Arata, Yukinobu et al. (2021) International Worm Meeting "Insulin signaling underlies a heavy-tailed temporal organization in C. elegans episodic swimming"

Jurica, Peter, Kimura, Hiroshi, Arata, Yukinobu, Ikeda, Yusaku, Kiyono, Ken, Sako, Yasushi, Shiga, Itsuki

[International Worm Meeting, 2021]

Episodic bouts of animal behaviors are not periodic, but temporally organized in a scale-free manner with long memory in many animal species including human. It is known that depression in human is associated with defects in the scale-free temporal organization of behaviors and impacts on social fitness, and accordingly exacerbates disease state. C. elegans crawls in on agar plate and swim in a solution while switching between actively-moving state and inactive state, which is referred to as episodic behavior. We have shown that the residence time in each state in episodic swimming has a scale-free or heavy-tailed property with long memory. Here, we studied how the residence times between two states are cross-correlated by using detrending moving-average cross-correlation analysis (DMCA) to activity time series obtained from a long-term culture of C. elegans. We found that residence times in the active and inactive states are correlated with each other over the time scale from minutes to hours, or at the ultradian time scale. This temporal cross-correlation disappeared when cultured in M9 buffer without food bacteria, and restored when cultured M9 buffer only with glucose. Additionally, the temporal cross-correlation disappeared in the mutant animals of daf-2 gene, which encodes the membrane receptor for insulin growth factor signal and in the mutant of daf-16, which encodes a downstream transcriptional repressor FOXO. Therefore, we conclude that the temporal cross-correlation is achieved by the insulin signaling. Glucose metabolism disorders and behavioral disorders are known to be closely related in human depression, raising the possibility that the temporal organization by insulin signaling found in C. elegans is conserved in humans.

Wang ST et al. (2021) Mol Microbiol "OmpR coordinates the expression of virulence factors of enterohemorrhagic Escherichia coli in the alimentary tract of Caenorhabditis elegans."

Wang ST, Kuo CJ, Chen JW, Chen CS, Lee TM, Huang CW

[Mol Microbiol, 2021]

Enterohemorrhagic Escherichia coli (EHEC), an enteropathogen that colonizes in the intestine, causes severe diarrhea and hemorrhagic colitis in humans by the expression of the type III secretion system (T3SS) and Shiga-like toxins (Stxs). However, how EHEC can sense and respond to the changes in the alimentary tract and coordinate the expression of these virulence genes remains elusive. The T3SS-related genes are known to be regulated by the locus of enterocyte effacement (LEE)-encoded regulators, such as Ler, as well as non-LEE-encoded regulators in response to different environmental cues. Herein, we report that OmpR, which participates in the adaptation of E. coli to osmolarity and pH alterations, is required for EHEC infection in Caenorhabditis elegans. OmpR protein was able to directly bind to the promoters of ler and stx1 (Shiga-like toxin 1) and regulate the expression of T3SS and Stx1, respectively, at the transcriptional level. Moreover, we demonstrated that the expression of ler in EHEC is in response to the intestinal environment and is regulated by OmpR in C. elegans. Taken together, we reveal that OmpR is an important regulator of EHEC which coordinates the expression of virulence factors during gastrointestinal infection in vivo.

Lee JH et al. (2014) Biofouling "Thermoresponsive oligomers reduce Escherichia coli O157:H7 biofouling and virulence."

Kim YG, Kim SC, Lee J, Cho MH, Cho HS, Kim J, Lee JH

[Biofouling, 2014]

Thermoresponsive polymers have potential biomedical applications for drug delivery and tissue engineering. Here, two thermoresponsive oligomers were synthesized, viz. oligo(N-isopropylacrylamide) (ONIPAM) and oligo(N-vinylcaprolactam) (OVCL), and their anti-biofouling abilities investigated against enterohemorrhagic E. coli O157:H7, which produces Shiga-like toxins and forms biofilms. Biofilm formation (biofouling) is closely related to E. coli O157:H7 infection and constitutes a major mechanism of antimicrobial resistance. The synthetic OVCL (MW 679) and three commercial OVCLs (up to MW 54,000) at 30 g ml(-1) were found to inhibit biofouling by E. coli O157:H7 at 37 C by more than 80% without adversely affecting bacterial growth. The anti-biofouling activity of ONIPAM was weaker than that of OVCL. However, at 25 C, ONIPAM and OVCL did not affect E. coli O157:H7 biofouling. Transcriptional analysis showed that OVCL temperature-dependently downregulated curli genes in E. coli O157:H7, and this finding was in line with observed reductions in fimbriae production and biofouling. In addition, OVCL downregulated the Shiga-like toxin genes stx1 and stx2 in E. coli O157:H7 and attenuated its in vivo virulence in the nematode Caenorhabditis elegans. These results suggest that OVCL has potential use in antivirulence strategies against persistent E. coli O157:H7 infection.

Chou TC et al. (2013) Cell Microbiol "Enterohaemorrhagic Escherichia coli O157:H7 Shiga-like toxin 1 is required for full pathogenicity and activation of the p38 mitogen-activated protein kinase pathway in Caenorhabditis elegans."

Chiu WT, Chen CS, Wu CM, Kuo CJ, Syu WJ, Chiu HC, Chou TC

[Cell Microbiol, 2013]

Enterohaemorrhagic Escherichia coli (EHEC) causes life-threatening infections in humans as a consequence of the production of Shiga-like toxins. Lack of a good animal model system currently hinders in vivo study of EHEC virulence by systematic genetic methods. Here we applied the genetically tractable animal, Caenorhabditis elegans, as a surrogate host to study the virulence of EHEC as well as the host immunity to this human pathogen. Our results show that E.coli O157:H7, a serotype of EHEC, infects and kills C.elegans. Bacterial colonization and induction of the characteristic attaching and effacing (A/E) lesions in the intact intestinal epithelium of C.elegans by E.coli O157:H7 were concomitantly demonstrated in vivo. Genetic analysis indicated that the Shiga-like toxin 1 (Stx1) of E.coli O157:H7 is a virulence factor in C.elegans and is required for full toxicity. Moreover, the C.elegans p38 mitogen-activated protein kinase (MAPK) pathway, an evolutionarily conserved innate immune and stress response signalling pathway, is activated in the regulation of host susceptibility to EHEC infection in a Stx1-dependent manner. Our results validate the EHEC-C.elegans interaction as suitable for future comprehensive genetic screens for both novel bacterial and host factors involved in the pathogenesis of EHEC infection.

Hwang SB et al. (2018) PLoS One "In vivo screening platform for shiga toxin-producing Escherichia coli (STEC) using Caenorhabditis elegans as a model."

Hwang SB, Park BJ, Choi JG, Wei S, Oh DH, Chelliah R

[PLoS One, 2018]

Shiga toxin-producing Escherichia coli (STEC) strains are the main cause of bacillary dysentery, although STEC strains generally induce milder disease symptoms compared to Shigella species. This study aimed to determine the virulence of STEC using the nematode Caenorhabditis elegans as a model host. Worm killing, fertility and bacterial colonisation assays were performed to examine the potential difference in the virulence of STEC strains compared to that of the control E. coli OP50 strains on which worms were fed. A statistically significant difference in the survival rates of C. elegans was observed in that the STEC strains caused death in 8-10 days and the E. coli OP50 strains caused death in 15 days. STEC strains severely reduced the fertility of the worms. The intestinal load of bacteria in the adult stage nematodes harbouring the E. coli OP50 strains was found to be 3.5 log CFU mL-1. In contrast, the STEC strains E15, E18 and E22 harboured 4.1, 4.2 and 4.7 log CFU ml-1 per nematode, respectively. The heat-killed STEC strains significantly increased the longevity of the worms compared to the non-heated STEC strains. In addition, PCR-based genomic profiling of shiga toxin genes, viz., stx1 and stx2, identified in selected STEC strains revealed that these toxins may be associated with the virulence of the STEC strains. This study demonstrated that C. elegans is an effective model to examine and compare the pathogenicity and virulence variation of STEC strains to that of E. coli OP50 strains.

Fujisawa K et al. (2007) Science "The slit receptor EVA-1 coactivates a SAX-3/Robo mediated guidance signal in C. elegans."

Wrana JL, Fujisawa K, Culotti JG

[Science, 2007]

The SAX-3/roundabout (Robo) receptor has Shiga-like toxin 1 (SLT-1)/Slit-dependent and -independent functions in guiding cell and axon migrations. We identified enhancer of ventral-axon guidance defects of unc-40 mutants (EVA-1) as a Caenorhabditis elegans transmembrane receptor for SLT-1. EVA-1 has two predicted galactose-binding ectodomains, acts cell-autonomously for SLT-1/Slit-dependent axon migration functions of SAX-3/Robo, binds to SLT-1 and SAX-3, colocalizes with SAX-3 on cells, and provides cell specificity to the activation of SAX-3 signaling by SLT-1. Double mutants of eva-1 or slt-1 with sax-3 mutations suggest that SAX-3 can (when slt-1 or eva-1 function is reduced) inhibit a parallel-acting guidance mechanism, which involves UNC-40/deleted in colorectal cancer.

Kuo, Cheng-Ju et al. (2013) International Worm Meeting "Shiga-like toxin 1 confers the full pathogenicity of Enterohaemorrhagic Escherichia coli and activation of the p38/MAPK pathway in Caenorhabditis elegans."

Chiu, Wen-Tai, Chen, Chang-Shi, Chiu, Hao-Chieh, Kuo, Cheng-Ju, Syu, Wan-Jr, Chou, Ting-Chen

[International Worm Meeting, 2013]

Enterohaemorrhagic Escherichia coli (EHEC), a major foodborne pathogen, is responsible for life-threatening diseases in humans as a consequence of the production of Shiga-like toxins. Currently, there is no specific treatment for EHEC infection and the use of antibiotics is contraindicated. Moreover, lack of a good animal model system hinders the study of EHEC virulence with systematic methods in vivo. To these regards, we applied the genetic tractable animal model, Caenorhabditis elegans, as a surrogate host to study the virulence of EHEC as well as the host innate immunity to this human pathogen. Our results show that E. coli O157:H7, a serotype of EHEC, infects and kills C. elegans. We also demonstrate that E. coli O157:H7 colonizes and replicates in C. elegans, stimulates the ectopic expression of microvillar actin, and induces the characteristic attaching and effacing (A/E) lesions in the intact intestinal epithelium in vivo. Moreover, our genetic analyses corroborate that the Shiga-like toxin 1 (Stx1) of E. coli O157:H7 is required partly for its toxicity. The C. elegans p38 MAP kinase signaling pathways, an evolutionally conserved innate immune signaling pathways, are activated and mediated in the regulation of host susceptibility to EHEC infection in a Stx1-dependent manner. Given that the bacterivore C. elegans and bacterium, in this case E. coli, may have generated a diverse arsenal for both parties to defend each other during the long-term evolution of this predator-prey relationship, our results suggest that this EHEC-C. elegans model is suitable for future comprehensive genetic screens for both novel bacterial and host factors involved in the pathogenesis of EHEC infection.

Lee Y et al. (2008) FEMS Microbiol Lett "The role of disulfide bond isomerase A (DsbA) of Escherichia coli O157:H7 in biofilm formation and virulence."

Yeom S, Kim S, Lee Y, Kim Y, Park W, Park S, Jeon CO

[FEMS Microbiol Lett, 2008]

The role of periplasmic disulfide oxidoreductase DsbA in Shiga toxin-producing Escherichia coli O157:H7 (STEC) was investigated. Deletion of dsbA (DeltadsbA) significantly decreased cell motility and alkaline phosphatase activity in STEC. STEC DeltadsbA also showed greater sensitivity to menadione and under low pH conditions. Significant reductions in surface attachment to both biotic (HT-29 epithelial cells) and abiotic (polystyrene and polyvinyl chloride) surfaces were observed in STEC DeltadsbA. In addition, no biofilm formation was detected in STEC DeltadsbA compared to wild-type cells in glass capillary tubes under continuous flow-culture system conditions. In the nematode model Caenorhabditis elegans-killing assay, the deletion of dsbA in STEC resulted in attenuated virulence compared to wild-type cells. STEC DeltadsbA was also found to have a reduced ability to colonize the nematode gut. These results suggest that DsbA plays important roles in biofilm formation and virulence in STEC cells.

Lee JH et al. (2014) Phytomedicine "Coumarins reduce biofilm formation and the virulence of Escherichia coli O157:H7."

Cho HS, Lee JH, Kim YG, Ryu SY, Lee J, Cho MH

[Phytomedicine, 2014]

E. coli O157:H7 is the most common cause of hemorrhagic colitis, and no effective therapy exists for E. coli O157:H7 infection. Biofilm formation is closely related to E. coli O157:H7 infection and constitutes a mechanism of antimicrobial resistance. Hence, the antibiofilm or antivirulence approach provides an alternative to antibiotic strategies. Coumarin and its derivatives have a broad range of biological effects, and in this study, the antibiofilm activities of nine coumarins were investigated against E. coli O157:H7. Coumarin or umbelliferone at 50g/ml was found to inhibit biofilm E. coli O157:H7 formation by more than 80% without affecting bacterial growth. Transcriptional analysis showed that coumarins repressed curli genes and motility genes in E. coli O157:H7, and these findings were in-line with observed reductions in fimbriae production, swarming motility, and biofilm formation. In addition, esculetin repressed Shiga-like toxin gene stx2 in E. coli O157:H7 and attenuated its virulence in vivo in the nematode Caenorhabditis elegans. These findings show that coumarins have potential use in antivirulence strategies against persistent E. coli O157:H7 infection.

Kim Y et al. (2006) Lett Appl Microbiol "Inhibitory effects of Lactobacillus acidophilus lysates on the cytotoxic activity of shiga-like toxin 2 produced from Escherichia coli O157:H7."

Park S, Han KS, Kim SH, Oh S, Imm JY, You S, Kim Y

[Lett Appl Microbiol, 2006]

Aims: The purpose of this study was to characterize the degree to which four cell lysates obtained from Lactobacillus acidophilus strains affected the cytotoxic activity of Escherichia coli O157:H7 in vitro and in vivo. Methods and Results: In a cytotoxic inhibition test that used 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and toxin-binding ELISA assays, the activity of shiga-like toxin 2 (Stx-2) was inhibited profoundly by the cell lysates (10 mg ml(-1)) from two strains of L. acidophilus A4 and 30SC (>85% of survival rates compared with the control) among the five strains tested. In particular, a significant decline in the virulence level of E. coli O157:H7, under the presence of the cell lysates of L. acidophilus A4, was observed by killing assay of Caenorhabditis elegans in vivo model. Conclusions: According to our results, L. acidophilus strains might be capable of attenuating the virulence of Stx-2 produced from E. coli O157:H7. Significance and Impact of the Study: The cell lysates of L. acidophilus can be applied to a variety of foods, and can be used as adjuncts for the inhibition of Stx-2-mediated cytotoxicity.