysis, B.Y.W. and Z.L.; Resources, B.Y.W., S.W.W., C.D., W.R. and F.Y.; Data Curation, Y.F.; Visualization, Y.F.; Writing-Original Draft, B.Y.W., Z. L. and Y.F.; Writing-Review & Editing, B.Y.W., Z.L, and J.N.Z.; Supervision, G.C., and J.N.Z. All authors contributed to the article and approved the submitted version.This study was funded by the Innovation Cultivation Fund of the Sixth Medical Center of PLA General Hospital (CXPY201913) and the Youth Science Foundation Project of the National Natural Science Foundation of China (81801165).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.


Polyalthopsis Chaowasku is a recently newly described genus in Annonaceae. Polyalthopsis verrucipes (C.Y.Wu ex P.T.Li) B.Xue & Y.H.Tan is distributed in Southern Yunnan, China. In this article, we report the complete chloroplast (cp) genome of Polyalthiopsis based on Illumina sequencing data. The whole cp genome of this species is 159,965 bp in length, consisting of two inverted repeat regions , one large single-copy region , and one small single-copy region . A total of 130 genes were annotated for the cp genome, including 83 protein-coding genes, 37 tRNAs, and 8 rRNAs. Phylogenetic analysis indicated that P. verrucipes was closely related to Meiogyne hainanensis. Polyalthiopsis Chaowasku is a recently published small genus in Annonaceae  B.Xue & Y.H.Tan is distributed in Menghai, Yunnan, with restricted distribution and small populations. It was transferred from the genus Polyalthia Blume  genome sequence of Polyalthopsis verrucipes were collected from Menghai, Xishuangbanna, Yunnan province, China . The total genomic DNA of the species was extracted using the modified CTAB method (Doyle and Doyle B. Xue & H.B. Ding XB311) was deposited in the herbarium of South China Botanical Garden, Chinese Academy of Sciences (IBSC) with the barcode number IBSC0860085. Library construction and sequencing were performed by BGI-Shenzhen , using an Illumina HisSeq 2500 Sequencing System following the manufacturer’s instructions. The cp genome was assembled by the GetOrganelle  (recorded as Chieniodendron hainanense in Genbank) as a reference, the annotations were implemented using the online software Geseq  region of 89,030 bp, a small single-copy (SSC) region of 18,987 bp, and two inverted repeat (IR) regions of 25,974 bp each. The whole cp genome has 130 genes in total, including 85 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. In addition, the overall GC content of the genome is 39.05%. In order to investigate the phylogenetic relationships of P. verrucipes with other species in Annonaceae and outgroup species, another two complete cp genomes from Annonaceae and 11 complete cp genomes from several closely related families Aristolochiaceae, Magnoliaceae, Myristicaceae, and Lauraceae were obtained from GenBank and aligned with P. verrucipes using MAFFT (Katoh and Standley Aristolochia macrophylla (GenBank Accession NC_041453.1) and Aristolochia tubiflora (Genbank Accession NC_041456.1) were selected as outgroups. The phylogeny showed that P. verrucipes was closely related to genus Meiogyne in Annonaceae; the family Annonaceae is closely related to Magnoliaceae and Myristicaceae (The complete cp genome of ticaceae .