Enterohemorrhagic Escherichia coli (EHEC) strains adhere to the intestinal mucosa and produce an attaching and effacing (A/E) lesion. Most of the genes required to produce A/E lesions are thought to be encoded by the 36-kb pathogenicity island termed the locus for enterocyte effacement (LEE). Although the mechanisms underlying the bacterial adherence, including the genes involved, are still poorly understood, the preferential adherence phenotype of EHEC is thought to depend on the nature of the genes and/or the response of these genes to changes in environmental conditions. To explore the environmental factors affecting EHEC adherence, we used an O157:H7 strain and investigated the optimal growth conditions for its adherence to Caco-2 cells. We observed that EHEC grown in Dulbecco's modified Eagle's medium (DMEM) adhered more efficiently to Caco-2 cells than EHEC grown in Luria-Bertani (LB) broth. Among the components of DMEM, only NaHCO 3 was found to remarkably stimulate bacterial adherence. When bacteria were grown in LB broth containing NaHCO 3 , the production of intimin, Tir, EspA, and EspB was greatly enhanced compared with the production in LB broth. Indeed, the transcription of ler required for LEE-encoded gene expression was promoted in response to the concentration of NaHCO 3 in LB broth. Since the concentration of NaHCO 3 in the lower intestinal tract has been shown to be relatively high compared with that in the upper small intestine, our results may imply that NaHCO 3 is an important signaling factor for promoting colonization of EHEC in the lower intestinal tract in humans.Enterohemorrhagic Escherichia coli (EHEC) is a leading cause of hemorrhagic colitis, bloody diarrhea, and hemolytic uremic syndrome (31, 42). Early in infection leading to illness, the ability of the bacteria to colonize the intestinal epithelial surface and cause histopathological alterations at so-called attaching and effacing lesions (A/E lesions) is thought to be vital. The formation of A/E lesions is characterized by localized destruction of the brush border microvilli, intimate attachment to the host cell, and reconstitution of cytoskeletal components beneath the attached bacteria (11). Thus, EHEC shares pathogenic features with enteropathogenic E. coli (EPEC), rabbit enteropathogenic E. coli, and Citrobacter rodentium, although the extent to which A/E lesion formation, including the target host tissue, varies among the pathogens is not clear.Most of the genes necessary to form A/E lesions are located in a pathogenicity island termed the locus for enterocyte effacement (LEE) (7, 27, 32). This locus contains (i) sep and esc genes encoding a type III secretion system (17), (ii) eae encoding an adhesin called intimin that is necessary for intimate attachment to epithelial cells (18), (iii) espA, espB, espD, and tir genes encoding proteins secreted by the type III secretion system, including EspA, EspB, EspD, and Tir (3,20,22,23,25), and (iv) ler encoding a positive regulator of LEE (LEEencoded regulator) (6, 30). Esp proteins...

Adherence of enterohemorrhagic Escherichia coli (EHEC) to intestinal epithelium is essential for initiation of the infection.To identify genes involved in adherence, an EHEC O157:H7 strain (O157Sakai) was mutagenized by mini-Tn5Km2, where Km refers to kanamycin resistance, and 4,677 insertion mutants were screened for their ability to form microcolonies (MC) on Caco-2 cells. The less adherent mutants were divided into three groups: those with no adherent ability (designated as class 1 mutants, n ‫؍‬ 10), those less adherent than the wild type (class 2 mutants, n ‫؍‬ 16), and those unable to form MC but which adhered in a diffuse manner (class 3 mutants, n ‫؍‬ 1). The sites of insertion in class 1 mutants were all found within genes of the locus for enterocyte effacement (LEE) thought to be required for type III protein secretion. Indeed, the class 1 mutants failed to secrete type III secreted proteins such as EspA and Tir into the culture medium. The insertions in class 2 mutants were outside the LEE, and all the mutants except one were able to secrete type III proteins into the culture medium. The class 3 mutant had the insertion in the tir gene in the LEE and was deficient in Tir and intimin expression, suggesting that in the absence of intimin-Tir, O157Sakai can still adhere to Caco-2 cells but in a diffused manner. This was confirmed by construction of a nonpolar eae (encoding intimin) mutant. Examination of the eae mutant together with O157Sakai and one of the class 1 mutants for the ability to form MC revealed that EHEC initially adhered diffusely at 1.5 h after infection. Following washing out of the nonadherent bacteria, while wild-type EHEC bacteria developed MC for another 2 to 3 h on Caco-2 cells, the eae mutant diffusely adhered throughout the infection without forming MC. MC with O157Sakai but not the diffusely adherent eae mutant could evoke F-actin condensation beneath the bacterium. Our results suggest that EHEC encodes additional adherence-associated loci and that the type III secreted proteins are involved in the initial diffuse adherence, while the intimin-Tir interaction is required for the subsequent development of MC.

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging hemorrhagic fever caused by the SFTS phlebovirus (SFTSV). SFTS patients were first reported in China, followed by Japan and South Korea. In 2017, cats were diagnosed with SFTS for the first time, suggesting that these animals are susceptible to SFTSV. To confirm whether or not cats were indeed susceptible to SFTSV, animal subjects were experimentally infected with SFTSV. Four of the six cats infected with the SPL010 strain of SFTSV died, all showing similar or more severe symptoms than human SFTS patients, such as a fever, leukocytopenia, thrombocytopenia, weight loss, anorexia, jaundice and depression. High levels of SFTSV RNA loads were detected in the serum, eye swab, saliva, rectal swab and urine, indicating a risk of direct human infection from SFTS-infected animals. Histopathologically, acute necrotizing lymphadenitis and hemophagocytosis were prominent in the lymph nodes and spleen. Severe hemorrhaging was observed throughout the gastrointestinal tract. B cell lineage cells with MUM-1 and CD20, but not Pax-5 in the lesions were predominantly infected with SFTSV. The present study demonstrated that cats were highly susceptible to SFTSV. The risk of direct infection from SFTS-infected cats to humans should therefore be considered.