Locusta migratoria manilensis♀1C = 6.60 pg, Angaracris rhodopa♀1C = 16.36 pg) to ascertain the influence of piRNAs.Transposable elements (TEs) have been likened to parasites in the genome that reproduce and move ceaselessly in the host, continuously enlarging the host genome. However, the Piwi-interacting RNA (piRNA) pathway defends animal genomes against the harmful consequences of TE invasion by imposing small-RNA-mediated silencing. Here we compare the TE activity of two grasshopper species with different genome sizes in Acrididae  were shared in both species. The two species exhibited distinct “landscapes” of TE divergence. The TEs outbreaks in the small-genome grasshopper occurred at more ancient times, while the large-genome grasshopper maintains active transposition events in the recent past. Evolutionary history studies on TEs suggest that TEs may be subject to different dynamics and resistances in these two species. We found that TE transcript abundance was higher in the large-genome grasshopper and the TE-derived piRNAs abundance was lower than in the small-genome grasshopper. In addition, we found that the piRNA methylase HENMT, which is underexpressed in the large-genome grasshopper, impedes the piRNA silencing to a lower level.We discovered that repetitive sequences accounted for 74.56% of the genome in Our study revealed that the abundance of piRNAs is lower in the gigantic genome grasshopper than in the small genome grasshopper. In addition, the key gene HENMT in the piRNA biogenesis pathway (Ping-Pong cycle) in the gigantic genome grasshopper is underexpressed. We hypothesize that low-level piRNA silencing unbalances the original positive correlation between TEs and piRNAs, and triggers TEs to proliferate out of control, which may be one of the reasons for the gigantism of grasshopper genomes.The online version contains supplementary material available at 10.1186/s12915-022-01441-w. Metazoan haploid nuclear genome sizes  range from 0.02 to 132.83 pg and exhibit more than 6600-fold variation , salamanTEs, as selfish DNA, have the ability to move around and replicate themselves within the genomes –20. At tDrosophila melanogaster, post-transcriptional silencing of TEs is based on Aubergine (AUB) and Argonaute3 (AGO3) binding directly to piRNAs, guided by the specific binding of piRNAs to TE transcripts [The Piwi-interacting RNA (piRNA) pathway is a critical regulator of germline TE activity. This host defense system relies on piRNAs that bind to PIWI-clade proteins and suppress TE activity transcriptionally and post-transcriptionally –36. In Dnscripts –44. Antinscripts , 45. Thenscripts , 47. Addnscripts , 48, 49.TE activity is highly dynamic during evolution, and the host genome faces a constant onslaught of reactivated or horizontally transferred TE families . The hopLocusta migratoria manilensis♀1C = 6.60 pg, Angaracris rhodopa♀1C = 16.36 pg) to investigate the genome repeat composition and evolutionary history of the TEs found in the two species using low-coverage Illumina sequencing short reads. The intraspecies comparison of the abundance of retrotransposon transcripts in different tissue types was also investigated using RNA sequencing data. Combined with the genomic content of retrotransposons, we compared the abundance of retrotransposon transcripts and piRNAs between the two species. Finally, we discuss the relationship between TEs and piRNAs within and between species. Our study provides new insights into the mystery of grasshopper genome gigantism.Orthoptera is the only known group of in the Insecta class with a significantly enlarged genome , 54 and A. rhodopa (16.36pg) account for 74.56% of the genome and 56.83% in L. migratoria (6.60pg) . The results show that repetitive elements in L. migatoria, and the other is that LINEs proliferated more slowly than other TE subclasses in the process of increasing grasshopper genome size.Within the TE subclasses, the proportions of DNA transposons, short interspersed nuclear elements (SINEs), and LTRs in the large-genome grasshopper are higher than those in the small-genome grasshopper Fig. . Howevery-axis for A. rhodopa than for L. migratoria . TE copies in L. migratoria deviate significantly from that of the consensus sequence (K2P = 5–10%). We discovered that A. rhodopa has a higher genome abundance than L. migratoria when comparing the least divergent (divergence < 1%) TEs elements from their consensus sequences. The results suggest that A. rhodopa has more newly proliferated TE copies, with little divergence from the consensus sequence. Assuming a molecular clock for nucleotide substitutions within duplicated TEs, a smaller divergence from the consensus sequence indicates a recent active transposition event [A.rhodopa, while more ancient proliferation events occurred in L. migratoria.To test whether the observed abundance patterns of specific TEs were driven by ancient proliferation events or by recent activities, we first generated divergence “landscapes” for TEs within each genome using dnaPipeTE (see the “Methods” section). The landscapes measure the amount of sequence divergence between each copy of TE. Histograms of the resulting Kimura 2-parameter distance (K2P) provide insights into the evolutionary history of TE activity –57. The on event . We consA.rhodopa.TEs are extremely unconserved within and between species, and variation exists even between TE copies. In the TE comparative analysis of two species, it is crucial to find the consensus sequence shared between the two species. For this, we used the comparative analysis script of dnaPipeTE (see the “Methods” section), and the results revealed 162 shared repeats (copy number > 100) in the two species . The top 10 TEs with the highest number of copies are shown in Fig. In the TE divergence landscapes, we inferred that the sequences with K2P deviation < 5% have recently undergone frequent transposition events. In addition, the transposons transcriptome results can better illustrate the transcriptional activity of TEs.A. rhodopa from four subclasses of retrotransposon transcripts  . In contrast, LINE and Penelope elements did not display tissue-specific expression  in both species . We obtained sequencing data for three biological replicates per tissue sample. After assembly, we used the DANTE tool and extracted the domains of group-specific antigen (GAG), protease (PROT), reverse transcriptase (RT), ribonuclease H (RH), and integrase (INT) for retrotransposon structural annotation (see Methods). Finally, we discovered 101 retrotransposon transcripts (TPM>1) in L. migratoria and 154 transcripts (TPM>1) in Transcription of retrotransposons is only the first step in the entire transposition event, which is followed by reverse transcription and integration. We also investigated the expression differences of reverse transcriptase and integrase in tissues. Analysis of transcript abundance of the RT and INT domains revealed that reverse transcriptase and integrase were highly expressed in testis tissue Fig. c, d.A. rhodopa  and 10.13% (A. rhodopa testis) of clean data, respectively. After that, we performed length statistics on the remaining small RNAs. The small RNAs in the testis of the two species showed different length distributions , and found that the abundance of miRNAs in A. rhodopa was higher than that in L. migratoria. The A. rhodopa small RNAs consisted of 41.46% of miRNAs. Meanwhile, miRNAs in L. migratoria only accounted for 21.41% of total small RNAs. The remaining small RNAs were aligned with TEs to identify TE-derived piRNAs (see the “Methods” section). We selected aligned small RNAs with a length of 23–31 nt and identified them as TE-derived piRNAs. We performed base bias analysis on the identified TE-derived piRNAs. 82.3% and 72.3% of the TE-derived piRNAs in L. manilensis and A. rhodopa, respectively, had uridine in the first position of the 5′-end (referred as “1U”) . Reads of mapped rRNA accounted for 4.90% , while the large-genome grasshopper has a lower TE-derived piRNAs abundance of 48,702.81 (RPM) , and HEN methyltransferase 1 (HENMT) in the gonads [To explore what causes the low abundance of piRNAs in the large-genome grasshopper, we analyzed key genes in the piRNA pathway, including AGO3, PIWI2, PIWI3 , and Ty3_gypsy elements showed relatively weak correlations    Fig. b. We susNAs Fig. b.L. migratoria but no correlation in A. rhodopa. We also found that the slope of the fitted line was always greater for L. migratoria than for A. rhodopa  [Clunio tsushimensis, 1C value =0.07 pg) [Insect genomes are characterized by high heterozygosity and duplication , and gen8.64 pg) , which i0.07 pg) –75. Thus0.07 pg) , studies0.07 pg) –79.Our study of the TEs divergence landscape found that TEs exhibited distinct burst patterns between species Fig. a, b. TheA. rhodopa. In scaled profiles, we consider the position with the highest read coverage as the TE origin, which is accompanied by an increase in copy number and extension of the sequence end as TE jumps and proliferates. We found that the large-genome grasshopper has more copies of end extensions.To compare differences in TE copy accumulation across species, we performed a scaled-profiles analysis for shared TEs. On a copy number scale, the giant genome grasshopper species has a higher copy number of repetitive elements. It is assumed that these shared-TEs existed during the common ancestor of the two species, and these TEs have undergone the same temporal evolution in different hosts. We observed that these TEs accumulate more copies with a faster expansion rate in A. rhodopa, whereas degeneration or inactivation of TEs has occurred in L. migratoria. The different landscape patterns in the two species illustrate that TEs are subject to different dynamics and resistances as they expand. TEs suffer different fates, which may be related to the host defense mechanism against TE invasion.In addition, the TE divergence landscapes showed that active transposition events have recently occurred in L. migratoria and A. rhodopa. Similar results were found in a study of D. melanogaster and D. simulans [TEs choose the main battlefield in the germline, where even the most harmful TE insertions are heritable , 82. Somsimulans .The transposition of TE in somatic cells cannot be ignored. Although its transposition in somatic cells will not be inherited, it will harm the adaptability of the host. In humans, the expression and transposition of LINE elements have been detected in various somatic contexts, including early embryos and certain stem cells , 86. SomL. migratoria: r = −0.13, p = 0.42; A. rhodopa: r = −0.18, p = 0.27; Pearson correlation coefficient) , but there was a negative linear correlation in L. migratoria  , which triggers the production of piRNAs –94. BaseThe germline-specific ping-pong amplification cycle has been demonstrated to produce antisense piRNAs from piRNA precursor transcripts, and the TE transcript is degraded to form sense piRNAs through a “secondary” piRNA pathway , 95. MorD. melanogaster revealed the existence of an evolutionary arms race between the copy numbers of TEs and the abundance of piRNAs [It is interesting to discuss the relationship between piRNAs and TEs within species because piRNAs originate from TE-rich regions of the genome and can inhibit TE transposition. A study of the Global Diversity Lines (GDL) of f piRNAs . In addif piRNAs . These rf piRNAs , 98.L. migratoria fitted line is always greater than that of A. rhodopa, suggesting that piRNAs have better control over TE transcripts in the small-genome grasshopper. For example, if TE transcripts have the same abundance in both species, the abundance of corresponding piRNAs is higher in L. migratoria than that in A. rhodopa.Our findings in the grasshopper species with the smaller genome are consistent with the above hypothesis, but not in the grasshopper with the larger genome. We speculate that the uncorrelated association between TE mRNA abundance and TE-derived piRNA abundance in large-genome grasshoppers is due to weaker suppression of particular TE transcripts by low-abundance piRNAs, which leads to rapid accumulation of these TEs. The slope of the Drosophila and L. migratoria is considered a balanced relationship for the host to counteract the damage suffered by TE invasion under normal conditions. The low-level piRNA silencing in the large-genome grasshopper species disrupts the original balance between TEs and piRNAs, causing some TEs to be out of control and continue to expand.The positive correlation between TEs and TE-derived piRNAs found in Natural selection and genetic drift are powerful forces shaping the distribution and accumulation of TEs . Some stA. rhodopa species was collected at higher altitude areas . At present, we do not have enough evidence to prove this conjecture, and we need more samples from extreme living environments. Many questions about the host’s response to TE invasion remain unanswered. Whether this low-level piRNA silencing is unique to gigantic genome grasshopper species, or is an evolutionary process of Acrididae insects, requires more species data to reveal.There may be another possibility that the expansion of TEs may bring some evolutionary advantages to the host. Large amounts of DNA insertion or deletion would result in a high genome plasticity . ResearcA. rhodopa, which has a larger genome (16.36 pg). We found that the expansion of TEs in the large-genome grasshopper species was more rapidly manifested by the accumulation of more repeat copies. The different levels of the two hosts in response to TE invasion may be the main reason for the different expansion patterns of TEs in the two grasshopper species. By comparing the piRNA silencing mechanisms of the two species, we found that the piRNA methylase HENMT, which is underexpressed in the large-genome grasshopper, made piRNA abundance lower than that in the small-genome grasshopper, breaking the original balance between TEs and piRNAs. In summary, we hypothesize that low levels of piRNA silencing lead to an imbalance in the relationship between TEs and piRNAs in the host, resulting in a rapid expansion of TEs leading to genomic gigantism.In this study, we analyzed the activity of TEs in two grasshopper species using genomic and transcriptomic datasets. The results showed that the transposition of TEs was more active in L. migratoria under experimental rearing conditions and A. rhodopa were collected at the Tibetan Autonomous Prefecture of Haibei, Qinghai, China  in August 2020. Live adults were taken to the laboratory for dissection. The DNA-grade samples were added to 95% ethanol and stored in a −20°C freezer. The RNA-grade tissues from males  and females  were dissected and stored in RNAlater  stored at −80°C until subsequent RNA extraction. The head, thorax, and legs of individual genders were mixed into one sample as a body tissue for RNA extraction. We sampled three biological replicates for each tissue sample. Furthermore, the freshly collected samples were used to estimate the genome size using flow cytometry (FCM) of propidium iodide-stained nuclei following the standard protocol [Samples of protocol , 108.A. rhodopa from the hind leg of one female using an SDS-based lysis method and purified the DNA with chloroform. The extracted DNA was sonicated to a fragment size of 350 bp. The library was fixed onto a microarray by bridge PCR and sequenced using the Illumina HiSeq 2500 sequencing platform (PE150bp). The genomic data for L. migratoria was downloaded in the Sequence Read Archive (SRA), accession number SRR764584.We extracted the genomic DNA of RNA sequencing libraries were generated using NEBNext Ultra RNA Library Prep Kit for Illumina . The clustering of the index-coded samples was performed on a cBot Cluster Generation System using TruSeq PE Cluster Kit v3-cBot-HS (Illumina). The library preparations were sequenced on an Illumina NovaSeq 6000 platform, and paired-end reads were generated. Small RNA Sequencing libraries were generated using NEBNext Multiplex Small RNA Library Prep Set for Illumina (NEB). After that, the different libraries are pooled according to the effective concentration and the target amount of data off the machine, and 50 bp single-end reads are generated by Illumina NovaSeq 6000 sequencing.We used 0.1× genome coverage sequencing data for repeat sequences analysis with dnaPipeTE software . The dnahttps://github.com/clemgoub/dnaPT_utils) in dnaPipeTE to perform the comparative analysis of repeat sequences in the two species (dnaPT_compare.sh -A Locus_dnaPipeTE.OUT -a Locus -B Angar_dnaPipeTE.OUT -b Angar -o compare.out -T -e 500 -E -C 36). We selected shared TEs with more than 500 copies in both species for display.TE landscapes are automatically generated in the dnaPipeTE output file. The consensus sequences used in the TE divergence landscape analysis are the respective annotated TEs in each species. We used dnaPT_compare.sh (https://github.com/johnssproul/RepeatProfiler) for visualizing and comparing repetitive DNA profiles of 41 shared TEs from 0.5× coverage short-read sequence data [We used the RepeatProfiler tool  to annotate the assembly results. UniProtKB (https://www.uniprot.org/) and Pfam [https://repeatexplorer-elixir.cerit-sc.cz/galaxy). We choose taxon and protein domain database version as REXdb (Metazoa_version_3.1). The minimum similarity parameter is set to 0.65, and the minimum alignment length parameter is set to 0.7. The structure of LTR retrotransposons and retroviruses are very similar, and they also encode a viral particle coat (GAG) and reverse transcriptase (RT), ribonuclease H (RH), and integrase (IN) [Raw reads from all libraries were processed to remove sequencing adaptors and low-quality bases on the 3′ end using trimmomatic v0.39 , and cleand Pfam  referencand Pfam  (diamondase (IN) . Accordiase (IN) , 116. Wease (IN) .https://github.com/trinityrnaseq/trinityrnaseq/tree/master/util) script . Then we extracted the TPM normalized expression matrices of retrotransposons and piRNA pathway genes separately based on DANTE, Pfam, and UniProtKB annotation results. Boxplots of each transcript abundance of retrotransposons and piRNA pathway genes were plotted by R packages (“ggplot” and “ggboxplot”), and significant differences were performed using T-test.Quantitative analysis of all transcripts was performed through the align_and_estimate_abundance.pl  [L. migratoria [We deep-sequenced small RNAs from testis, ovaries, and bodies of t (v3.3)  softwaret (v3.3)  databaset (v3.3)  to removgratoria –123. Thegratoria , 124 ) + geom_point + geom_smooth  + stat_cor).http://repeatmasker.org) with the “-a” option and the RMBlast search engine to estimate the divergence of each shared-TEs (RepeatMasker 0.1x.fa -lib 41sharedTEs.fa -a -e rmblast)  (createSatellitome1Landscape.pl -div name.divsum -g genome_size).We used RepeatMasker . Table S4. Annotations of Class I retrotransposon transcript (TPM>1). Table S5. Retrotransposon transcript quantification matrix  of L. migratoria.Table S6. Retrotransposon transcript quantification matrix  of A. rhodopa.Table S7. Abundance of piRNAs corresponding to 41 shared TEs . Table S8. Abundance of sense piRNAs corresponding to L. migratoria retrotransposon transcripts . Table S9. Abundance of antisense piRNAs corresponding to L. migratoria retrotransposon transcripts . Table S10. Abundance of sense piRNAs corresponding to A. rhodopa retrotransposon transcripts . Table S11. Abundance of antisense piRNAs corresponding to A. rhodopa retrotransposon transcripts . Table S12. List of K2P divergence and abundance of 41 shared-TEs.


Artocarpus heterophyllus) is an important cultivated crop in tropical regions of South and Southeast Asia. The species has been cultivated in China for more than 1000 years, but the history of its introduction to the country remains unclear. We assembled a high-quality chromosome-level genome of jackfruit into 985.63 Mb with scaffold N50 of 32.81 Mb. We analyzed whole-genome resequencing data of 295 landraces to investigate the domestication history in China and agronomic trait evolution of jackfruit. Population structure analysis revealed that jackfruits of China could be traced back to originate from Southeast Asia and South Asia independently. Selection signals between jackfruit and its edible congener, cempedak (Artocarpus integer), revealed several important candidate genes associated with fruit development and ripening. Moreover, analyses of selective sweeps and gene expression revealed that the AhePG1 gene may be the major factor in determining fruit texture. This study not only resolves the origins of jackfruit of China, but also provides valuable genomic resources for jackfruit breeding improvement and offers insights into fruit size evolution and fruit texture changes.As the largest known tree-borne fruit in the world, jackfruit ( Published by Oxford University Press on behalf of Nanjing Agricultural University. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

Received: 29 March 2022; Revised: 21 June 2022; Accepted: 25 July 2022
 Artocarpus comprises ~70 species of evergreen flowering trees in the mulberry family (Moraceae) and is widely distributed from South and Southeast Asia into Oceania  throughout the whole genome with 10-kb sliding windows. In total, we identified 1074 genes of 4290 CDRs as being under selection   and two unannotated genes  . It is a–3.5 kg) . To idenelection . These g = .003) , contained genes . CalS ishomology . Moreovehomology . The jacAHE.Chr18.535, AHE.Chr18.642), and one auxin transport protein, BIG (AHE.Chr22.1004), within the regions under selection , phosphatidylinositol-bisphosphatase , and polygalacturonase   and AhePG2 (AHE.Chr24.22) loci separated most of the firm-type accessions from the soft-type accessions  . Specifickground  and b. Awo types  and d. Wirm type  and f. Tcessions , indicatAhePG1 gene increased sharply along with the decrease in jackfruit firmness and the enhancement of fruit softening. The gene expression of AhePG1 was significantly negatively correlated with fruit firmness  and South Asia (Bangladesh), both of which might trace back to the Silk Road. The Silk Road was a grouping of major historical trade arteries and cultural transmission routes in Asia, and primarily consisted of two major routes, the Overland Silk Road and the Maritime Silk Road . These rArabidopsis, five out of the 12 CalSs, including CalS5, CalS9, CalS10, CalS11, and CalS12, play important roles during sporophytic development and pollination } as CDRs in populations of A. heterophyllus; Z-transformed FST = (per window FST − mean FST)/standard deviation of FST  in the 500-kb region of chromosome 24, and performed KEGG analysis with the R package clusterProfiler.We found high population differentiation in the 500-kb region of chromosome 24 between the firm type (FF group) and soft type (SF group) of jackfruit. To test for possible positive selection, we measured the cross-population extended haplotype homozygosity (XP-EHH) , two-pophttp://ccb.jhu.edu/software/hisat/index.shtml), and assembled them based on the alignment results with StringTie [AhePG1 gene expression and fruit firmness using the cor function in R and fitted the result by linear regression using the lm function. We removed the genes of FPKM value <1 in all samples, and drew a gene expression heat map of the remaining genes using the R package ComplexHeatmap (http://www.bioconductor.org/packages/3.8/bioc/html/ComplexHeatmap.html).To investigate the expression pattern of genes associated with jackfruit texture, we collected jackfruit pulps of different firmness during fruit softening. We measured the hardness value of jackfruit pulps using a hardness tester, taking six biological replicates for each group. Then, we mixed the samples within each group and subjected them to transcriptome analysis. We trimmed the RNA sequences to remove adaptors and contaminating sequences, and preprocessed to remove low-quality sequences with QC script . Then wetringTie . Next, wtringTie . We perfThis work was supported by the Institution of South China Sea Ecology and Environmental Engineering, Chinese Academy of Sciences (No. ISEE 2018YB02), Projects of Enhancing School with Innovation of Guangdong Ocean University , and Science and Technology Projects of Guangzhou City (202102020341).M.K. and Y.L. designed and supervised the project. T.L. collected materials. X.L. performed the experiments and bioinformatics analysis. X.L. designed and visualized the figures. X.L. and M.K. wrote the manuscript. M.K., C.F., and A.J.H. revised the manuscript. All authors read and proved the final version of this manuscript.The raw genomic Illumina sequences, PacBio sequences, and transcriptome data have been deposited in the NCBI Sequence Read Archive under accession numbers PRJNA788174 and PRJNA791757.The authors declare no conflict of interests.Horticulture Research online.Web_Material_uhac173Click here for additional data file.


Biologics have become an important area of medical research generating therapeutics essential for the treatment of many disease states. Biologics are defined as biologically active compounds manufactured by living cells or through biological processes termed bioprocessing. Compared to small molecules which are chemically synthesised they are relatively complex and therapeutically specific molecules. Biologics include hormones, vaccines, blood products, monoclonal antibodies, recombinant therapeutic proteins, enzymes, gene and cellular therapies amongst others. For biologic production prokaryotic and eukaryotic cells  are used as expression systems. Eukaryotic expression systems offer many advantages over prokaryotic based systems. The manufacture of high-quality proteins for human clinical use via recombinant technologies has been achieved in yeast and filamentous fungal systems. Advances in bioprocessing such as genetic engineering, bioreactor design, continuous processing, and quality by design has allowed for increased productivity and higher yield in in these non-mammalian eukaryotic systems with protein translation similar to mammalian systems. The application of eukaryotic expressions systems for the manufacture of biologics of therapeutic importance are described herein. Escherichia coli, eukaryotic yeast and fungal systems, e.g., Saccharomyces cerevisiae, Aspergillus, plant systems, insect systems, mammalian and human expression systems and cell lines [The manufacture of biologics is a rapidly growing industry as these specific therapeutics offer targeted treatment approach for many chronic and prevalent medical conditions including cancer, cardiac disease, neurological disease and autoimmunity. According to the Food and Drug Administration (FDA), a biologic is a therapeutic substance produced by a biological process using biological systems as opposed to the process of chemical synthesis  and includes vaccines, antibody therapies, non-vaccine therapeutic immunotherapies, gene therapies and cell therapy . While bll lines . Currentll lines . The bioll lines . Indeed,ll lines . The bioll lines . In cancll lines . More rell lines . Recentlll lines .Aspergillus and Penicillium are considered as GRAS by the FDA and are used as expression systems by many biotechnology companies to produce varied biological products [Bioprocessing occurs in 4 phases: strain/cell line selection and propagation, upstream processing (fermentation), downstream processing, and drug formulation with one biologic usually produced from 1 cell strain . The bioproducts . In contproducts . This reEukaryotic cell lines, including CHO cells, human cells and insect cells, are invaluable expression systems for the production of many recombinant proteins . MammaliSaccharomyces cerevisiae is used for recombinant therapeutic protein production having been applied in the production of hormones glucagon and insulin and Hepatitis B vaccine, at industrial scale [S. cerevisiae [Pichia pastoris (renamed to Komagataella phaffii) are the expression systems used in vaccine development, protozoal proteins and tumour antigens [S. cerevisiae also displays a broad range of pH tolerance and elevated osmotic pressure [S. cerevisiae can be done using 3 types of vectors: integration plasmids (YIp), episomal plasmids (YEp), and centromeric plasmids (YCp) [P. pastoris is also amenable to bioengineering where genetically engineered strains are able to produce heterologous biologics with human glycosylation profiles [P. pastoris produces a higher protein yield than S. cerevisiae due to its biochemical processes relating to a lack of ethanol production under aerobic conditions allowing for a higher biomass and protein production [P. pastoris favours respiration over fermentation, S. cerevisiae produces ethanol under aerobic conditions with glucose as a food source reducing protein yield, termed the Crabtree effect [P. pastoris is currently used in the production of interleukin 1-β for the treatment of autoimmune disease, interferon-α for the treatment of hepatitis B and C and cancer and macrophage colony-stimulating factor (M-CSF) for the treatment of hematopoietic disorders [P. pastoris has also been implemented in the production of human insulin, human serum albumin, hepatitis B vaccine, trypsin, and collagen, among others [P. pastoris however, remains a non-conventional yeast where the genome has not been fully elucidated which hinders genetic engineering unlike the well-established conventional S. cerevisiae [P. pastoris can efficiently produce and secrete fully active heterologous proteins, e.g., pochymosin, poorly secreted by S. cerevisiae [Yarrowia lipolytica is recognised for its ability to produce heterologous recombinant proteins. Y. lipolytica is a human commensal, obligate aerobe, GRAS and is capable of folding and secreting large and/or complex heterologous proteins in contrast to S. cerevisiae [S. cerevisiae recombinant proteins [Y. lipolytica strains produce several heterologous valuable metabolites including carotenoids, terpenes, polyketides, aromatic amino-acid-derived molecules and therapeutic biologics interferon α, epidermal growth factor, blood coagulation factor XIIIa, proinsulin and insulinotropin, cytochrome P450 enzymes, and oestrogen receptor α [Y. lipolytica expression systems however, only 25 of these are produced at industrial scale or bioreactor scale despite this species having similar yield and productivity as S. cerevisiae and P. pastoris [Ogataea polymorpha (formerly Hansenula polymorpha) has been used as an expression system for the production of hepatitis B vaccines and insulin-like growth factors [H. polymorpha is thermotolerant with a temperature range of 30 to 50 °C allowing proteins with biological activity at 37 °C as obtained with other yeast including S. cerevisiae with PTM and reduced hyperglycosylation [Yeast are single celled microorganisms within the Fungus kingdom, being defined as unicellular fungi. Yeast are eukaryotic microbial species having a cell wall and membrane bound organelle unlike bacteria being prokaryotic. Fungi growing as a yeast morphology have historically been used in the production of food and beverages . Due to al scale . Indeed,revisiae . Currentantigens . S. cerepressure . Productds (YCp) . P. pastprofiles . P. pastoduction . While Pe effect . P. pastisorders . P. pastg others . P. pastrevisiae . P. pastrevisiae . The abirevisiae . The nonrevisiae . Indeed,proteins . Engineeceptor α . Currentpastoris . Scale upastoris . The non factors . H. polysylation .Aspergillus terreus a filamentous fungus, where yeast S. cerevisiae and P. pastoris may offer a more favourable non-pathogenic production system [Polyketides and non-ribosomal peptides constitute a group of small molecules having complex chemical structures produced by microbial species, plants and marine organisms which allow for environmental adaption, communication between species and self-protection . Polyketn system .S. cerevisiae offers many advantages in the production of vaccines including strong adjuvant properties, long term antigen stability, ease of GM and ability to survive the gastrointestinal tract allowing for oral administration [S. cerevisiae cells expressing foreign antigens can activate dendritic cells (DCs), robust antigen-specific cytotoxic T lymphocyte (CTL) responses, and confer protective cell-mediated immunity in animal studies regardless of yeast cell viability [Recent advances have moved towards protein-based vaccines, virus-like particle vaccines, viral vector and nucleic acid-based vaccine production in an attempt to mitigate the time constraints, pathogenicity, immunogenicity and biocompatibility issues observed with traditional vaccine types . Recombistration . Studiesiability . WYVs baiability .S. cerevisiae results in an immunogenic reaction in humans [P. pastoris is less prone to this mannose hyperglycosylation [E. coli and Bacillus subtillis and yeast S. cerevisiae and P. pastoris have been used to express rAbs [S. cerevisiae as an in vitro nanobody platform [Monoclonal antibodies (mAb) have many therapeutic applications in the treatment of infectious disease, inflammation and cancer where their glycosylation is key to biologically . The pron humans . P. pastsylation . Experimsylation . Geneticsylation . Nonethesylation . Therapysylation . Biologisylation . RDNA teess rAbs . In ordeess rAbs . Antibodess rAbs . Antibodess rAbs . The PD-ess rAbs . The disess rAbs . Nanobodess rAbs . Studiesplatform .S. cerevisiae. Additionally, fungi expression systems are a cheap fermentation process, where the extracellular proteins are easier purified compared to intracellular proteins [Fungal species have a long history in the food and beverage industries; however, many enzymes, organic acids, antimicrobial peptides and antibiotics are also produced by filamentous fungi. The filamentous fungi including moulds and mushrooms having extremely diverse metabolic capabilities producing organic acids, antibiotics, proteins, enzymes, vitamins, fatty acids amongst other compounds leading to their application in many industries and more recently as meat alternatives and vegan leather products . As prodproteins .Aspergillus niger, Aspergillus oryzae, Trichoderma reesei and Neurospora crassa being well characterized by whole-genome-sequencing, transcriptomics, gene annotations, and genetic engineering tools including adaptive laboratory evolution, and radiation [While GM is more difficult in fungi and the development of filamentous fungal expression platforms is much more complex and time-consuming, the protein yield is 10–1000-fold that of yeast or mammalian cells making them worthy of development . For recadiation . Unlike adiation . Productadiation .Fungal bioprocessing is typically performed in either submerged fermentation bioreactors using free flowing liquids or solid-state fermentation systems (SSF) better mimicking the natural growth patterns of filamentous fungi  or as biFusarium oxysporum and A. niger [Mucor hiemalis, A. niger, A. flavus, A. nidulans, A. terreus [Trichoderma has been cultivated using submerged, solid-state and biofilm fermentation to produce the secondary metabolite and immunosuppressant cyclosporin [E. coli expression systems where death is a risk to patients [Filamentous fungi produce a vast array of molecules  which maA. niger . To dateA. niger . Asparag terreus . Trichodlosporin . Optimispatients . Filamenpatients . Additiopatients .P. pastoris and H. polymorpha can use methanol as a carbon source allowing for the use of methanol inducible promoters [S. cerevisiae. There is a need to elucidate the genetic profile, metabolic and biochemical systems of the non-conventional yeast in order to better apply strains such as Y. lipolytica in bioprocessing where their advantages over S. cerevisiae can be harnessed. For example, sequencing of the genome of Y. lipolytica has determined that this species shares genomic traits with filamentous fungi where this species is considered a dimorphic yeast [Y. lipolytica has not been elucidated in bioreactors where the dimphorism variability impacts on heat and mass transfer in the reactor impacting protein formation [P. pastoris, K. lactis, Y. lipolytica, H. polymorpha and S. pombe for the synthesis of recombinant proteins [The first step in yeast recombinant protein production is construction and genetic engineering of the strain for scale up in bioreactors . The GM romoters . At induromoters . The mosromoters . The conromoters . A codonromoters . Studiesromoters . In yeasromoters . In the ic yeast . This moormation . Other nproteins .E. coli and S. cerevisiae are most frequently utilised due to their broad ranging advantages [The operation of bioreactors is affected by cell type, media composition, substrate concentration, cell density of the biocatalyst, product inhibition, pH, temperature with productivity increased in continuous mode over batch systems . Optimisvantages . Synthetvantages .S. cerevisiae, a non-methylotroph, P. pastoris, and Hansenula polymorpha being methylotrophic yeasts are implemented for recombinant protein production. Yeast such as Komagataella sp., Kluyveromyces lactis, and Y. lipolytica have also emerged as advantageous expression systems. As non-mammalian eukaryotic expression systems, S. cerevisiae is a preferred production system for a variety of biologics due to its innate ability to secret proteins, is toxin free, good expression levels, and ability to perform PTMs. PTMs relates to protein biological activity as it impacts on protein folding where misfolded proteins lose activity and stability. While glycosylation in yeast is different than in mammalian cells, the use of RDNA technology has allowed for genetically engineered strains enabling some humanization of yeast glycosylation. S. cerevisiae has also some important disadvantages as this species is not suitable for high-density culture, regulation promoter deficits, and low secretion of proteins greater than 30 kDa. Such limitations have encouraged the application of non-conventional yeasts including P. pastoris as expression systems. Filamentous fungi as expression systems for biologics offers many advantages including glycosylation and other PTMs more similar to human proteins, high density growth adaptability and a high rate of protein expression with low media requirements. Indeed, the secretory potential of filamentous fungi, e.g., Aspergillus sp. and Trichoderma sp. is described as being ten times higher than that of S. cerevisiae. The use of non-mammalian eukaryotic cells also contributes to more environmentally friendly industrial production processes requiring less energy input and generating less waste than mammalian systems. The use of yeast expression systems to produce recombinant protein, virus-like particles, and yeast surface display for development as oral vaccines offers a new strategy for vaccine administration. Optimising eurkaryotic production platforms may provide increased productivity of key therapeutics in the near future. Due to their high-cost comparative to small molecule therapy biologics are relatively inaccessible to many patients and so are not chosen for treatment. Reducing production costs may allow for increased accessibility to these potent therapeutics allowing for better access and improved health care globally.For the production of biologics yeast are classified as either non-methylotroph and methylotroph species. At industrial scale


Correction: BMC Microbiol 22, 168 (2022)https://doi.org/10.1186/s12866-022-02580-4In the originally published version of this article , we wereIn the Table 3 section:S.S.S.S.S.S.S.S.aureus” should be “S. aureus”.“In the Materials and Methods section:Antibacterial and antioxidant activities of compounds“The raw data supporting the conclusions of this manuscript will be made available to the authors, without undue reservation, to any qualified researcher. Publicly available datasets were analyzed in this study. This data can be found here: The obtained ITS gene sequences were deposited in the GenBank database under accession numbers OK184563-OK184606 and OK285068. The obtained 16S rRNA gene sequences were deposited in the GenBank database under accession numbers OK147622-OK147645 and OK169608-OK169611. Data of high-throughput sequencing by paired-end Illumina technology of ITS 1 gene amplicons can be retrieved from the NCBI Short Read Archive under accession number PRJNA817087 and PRJNA817099”. It should be as follows.Antibacterial and antioxidant activities of compoundsE. coli, M. tetragenus, and S. aureus were used to assess the antibacterial activity of compounds. Compounds were dissolved separately in acetone to get a concentration of 6 mg/mL. Acetone and gentamicin sulfate served as negative and positive control, respectively. In addition, compound 1 at different concentrations was selected for the antioxidant experiment. Methanol and ascorbic acid were used as negative and positive controls, respectively. Each sample was performed in triplicate.The IZD of the compounds isolated from MFFC22 were determined using the method as previously described.


Pseudomonas aeruginosa is a Gram-negative opportunistic pathogen frequently isolated from chronic infections of the cystic fibrosis lung and burn wounds, and is a major cause of antimicrobial-resistant nosocomial infections. P. aeruginosa is frequently co-isolated with the opportunistic fungal pathogen Candida albicans, with the presence of C. albicans in dual-species biofilms promoting tolerance to meropenem. Here, transcription profiling of mature P. aeruginosa single- or dual-species biofilms was carried out to understand the molecular mechanism(s) by which C. albicans enhances meropenem tolerance. C. albicans appeared to have a mild impact on the transcriptome of P. aeruginosa mature biofilms, with most differentially regulated genes being involved in interkingdom interactions (i.e. quorum sensing and phenazine biosynthesis). The addition of meropenem to mature single- or dual-species biofilms resulted in a significant bacterial transcriptional response, including the induction of the beta-lactamase, ampC, genes involved in biofilm formation. P. aeruginosa elicited a similar transcriptional response to meropenem in the presence of C. albicans, but C. albicans promoted the expression of additional efflux pumps, which could play roles in increasing the tolerance of P. aeruginosa to meropenem. Pseudomonas aeruginosa is a Gram-negative bacterial pathogen associated with chronic infections in the cystic fibrosis (CF) lung  in Mueller–Hinton broth. Each well contained 3 ml C. albicans and 300 µl of P. aeruginosa in a total of 6 ml. Plates were incubated at 37 °C for 2 h to allow cells to adhere, at which point the media were replaced with fresh sterile media, and plates were incubated statically at 37 °C for 24 h. Cells not part of the biofilm were removed, media were replaced with fresh MHB containing 0 or 5 µg ml−1 meropenem, and plates were incubated for 4 h. Media were replaced with 2 ml PBS containing 50 µg ml−1 DNase I and plates were incubated at 37 °C for 1 h to degrade the extracellular matrix. Biofilms were detached from the plate by scraping, serially diluted and plated onto selective agar .Biofilm assays were based on previously described methodology , but sca−1 tetracycline to check for contamination. Remaining biofilm cells were centrifuged at 3500 r.p.m at 4 °C for 5 min and pellets snap frozen in liquid nitrogen. Four biological replicates were shipped to GeneWiz, UK, for RNA extraction sequencing and basic bioinformatic analysis.Biofilms were formed as described above and triplicate biofilms were pooled, 50 µl serially diluted and plated on cetrimide agar or YPD supplemented with 100 µg mlRNA was extracted from the biofilms by GeneWiz using the Qiagen RNeasy Plus mini kit. Library preparation was done in the following stages: (a) ribosomal RNA depletion; (b) RNA fragmentation and random priming; (c) first and second strand cDNA synthesis; (d) end repair, 5′ phosphorylation and dA-tailing; (e) adapter ligation, PCR enrichment and sequencing. Paired-end sequencing was performed using Illumina HiSeq 4000 .P. aeruginosa reference genome, available on ENSEMBL, using the STAR aligner v.2.5.2b. For dual-species biofilms, samples were treated the same as single-species biofilms. Reads were mapped to the PAO1 genome, and non-mapped reads were discarded and later aligned to the C. albicans reference genome. Unique gene hit counts were calculated using featureCounts from the Subread package v.1.5.2. Only unique reads that fell within exonic regions were counted . For the analysis, only read counts for genes in the P. aeruginosa genome were used and the TPM values were calculated as follows: each read count was divided by the length of each gene in kb to generate reads per kb (RPK), then all the RPK values in the sample were counted and divided by 1 000 000 to generate the scaling factor, and finally the RPK values were divided by the scaling factor to generate the TPM value. Differential gene expression analysis was performed using DESeq2 and the comparisons listed in C. albicans had a reduced impact on the P. aeruginosa transcriptome, resulting in great spread of the data. DESeq2 output files for each comparison, and files containing summary raw and normalized reads and TPM values for each gene are available at the Gene Expression Omnibus (GEO) database (https://www.ncbi.nlm.nih.gov/geo/), under accession number GSE167137.The sequence quality of each sample was evaluated by determining the number of reads, the yield (Mbases), the mean quality score and the percentage of reads over 30 bases in length. FastQC software was used to determine per-base-sequence quality and per-sequence GC content. Sequence reads were trimmed to remove adapter sequences and nucleotides with poor quality using Trimmomatic v.0.36. The trimmed reads were mapped to the P. aeruginosa transcriptomic analysis, differential expression of genes between conditions was considered significant if the adjusted P-value (Padj) was ≤0.05. Gene ontology (GO) analysis was performed using KOBAS 3.0 software [Kyoto Encyclopaedia of Genes and Genomes (KEGG) Orthology Based Annotation System] [For  System] .Biofilm data were analysed in GraphPad Prism (version 9.1.0) using two-way analysis of variance (ANOVA) and Holm–Sidak’s multiple comparisons test.P. aeruginosa is grown in a dual-species biofilm with the fungal pathogen C. albicans, the tolerance of P. aeruginosa to meropenem is increased [P. aeruginosa to meropenem in the presence of C. albicans was still enhanced even at this early timepoint , which encodes a beta-lactamase, important for carbapenem resistance. GO term enrichment analysis of the differentially regulated genes confirmed that meropenem resulted in the significant upregulation of genes involved in the maintenance of the bacterial cell membrane, cell wall, biofilm formation and extracellular matrix production, siderophore production, type IV pilus formation and type VI secretion system, while genes involved in putrescine transport were significantly downregulated  and PA5384 , which were both twofold upregulated in the presence of C. albicans. GO term enrichment analysis  production, pyridocal phosphate biosynthesis, and siderophore transport and pyoverdine biosynthesis were significantly upregulated, while genes involved in type IV pilus biogenesis, the general secretory (Sec) pathway and amino acid biosynthesis were significantly downregulated. Therefore, P. aeruginosa appears to upregulate processes that might provide the bacterium with a competitive advantage when growing in the presence of C. albicans.To determine how the presence of egulated . Althouganalysis  and KEGGanalysis  of the 3P. aeruginosa single-species biofilms treated with meropenem to the list of DEGs identified in untreated dual-species biofilms. Seven genes  were upregulated, while five genes  were downregulated under both conditions. However, as the majority of the genes encode hypothetical proteins, the impact of these transcriptional responses is unknown.To identify whether the presence of the fungus was inducing a similar transcriptional response to the drug treatment we compared DEGs identified in C. albicans affected the response of P. aeruginosa to meropenem, we compared the transcriptional profile of untreated dual-species biofilms to that of meropenem-treated dual-species biofilms. In response to meropenem, a total of 620 genes were differentially regulated, with 304 genes being significantly upregulated and 316 being significantly downregulated . GO term enrichment analysis  were shared with dual-species biofilms  were upregulated in both single- and dual-species biofilms, while 173 (36.3 %) and 123 (25.8 %) were unique to single- and dual-species biofilms, respectively  [ddlA), three were involved in biofilm formation  and three were involved in efflux . This suggests that the presence of C. albicans may increase the ability of P. aeruginosa to form robust biofilms, to secrete molecules via OMVs and upregulate the MexCD-OprJ efflux pump, which may play a role in increasing the tolerance of the bacterium to meropenem.Although ectively . Among t PA5441) , one wasP. aeruginosa biofilm cells, while 227 (41.8 %) and 34 (6.3 %) were unique to single- and dual-species biofilms, respectively . This suggests that the presence of C. albicans may result in reduced production of the MexEF-OprN efflux pump in P. aeruginosa biofilm cells.Of the genes that were significantly downregulated in response to meropenem, 282 (51.9 %) genes were shared in common between single- and dual-species ectively . Among tC. albicans enhances the tolerance of P. aeruginosa to meropenem, a commonly used carbapenem for the treatment of chronic P. aeruginosa infections. Therefore, transcriptional analysis was carried out to understand how C. albicans promotes meropenem tolerance.Biofilms are medically important, as they result in antimicrobial resistance and protect microbes from the actions of the immune system, making infection harder to treat. Furthermore, biofilms are normally mixed-species communities, with our understanding of how microbe–microbe interactions within these communities affect disease progression being limited. We have previously shown that P. aeruginosa single-species biofilms resulted in the significant induction of ampC, a beta-lactamase precursor. Differential regulation of ampC is linked to the natural resistance of P. aeruginosa to this class of antibiotic [P. aeruginosa infections, also induces the expression of ampC [ampC induction as a conserved response to carbapenems. In addition to ampC, the outer-membrane porin, OprD, has also been associated with meropenem resistance. OprD facilitates entry of carbapenems into bacterial cells, and is therefore frequently downregulated in meropenem-resistant strains [oprD was not differentially regulated in response to meropenem treatment. However, there is often a poor correlation between oprD mRNA levels and protein levels [Mechanisms of resistance to the carbapenem class of antibiotics include decreased outer membrane permeability, beta-lactamase expression, increased efflux and target modification. In agreement with this, the addition of meropenem to mature tibiotic . Imipene of ampC , highlig strains . In our n levels . TherefoP. aeruginosa biofilms with either meropenem or imipenem, results in the significant induction of biofilm associated genes [pslB, pslD, pslE and pslF were significantly upregulated. The induction of these polysaccharide genes results in enhanced intracellular and cell–substrate interactions, and the expression of subsets of these genes are commonly enhanced in biofilm-forming clinical isolates. Psl is thought to enhance the tolerance of P. aeruginosa biofilms to a range of antimicrobial agents including colistin, tobramycin and ciprofloxacin [Interestingly, treatment of ed genes . For exafloxacin .ppkA, pppA, clpV1 icmF1, vgrG1 and hcp1. The structure of the T6SS bears similarities to the cell-puncturing needle of bacteriophage viruses; the roles of the T6SS in P. aeruginosa include virulence within hosts, delivery of toxins to neighbouring microbes that are competing for resources, and biofilm formation [P. aeruginosa, the T6SS has been implicated in drug resistance in A. baumannii and K. pneumoniae [P. aeruginosa, the T6SS is upregulated during competition with other bacteria as a result of kin cell lysis [In response to meropenem, genes involved in the type 6 secretion system (T6SS) were also significantly upregulated, including ormation . Althougeumoniae . Howeverll lysis . TherefoP. aeruginosa upregulates genes required for quorum sensing, with all genes from the pqsABCDE operon being significantly upregulated. The pqs operon encodes several enzymes required for production of the Pseudomonas quinolone signal (PQS) [C. albicans-dependent regulation of PQS production is complex. The fungal quorum-sensing molecule, farnesol, inhibits PQS production in planktonic interactions through modulation of PqsR-dependent transcription of the pqs operon [lasR-deficient P. aeruginosa strains, C. albicans restores PQS production through farnesol-induced ROS-dependent production of C4-HSL [During growth in dual-species biofilms, al (PQS) . C. albis operon . Howeverf C4-HSL . Increasf C4-HSL . Howeverf C4-HSL , resultiP. aeruginosa tolerance to the antibiotic ciprofloxacin [Phenazines are redox-active molecules that affect the redox balance of cells, the uptake of metabolites and gene expression, and have also been shown to enhance floxacin . Phenazifloxacin . PathwayP. aeruginosa drug resistance [C. albicans alters the expression of several efflux pumps, which may contribute to carbapenem tolerance in dual-species biofilms. The P. aeruginosa genome encodes for at least 12 RND efflux pumps [P. aeruginosa may increase efflux through MexCD-OprJ. Phenazines and their derivatives have been shown to directly regulate the expression of several efflux pumps, including mexG [P. aeruginosa to meropenem. In agreement with this, enhanced production of the phenazine, pyocyanin, promotes resistance to the beta-lactam class of antibiotics through reduced drug influx [Our transcriptional analysis identified several RND and ABC transporters to the differentially regulated in dual-species biofilms in the presence of meropenem. Little is known about the role of ABC transporters in sistance , but it ux pumps , 3 of whux pumps . MexC waing mexG . Therefog influx .pslH, pslI and algR) and outer-membrane vesicles (OMVs). AlgR is a transcriptional regulator of genes involved in alginate production [C. albicans may function to increase the composition and complexity of the extracellular matrix, which together with enhanced beta-lactamase secretion would increase the tolerance of P. aeruginosa to meropenem.Other differences between the transcriptional profiles for single- and dual-species biofilms in response to meropenem include genes involved in biofilm formation (oduction . Therefooduction , but hasoduction . TherefoP. aeruginosa biofilms upregulate ampC and biofilm formation to provide protection from meropenem. Although dual-species biofilms exhibit enhanced tolerance to meropenem, the P. aeruginosa transcriptional responses from dual-species biofilms exposed to meropenem were similar to those of single-species biofilms treated with meropenem, suggesting that the presence of C. albicans in dual-species biofilms may limit the diffusion of or sequester the antibiotic, leading to increased bacterial tolerance (In summary, in response to meropenem, olerance .Click here for additional data file.


A patient presented with pancytopenia and hypercalcemia after intravesical immunotherapy with Bacillus Calmette-Guerin (BCG) for bladder cancer. Bone marrow biopsy performed six months later revealed noncaseating granulomas with negative stains for AFB. He was diagnosed with sarcoidosis and treated with prednisone. Hypercalcemia resolved, but mild pancytopenia persisted. One year later, he developed sepsis. Blood cultures six weeks later grew Mycobacterium tuberculosis complex, ultimately identified as Mycobacterium bovis. Despite triple antibiotic therapy, the patient progressively declined and expired. Granulomas are focal aggregates of macrophages, epitheliod cells, multinucleated giant cells, and lymphocytes that form in response to persistent inflammatory or infectious stimuli . Common One rare cause of granuloma is disseminated Bacillus Calmette-Guerin (BCG) infection. BCG is a vaccine developed from the bacterium Mycobacterium bovis (M. bovis). This organism was distinguished from Mycobacterium tuberculosis (MTB) by Robert Koch in early attempts to develop a vaccine against tuberculosis . CalmettA 69-year-old male presented to the hospital with recurrent episodes of hypercalcemia. He had a past medical history of head and neck squamous cell carcinoma status post resection and adjuvant cisplatin chemotherapy completed one year prior to admission and bladder cancer status post transurethral resection of bladder tumor followed by BCG intravesical immunotherapy completed six months prior to admission. The final BCG administration was complicated by extensive gross hematuria and a prolonged flu-like illness that lasted for weeks. Upon admission, he complained of nonspecific fatigue and weakness but no weight loss, nausea, vomiting, polyuria, abdominal pain, myalgia, or arthralgia. His temperature was 98.4, respiratory rate 18, blood pressure 163/80, pulse 66, and pulse oximetry 96% on room air. The physical exam was significant for a chronically ill-looking patient with mild pallor. He had no lymphadenopathy. His cardiovascular and respiratory system examinations were unremarkable. His abdominal examination revealed a soft, nontender abdomen with no hepatosplenomegaly, soft in consistency.Laboratory evaluation revealed pancytopenia with WBC 3.67 K/uL, hemoglobin (Hgb) 9.6 M/uL, and platelet 107 K/uL. His creatinine (Cr) was elevated at 1.94 mg/dL. Urinalysis didn't reveal proteinuria or hematuria. Calcium was elevated at 11.4 mg/dL. Parathyroid hormone (PTH) was appropriately low at 8.5 pg/dL, and parathyroid hormone-related protein (PTH-rP) was also low at 13 pg/mL. He had normal 25-OH Vit D with mildly elevated 1-25 vitamin D level at 77 pg/mL . He had a normal angiotensin-converting enzyme (ACE) level of 55 U/L (Table CT scan of the neck, chest, abdomen, and pelvis revealed mild hilar adenopathy but no recurrence of cancer. There was a small, calcified granuloma in the liver, which was not present on prior abdominal imaging. A skeletal survey revealed no destructive bone lesions. A bone marrow biopsy showed noncaseating granulomas without evidence of dysplasia or plasmacytoid process Figure . AFB staOne year later, the patient returned to the hospital with sepsis-like syndrome along with hypotension, fever, and persistent pancytopenia. Repeat bone marrow biopsy again showed non-caseating granulomas with negative AFB stain. Blood, urine, bone marrow, and sputum cultures were negative. Six weeks after his initial presentation, blood cultures for AFB became positive for Mycobacterium complex, subsequently confirmed to be M. bovis. Prednisone and mycophenolate given for sarcoid were discontinued, and he was started on isoniazid, ethambutol, rifabutin, and vitamin B6. Despite antibiotic therapy, the patient continued to deteriorate and expired one month later. The final diagnosis was systemic BCG infection 18 months after intravesical administration of BCG based on positive blood cultures for M. bovis and demonstration of granulomas in bone marrow and liver.BCG is a live attenuated strain of M. bovis. Intravesical administration is usually safe, however, local and systemic infections can occur . In the M. bovis-related symptoms may appear shortly after BCG administration or years later, making the diagnosis difficult. The finding of granulomas on a biopsy of involved organs is nonspecific, and AFB stains can be negative despite the presence of mycobacterium . The sloThe typical presentation of disseminated BCG infection includes fever, malaise, and pancytopenia, which occur in many other conditions. The finding of granulomas can easily point to sarcoidosis as a more likely diagnosis, as in our patient. Hypercalcemia has not previously been reported as a complication of M. bovis infection. We postulate that hypercalcemia was mediated by an increase in calcitriol as documented in other granulomatous diseases .In summary, although BCG is generally safe, disseminated M. bovis disease is a rare and serious adverse reaction that can occur following BCG vaccination. Symptoms of the infection can present one or more years after BCG administration. A high index of suspicion is required. The confirmation also becomes difficult due to M. bovis only growing in selective culture media under special laboratory conditions and at a very slow rate. A misdiagnosis with another granulomatous condition, such as Sarcoidosis, can occur given the similar clinical presentation with the presence of granulomas. A range of clinical presentations can occur. We add bone marrow involvement and hypercalcemia to the spectrum.


Bacillus subtilis variant natto after a gastrointestinal perforation in a patient in Japan. Genotypic and phenotypic studies of biotin identified B. subtilis var. natto. This case and 3 others in Japan may have been caused by consuming natto (fermented soybeans).We report a case of bacteremia caused by  Bacillus subtilis is a gram-positive, rod-shaped, spore-forming bacterium temporarily present in the human gastrointestinal tract  for a 2-day history of abdominal pain after having taken barium for gastric radiographic examination. The patient had a history of hypertension and ate natto (fermented soybeans) almost every day. At admission, the patient exhibited spontaneous abdominal pain, muscular defense, and rebound tenderness. Laboratory findings showed a decreased leukocyte count  and mildly increased C-reactive protein concentration . Contrast-enhanced computed tomography revealed contrast accumulation in the colon and free air around the sigmoid rectum. Lower gastrointestinal perforation and generalized peritonitis were suspected, and 2 sets of blood cultures were obtained. Emergency proctosigmoidectomy (Hartmann surgery) was performed on the same day, and perforation of the sigmoid colon was confirmed.B. subtilis was isolated from the culture by matrix-assisted laser desorption/ionization-time of flight mass spectrometry, and the antimicrobial drugs were changed to ampicillin/sulbactam (12 g/d) as indicated by antimicrobial susceptibility testing by broth microdilution . On day 5, because both blood culture sets were positive for gram-positive rod bacteria, teicoplanin (800 mg/d) was added. On day 11, only dilution  Table. BB. subtilis var. natto. DNA analysis showed that in the bioF region, the isolate was 100% homologous to the B. subtilis var. natto standard strain. Compared with the B. subtilis subspecies subtilis standard strain, the isolate had ≈50 fewer bases and the bioW region of the isolate had a single-nucleotide mutation that resulted in a termination codon for amino acid synthesis ; gram-positive bacilli rarely induce bacteremia (2.7%) (B. subtilis bacteremia typically originates from the gastrointestinal tract (B. subtilis bacteremia arising from the gastrointestinal tract (B. subtilis bacteremia caused by probiotics (B. subtilis bacteremia in Japan (We conclude that this case of bacteremia caused by B. subtilis var. natto resulted from gastrointestinal tract perforation. Genotypic and phenotypic studies on biotin effectively identified B. subtilis var. natto. In Japan, natto consumption is common, and B. subtilis bacteremia of gastrointestinal origin is most likely associated with B. subtilis var. natto.This case of bacteremia caused by Bacillus subtilis variant natto bacteremia of gastrointestinal origin, Japan.Supplemental results from study of patient with


Fusobacterium nucleatum (F. nucleatum) is an anaerobic oral commensal and the major coaggregation bridge organism linking early and late colonisers. In recent years, a large number of studies suggest that F. nucleatum is closely related to the development of various systemic diseases, such as cardiovascular diseases, adverse pregnancy outcomes, inflammatory bowel diseases, cancer, Alzheimer's disease, respiratory infection, rheumatoid arthritis, etc.F. nucleatum on systemic diseases and its possible pathogenesis and to open new avenues for prevention and treatment of F. nucleatum-associated systemic diseases.To review the effect of Fusobacterium nucleatum' OR 'Periodontal pathogen' OR 'Oral microbiota' OR 'Porphyromonas gingivalis' and was conducted in the major medical databases.The research included every article published up to July 2022 featuring the keywords 'Systemic diseases' OR 'Atherosclerotic cardiovascular diseases' OR 'Atherosclerosis' OR 'Adverse pregnancy outcomes' OR 'Inflammatory bowel disease' OR 'Ulcerative colitis' OR 'Crohn’s disease' OR 'Cancers' OR 'Oral squamous cell carcinomas' OR 'Gastrointestinal cancers' OR 'Colorectal cancer' OR 'Breast cancer' OR 'Genitourinary cancers' OR 'Alzheimer’s disease ' OR 'Rheumatoid arthritis' OR 'Respiratory diseases' AND 'F. nucleatum can induce immune response and inflammation in the body through direct or indirect pathways, and thus affect the occurrence and development of systemic diseases. Only by continuing to investigate the pathogenic lifestyles of F. nucleatum will we discover the divergent pathways that may be leveraged for diagnostic, preventive and therapeutic purposes. F. nucleatum, a Gram-negative anaerobe, is one of the most abundant species in the oral cavity for both diseased and healthy individuals [F. nucleatum has become a hot research topic because of its increasingly revealed associations with extraoral diseases. The surface adhesin FadA expressed by F. nucleatum could increase permeability and promote F. nucleatum penetration of endothelial cells by binding to vascular endothelial cadherin (VE-cadherin) [F. nucleatum-related researches, including our evolving understanding of its mechanistic role in promoting systemic diseases and the challenges of developing diagnostic and therapeutic methods for F. nucleatum-associated systemic diseases.Periodontal disease (PD), one of the most common inflammatory diseases in adults, comprises a wide range of inflammatory conditions that jeopardize the supporting structures of the teeth . PD is initiated by bacterial biofilm, which interacts with the host immune defense system, further aggravating the inflammatory response . Recentlividuals . It is aividuals ,11. Its ividuals . In the adherin) . This phadherin) . In thiset al. [et al. [Aggregatibacter actinomycetecomitans (A. actinomycetecomitans), Porphyromonas gingivalis , Tannerella forsythia (T. forsythia), Prevotella intermedia (P. intermedia), F. nucleatum, Campylobacter rectus, and Treponema denticola (T. denticola) [ACVDs, including coronary artery disease and stroke, are one of the most common causes of death in the elderly . Atheroset al. , as well [et al. ,21. Somenticola) , suggestnticola) ,23. Morenticola) .F. nucleatum is one of the oral bacteria detected in the atherosclerotic plaques [F. nucleatum in atherosclerotic plaques and blood vessels is directly related to the severity of PD [F. nucleatum was 34% [F. nucleatum alone did not promote AS [F. nucleatum would act synergistically with other organisms in the development of AS [ plaques ,25,26 anty of PD . In 31 c was 34% . Subsequomote AS , but in nt of AS ,30. Thernt of AS . Permeabnt of AS .F. nucleatum can promote the progression of AS by affecting endothelial cell permeability through various mechanisms. F. nucleatum possesses a best-characterized surface adhesion called FadA [F. nucleatum is often found in mixed infections at extra-oral sites. In addition, F. nucleatum challenge markedly impaired cell proliferation and apoptosis in endothelial cells, destroying the original equilibrium state and leading to endothelial damage [F. nucleatum infection was also observed to affect the expression of endothelial cell surface markers and modulate receptors for VEGF on endothelial cells, resulting in impaired tissue vascularization during inflammation [F. nucleatum further impairs vascular endothelial integrity by inhibiting endothelial cell proliferation, destroying endothelial cell structure and function, and inhibiting the vascularization of damaged tissues during inflammation, thus paving the way for the invasion of other bacteria [Firstly, led FadA . FadA haled FadA . Pre-Fadled FadA . This rel damage . F. nuclammation . On the bacteria .et al. proved that the heat-shock protein GroEL of F. nucleatum stimulated atherosclerotic risk factors through several mechanisms [F. nucleatum GroEL upregulated the expression of chemokines, such as interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1), and cell adhesion molecules, such as vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), and E-selectin. This is an important step in the pathogenesis of endothelial dysfunction. In addition, GroEL could increase procoagulant activity by upregulating tissue factor (TF) and downregulating TF pathway inhibitor (TFPI) in endothelial cells. This prothrombotic response may be associated with plaque progression and instability. GroEL also induced monocyte adhesion to and transmigration through endothelial cells in concert with increased uptake of lipids in atherosclerotic lesions, and promoted monocyte differentiation into pro-inflammatory macrophages and, eventually, foam cells.Secondly, many studies have reported that human heat shock proteins (hHSPs) are closely related to AS. Antibodies directed against bacterial GroEL cross-react with hHSP60 on endothelial cells may result in endothelial dysfunction and the subsequent development of AS . Lee et chanisms . F. nuclF. nucleatum as monoinfection alone does not promote the induction of AS, but may conversely inhibit plaque formation [F. nucleatum-infected mice developed minimal aortic plaque, which was significantly smaller than sham-infected mice, and fewer F4/80+ macrophages were detected in the intimal layer of 24-week-infected mice than 12-week-infected mice. Additionally, there was no increase in macrophage infiltration or T-cell infiltration into the inner and outer membrane layers in infected mice at 24 weeks, suggesting that chronic F. nucleatum infection reduces inflammation in the aorta, which may in part explain the minimal development of atherosclerotic plaques. Although all lipid fractions were statistically elevated in F. nucleatum-infected mice, serum NO was not altered. Consistent with the minimal plaque observed in 24-week-infected mice, the result demonstrated no vascular endothelial dysfunction in animal model.Thirdly, chronic oral infection with ormation . In thisF. nucleatum promoted the endothelial dysfunction and AS plaque progression by upregulating expression of different aortic Toll-like receptors (TLRs) and inflammasome signal transduction [F. nucleatum up-regulated pro-inflammatory factors such as IL-1α, IL-6 and TNF-α through activation of TLR-MyD88-NF -κB in endothelial cells, resulting in the upregulation of ICAM, VCAM and MCP-1 [Some scholars used the multi-bacterial infection models to show that the inclusion of sduction ,30. Subsnd MCP-1 .F. nucleatum may not be involved in AS alone, but rather act synergistically with other organisms, in particular by disrupting cell–cell junctions, breaking down endothelial integrity and leading to invasion [Due to the interdependence  of periodontal bacteria, no single periodontal bacterial species is effective in inducing aortic disease pathology, for which a multimicrobial consortium is responsible. In summary, more researches are needed to confirm these arguments that invasion .P. gingivalis and F. nucleatum in vitro can induce the expression of fatty acid-binding protein 4 (FABP4) mRNA and protein depending on the JNK/AP-1 pathway, causing an increase of lipid uptake and foam cell transformation in macrophages [P. gingivalis correlated with serum levels of FABP4 in humans, whereas no association occurred between F. nucleatum antibody titers and FABP4 levels, which might indicate that P. gingivalis is superior to F. nucleatum in inducing FABP4 in humans. Further study is needed to define the reason underlying the difference in cell-based models and human studies. The mechanisms between F. nucleatum and AS are summarized in Furthermore, we found that both rophages . Human sAPO is a broad term including preterm labor, chorioamnionitis, preterm premature rupture of membranes, preeclampsia, miscarriage, intrauterine growth retardation, low birth weight, stillbirth, neonatal sepsis, etc.F. nucleatum is one of the most prevalent species and by far the most prevalent oral species implicated in APOs and has been found to be significantly enriched in a wide variety of placental and fetal tissues including amniotic fluid, fetal membranes, cord blood, neonatal gastric aspirates, fetal lung and stomach, associated with preterm birth, preterm premature rupture of membranes, chorioamnionitis, early-onset neonatal sepsis, stillbirth and preeclampsia, either as the sole infectious agent or in mixed infections [fections .F. nucleatum identified in amniotic fluid and placenta appear to match those from the maternal or the partner subgingival sites, providing the human evidences that the bacteria originated from the mother’s subgingival plaque and translocated to the placenta and fetus, causing acute inflammation that eventually leads to APOs [F. nucleatum, demonstrating that the oral bacteria are capable of translocation to the fetal-placental unit [F. nucleatum in amniotic fluid and cord blood associated with preterm birth indicates its ability to spread to different placental and fetal compartments [The strains of  to APOs . Animal tal unit . Concurrartments .F. nucleatum translocates from the maternal oral cavity to the intrauterine cavity via hematogenous transmission [F. nucleatum was injected into the tail vein of pregnant mice to mimic dental bacteremia, which led to bacterial colonization in the decidua of the mouse placenta, followed by spread to the fetal membranes, mimicking chorioamnionitis in humans [It has been assumed that smission . This hysmission ,46. In an humans . A recenn humans .Periodontal pathogens can cause adverse pregnancy outcomes through two major mechanisms identified in the consensus report from the joint European Federation of Periodontology/American Academy of Periodontology workshop on periodontitis and systematic diseases: one is that periodontal pathogens directly enter the blood circulation system, or an ascending route via the genitourinary tract, thus invading the fetal-placenta unit; the other is that pro-inflammatory cytokines, such as IL-6, IL-8, and TNF-α, which are locally produced in periodontal tissues, enter the circulatory system, directly exerting negative influence on the fetal-placenta unit, or circulate to the liver and increase the systemic inflammatory state through the acute phase of protein reactions, which in turn affects the fetal placental unit ,49. In bF. nucleatum in the placenta of mice [F. nucleatum FadA to VE-cadherin not only increases the endothelial permeability, but also allows F. nucleatum and other oral bacteria to disseminate into and from the circulation. This may explain why F. nucleatum is frequently detected concurrently with other oral species in intrauterine infections in humans [In vivo, an F. nucleatum mutant without FadA is significantly defective in placental colonization, while its complement clone restores the colonization [Several adhesin molecules, including Fap2 and FadA, have been identified for colonization of the  of mice ,51–53. A of mice . Bindingn humans . In vivonization ,54.F. nucleatum in the mouse placenta led to preterm and/or term fetal death, occurring 2–3 days following bacterial injection, accompanied by placental neutrophil infiltration, similar to that observed in humans [F. nucleatum activates both TLR2 and TLR4 in vitro and in murine placentas, it induces inflammatory responses via TLR4, accompanied by neutrophil infiltration into the decidua [F. nucleatum colonizes the placenta to a similar extent as in untreated wild-type mice without eliciting inflammatory responses, resulting in a reduced fetal death rate, which suggests that inflammation rather than the bacteria per se is the cause of fetal demise. In contrast, the effect of TLR2 is insignificant because there is no change in fetal loss or inflammatory response in TLR2-knockout mice compared to wild-type mice.More studies reported that colonization of n humans . Althoug decidua . In miceF. nucleatum-induced production of IL-6 by activating NF-κB signaling in murine macrophages and human decidual stromal cells (hDSCs) [F. nucleatum infection results in APOs by inducing aberrant production of cytokines and chemokines through NOD1/NOD2-Ripk2-mediated signaling. In conclusion, although it would be beneficial to regulate Ripk2 signaling to prevent APOs caused by bacterial infections, further studies using animal models are needed to elucidate whether Ripk2 is involved in F. nucleatum-induced APOs and Ripk2 inhibitors are beneficial to prevent the occurrence of APOs.Scholars have confirmed that inflammatory biomarkers in the maternal serum may include IL-6,C-X-C motif chemokine 8 (CXCL8) and CC chemokine ligand 2 (CCL2) . Recepto (hDSCs) . Ripk2 aIBD, a heterogeneous set of inflammatory disorders of the gastrointestinal (GI) tract and a global public health issue of increasing importance, presents as two major clinical phenotypes: ulcerative colitis (UC) and Crohn’s disease (CD). UC is a continuous inflammation of the colonic mucosa and submucosa, usually involving the rectum initially, then gradually spreading to the whole colon, whereas CD is usually transmural and can affect any area of the gastrointestinal tract, which is a discontinuous full-layer inflammation, most commonly involving the terminal ileum, colon, and perianal . The etiet al. have reported that F. nucleatum isolated from inflamed biopsy tissue from IBD patients is significantly more common and invasive than from healthy controls [F. nucleatum is obviously enriched in feces of IBD patients and its abundance has a positive correlation with patients’ disease activity, and administration of F. nucleatum markedly exacerbates colitis in DSS mice model [Back in 2011, Strauss controls . It has ce model . This coce model . In addice model . Recent ce model .et al. discussed that the mode of the relocation of oral bacteria from the oral cavity to the gut mucosa may include hematogenous route, enteral route and other possible factors [Kitamoto  factors . First,  factors ,77. In a factors . Second, factors ,78. Mean factors ,79,80. T factors ,82. Yet F. nucleatum in IBD. Firstly, F. nucleatum could damage the epithelial barrier integrity and increase permeability by downregulating the expression of the tight junction proteins zonula occludens-1 (ZO-1) and occludin, which are markers of intestinal mucosal barrier function [F. nucleatum-mediated mucosal barrier damage could be promoted by targeting caspase activation and recruitment domain 3 (CARD3) which in term activated the endoplasmic reticulum stress (ERS) pathway [F. nucleatum could regulate M1 macrophage skewing to induce colitis [et al. also reported that F. nucleatum could exacerbate intestinal inflammation through upregulating cytokine secretion, such as IL-1β, IL-6, and IL-17, activating STAT3 signaling pathway, enhancing proliferation of CD4+ T cell and differentiation to Th1 and Th17 [et al. found that the presence of F. nucleatum and FadA gene increased in UC patients, especially in patients with severe colitis and pancolitis, suggesting that FadA may play a significant role in the pathogenesis of UC [F. nucleatum and FadA gene in UC.In this review, we only discuss the possible pathogenesis of function . Additio pathway . Secondl colitis . Liu et and Th17 . Li et ais of UC . Neverth−/− mice model, Yan et al. reported that non-surgical periodontal treatment triggered modulation of gut microbiota, facilitating recovery to a healthy microbiome situation [F. nucleatum and IBD, we reasonably assume that F. nucleatum abundance maybe indicative of development of IBD. Moreover, targeting F. nucleatum may be effective to shorten disease course and prevent the development of IBD. Since there is no explicit cure for IBD, understanding the intrinsic mechanism of F. nucleatum may provide a new insight in optimizing current therapeutic strategies.Using ApoEituation . It alsoF. nucleatum in intestinal tract may be the common pathogenesis of these two diseases [F. nucleatum and CRC will be discussed in detail later.In addition, IBD has been recognized as a risk factor for colorectal cancer (CRC), the infection of diseases . The conet al. firstly showed that F. nucleatum is associated with OSCC from an epidemiological perspective [Fusobacteria, Firmicutes, Actinobacteria and Proteobacteria. Most notably, a significant reduction in the abundance of Streptococcus species and an increase in the abundance of Fusobacterium species were observed in both primary and metastatic samples. Resphera Insight applied to saliva samples from HNSCC patients and healthy controls led to the first discovery that F. nucleatum enriched in a subset of saliva samples from HNSCC patients, when compared with controls [Porphyromonas and Fusobacterium species in OSCC tissue samples compared to samples from healthy mucosa [F. nucleatum is closely associated with the development of oral cancer by several mechanisms.OSCC is a malignant tumor occurring in the oral epithelium, which is the main type of head and neck squamous cell carcinoma (HNSCC). Al-Hebshi,spective . Subsequspective . The miccontrols . Additiocontrols . Nagy ety mucosa . F. nuclF. nucleatum invades gingival epithelial cells, NF-κB and NOD-like receptor 3 (NLRP3) are simultaneously activated [et al. further verified that F. nucleatum could promote IL-1β by increasing AIM2 and downregulating POP1 in HNSCC in vitro [P. gingivalis and adenosine triphosphate (ATP) with or without F. nucleatum upregulated NLRP3, IL-1β by downregulating POP1. Moreover, P. gingivalis and F. nucleatum can initiate the overexpressed NLRP3, activate upstream signal molecules of ataxia-telangiectasia and Rad3 related (ATR)-checkpoint kinase 1 (CHK1), promoting the growth and proliferation of oral cancers [Firstly, when ctivated . NF-κB wctivated . Furtherctivated . The infctivated ,95. Aralin vitro . P. ging cancers .F. nucleatum induces activation of protein kinase p38 in infected cells, promoting the secretion of matrix metalloproteinase-13 (MMP-13) and MMP-9, which contributes to tumor invasiveness [Secondly, siveness .F. nucleatum-infected OSCC cells due to DNA damage [F. nucleatum and Ku70 is yet to be explained.Thirdly, DNA damage plays a significant role in the development and progression of oral cancer. It has been reported that the Ku70/p53 signaling pathway may be involved in the excessive proliferation of A damage . If Ku70A damage ,101. NevF. nucleatum-infected cells, which leads to cell-cycle arrest in the S phase and to increased cell proliferation [In addition, the levels of tumor suppressor protein p27, a member of the cyclin-dependent kinase inhibitor (CDK) family, would be downregulated in feration . This prF. nucleatum was found to contribute to the induction of epithelial–mesenchymal transition (EMT) through lncRNA MIR4435-2HG/miR-296-5p/Akt2/SNAI1 pathway [Moreover, infection of oral epithelial cells with  pathway . EMT is  pathway .GI cancers are in the forefront of all malignant tumor in morbidity and mortality, including esophageal, gastric, colorectal, liver, and pancreatic cancer. A case-control study examined the salivary microbiota in patients with GI cancers and evaluated their differential distribution based on the cancer sites, suggesting the bacterial diversity and composition of saliva are related to GI cancers .F. nucleatum can not only promote the progression of GI tumor, but also contribute to the chemo-resistance of GI cancer [F. nucleatum-related GI cancers.It was reported that I cancer . We haveFusobacterium species in human CRCs and adenomas compared with adjacent normal tissue [F. nucleatum, suggesting that F. nucleatum is involved in the development of CRC. Castellarin et al. verified overabundance of F. nucleatum in colorectal tumor specimens versus matched normal control tissue by quantitative PCR analysis and observed that F. nucleatum was positively associated with lymph node metastasis [F. nucleatum can be used as a risk biomarker of CRC [F. nucleatum correlated with the genetic and epigenetic aberrancies in CRC, such as the CpG island methylator phenotype (CIMP), microsatellite instability (MSI), and MLH1 methylation [CRC is one of the most common malignant tumors. Its morbidity and mortality rank among the top three cancers in the world, and are increasing in recent years . Recent l tissue ,107–110,tastasis . Subsequr of CRC . Increashylation , 116–118F. nucleatum can alter the composition of the residual microbiota, with a consortium of inflammatory responses, virulence factors and impaired epithelial signaling in the context of a polymicrobial oral biofilm with synergistic properties, resulting in intestinal dysbiosis, contributing to the initiation and progression of CRC [n of CRC .F. nucleatum and CRC oncogenesis, two pathways must be mentioned. One of the pathways is the virulence factor FadA of F. nucleatum which would activate the β-catenin signaling pathway when present on the VE-cadherin, initiating inflammatory responses, thus boosting the genes of the transcription factors NF-κB, pro-inflammatory cytokines such as IL-6, IL-8, and IL-18, Wnt7a, Wnt7b, Wnt9a, transcription factors lymphoid enhancer factor (LEF) /T cell factor (TCF), Myc, and Cyclin D1, and the parts of the Wnt pathway [F. nucleatum also can activate β-catenin signaling through TLR4/P-PAK1 cascade [F. nucleatum mediated its enrichment in CRC by binding to tumor-overexpressed Gal-GalNAc, further intensifying the inflammatory response [F. nucleatum lipopolysaccharide (LPS) could activate TLR4/MyD88/NF-κB, inducing high expression of miR21; the elevated miR21 could reduce levels of RASA1, while the RASA1 could active MAPK pathway [F. nucleatum was found to inhibit the expression of miR-18* and miR-4802 through TLR4/MyD88 immune signals [F. nucleatum invades into CRC cells, reactive oxygen species (ROS) could be induced, which subsequently lead to DNA damage [In respect to association between  pathway . F. nucl cascade . In pathresponse . In path signals . The losA damage ,125.F. nucleatum modulates tumor immune microenvironment in CRC? On the one hand, F. nucleatum could selectively recruit tumor-infiltrating myeloid cells and promotes tumor progression [F. nucleatum could interact with TIGIT, an immunoregulatory signaling receptor on NK and T cells, and this Fap2-TIGIT interaction could reduce killing of tumor cells by NK and tumor-infiltrating lymphocytes and inducing lymphocyte apoptosis [The above findings bring us to the next question: how gression . On the poptosis .F. nucleatum and its relationship with CRC, studies on F. nucleatum are increasing gradually, but our understanding of F. nucleatum is still insufficient at present. Further researches are needed on how F. nucleatum can be a therapeutic target to reduce the risk of CRC. The researchers found that detection of fecal abundance of F. nucleatum can be used as a rapid and non-invasive diagnostic technique for CRC screening [F. nucleatum also have potential diagnostic value [F. nucleatum-associated CRC, but the risk of CRC did not seem to change in patients lacking F. nucleatum [F. nucleatum [F. nucleatum and only targeted at tumor tissue.Due to the biological characteristics of creening . Combineic value . Scholarucleatum . Moreoveucleatum . Howeveret al. reported that F. nucleatum enrichment was associated with a worse prognosis in esophageal cancer by activating chemokines, such as CCL20 [et al., the detection rate of Fusobacterium spp. in pancreatic cancer tissue specimens was 8.8% [F. nucleatum was not detected in pancreatic cancer tissue in another study by Yamamura et al. [et al. found that several bacterial groups, including Lactococcus, Veilonella, Fusobacterium, and Leptotrichia species were enriched in gastric cancer [et al. implied that Clostridium colicanis and F. nucleatum could be used as diagnosis biomarkers of gastric cancer [et al. found that Gal-GalNAC antigen was highly expressed in gastric cancer and esophageal cancer tissues, suggesting that gastric cancer and esophageal cancer could enrich F. nucleatum chemotaxis and affect disease progression [Yamamura as CCL20 . In a stwas 8.8% , while Fa et al. . Castan˜c cancer . Heish ec cancer . Abed etgression .F. nucleatum has recently been detected in human BC tissues and was shown to promote BC progression in a murine model [et al. reported that F. nucleatum colonizes not only CRC, but also BC through recognition of Gal-GalNAc by Fap2 [F. nucleatum-induced tumor exacerbation. Van der Merwe et al. have suggested that F. nucleatum may promote CRC and BC progression by activating the TLR4/ MyD88 pathway, and F. nucleatum also exhibits immunomodulatory effects [F. nucleatum to tumours [F. nucleatum [BC is the leading cause of cancer mortality in women and a type of cancer with different presentations among women . F. nuclne model ,141. Par by Fap2 . Also, m effects . These o tumours ,140; autucleatum ,143; PD-ucleatum .et al. have summarized epidemiological studies exploring the association between PD and GU cancers, and indicated that the presence of an oral-genitourinary axis and oral microbiota may be involved in the pathogenesis of GU cancers [et al. are the first to report that there exists possible association of Fusobacterium spp. with urothelial carcinomas, at least in some bladder cancer patients [F. nucleatum is indeed present in approximately one quarter of the tested samples by PCR-based analysis, and indicated the bacteria may play a key role in the exacerbation of the bladder cancer based on the 16S rDNA gene sequence approach. Nevertheless, more epidemiological studies and animal experiments are needed to elucidate the exact role of F. nucleatum in bladder cancer formation and progression. Alluri et al. have identified the periodontal pathogen F. nucleatum in prostate glands diagnosed with adenocarcinoma, but they have no evidence of whether the F. nucleatum found in the prostatic tissue of the individual originated from the oral cavity [F. nucleatum in pathological prostate changes is unclear yet. Huang et al. found that there was a distinct observation of higher levels of F. nucleatum in cervical cancer, especially for recurrent tissues [F. nucleatum intratumoral infiltration exhibited correspondingly poor rates of both overall survival and progression-free survival, suggesting that F. nucleatum might be one potential cervical cancer diagnostic and prognostic biomarker.Shuai Yuan  cancers ,148. Bučl cavity . Meanwhi tissues . PatientAD is the most common form of dementia in older adults and refers to a central nervous system disease characterized by progressive cognitive dysfunction and memory loss . In the Several putative mechanisms that could explain how periodontitis affects the central nervous system (CNS) homeostasis have been described , includiWith the increase of age, the permeability of the BBB increases in elderly patients. Therefore, pathogenic microorganisms could easily cross the BBB and enter the brain tissue, directly act on neurons, activate the inflammatory cascade reaction, and cause direct damage to the CNS. The bacteria entering brain tissue and their secreted virulence factors could activate a classic immune response similar in some aspects to that observed in AD, through TLR2 and TLR4 pathway, and turn inactive microglia into active ones ,158 WhenF. nucleatum can be detected in the serum of patients with AD or cognitive impairment [F. nucleatum were significantly more abundant in the AD group than controls [in vitro and animal experiments were conducted to preliminarily explore the pathogenesis of AD exacerbated by F. nucleatum [F. nucleatum LPS promoted the proliferation of microglia, which promoted the enhancement of inflammatory immune function, leading to changes in cell morphology and increased expression of inflammatory genes. F. nucleatum LPS could increase permeability of the BBB, and then affect the onset and development of AD. Previous studies have shown that local inflammation in the central nervous system could lead to cognitive impairment [F. nucleatum. In addition, in the presence of F. nucleatum infection, the expression of P38 protein in mouse brain tissue was significantly upregulated, as was phosphorylated P38 protein. Likewise, MyD88 protein was upregulated. F. nucleatum also stimulated the JNK pathway; the expression levels of phosphorylated JNK and JNK proteins were raised. Moreover, quantitative proteomics analyses were performed to detect the proteins in the brain tissues of 5XFAD mice with or without F. nucleatum infection [F. nucleatum affect the progression of AD remains unkonwn. On the whole, more efforts are required to elucidate the mechanism of F. nucleatum actions in AD in future studies.Earlier studies have reported that antibodies to pairment . Scholarcontrols ,166. Recucleatum . Scholarpairment . This renfection . The resnfection . FurtherF. nucleatum is associated with brain, lung, liver, and splenic abscesses [F. nucleatum was involved in rheumatoid arthritis (RA) and acute appendicitis [et al. firstly reported that a triple oral inoculation of pathobionts  combined with collagen could induce arthritis in the mouse, and oral inoculation with either F. nucleatum or A. actinomycetemcomintans alone could accelerate subsequent arthritis onset and progression [P. gingivalis, F. nucleatum and A. actinomycetecomitans inhaled into respiratory tract could promote the invasion of Pseudomonas aeruginosa into respiratory epithelial cells and induce cytokine production and cell apoptosis [F. nucleatum is one of the most common causative agents of Lemierre’s syndrome, a rare form of upper airways infection with a life-threatening secondary septic thrombophlebitis of internal or external jugular veins [Han et al. have suggested that bscesses . Recent ndicitis ,169–171.poptosis . F. nuclar veins .F. nucleatum has been considered as an opportunistic pathogen that interacts with other microorganisms and plays a crucial role in many infectious diseases. With the development of omics technology, the occurrence and development of many diseases are closely related to the infection of F. nucleatum. The latest trend has been to focus on exploring possible direct and indirect links between F. nucleatum and certain systemic diseases, especially ACVDs and CRC, and an increasing amount of evidence has been accumulated. F. nucleatum not only promotes inflammation, but also binds to or invades multiple cell types, including oral, colon and placental epithelial cells, T cells, keratinocytes, and macrophages [F. nucleatum can invade the human body and influence general health lies in several key virulence mechanisms which can be broadly classified into three groups  Cologenesis . In addive tract . (ii) Incascades ,48–50. Cent TLRs ,121,141,ent TLRs , and enhand Th17 . (iii) Sdiseases ,114,140.F. nucleatum on multiple systemic diseases, but also discuss the impact of targeting F. nucleatum on the treatment of related systemic diseases. However, many of the epidemiological studies only show the correlation but not necessarily causation between F. nucleatum and specific diseases. Precise pathogenicity of F. nucleatum-related systemic diseases has yet to be uncovered, and a few conclusions are controversial. Only by continuing to investigate mechanisms involved in transformation of this oral commensal organism into systemic pathogens will we discover possible strategies for diagnostic, preventive and therapeutic purposes. Based on current research, some recommendations can be given. Firstly, it must be emphasized that oral health should be maintained as an indispensable part of a healthy lifestyle to reduce risk of bacteremia, especially for immune-compromised patients. Secondly, by investigating into the epidemiological background of F. nucleatum infection, high-risk groups for various related diseases can be effectively revealed, facilitating early detection and prevention among the population. Thirdly, it is of great significance for F. nucleatum-associated cancer patients with poor prognosis or chemotherapy-resistance to apply multi-drug combination therapy against F. nucleatum and employ drug administration methods similar to Helicobacter pylori triple therapy.In this article, we not only summarize the adverse effects of