1510株肠道杆菌大肠菌素调查及其检测方法的研究

对７个菌属１５１０株肠道杆菌进行了大肠菌素检测，宋内氏菌Ｃｏｌ质粒检出率最高为７８．１８％，其次为普通大肠杆菌３３．４７％，福氏志贺氏菌、克雷伯氏菌、伤寒沙门氏菌分别为８．０％、３．５％、２．３２％，不同种属菌株大肠菌素检出率差异悬殊。药敏试验证明，９０．５％（１０５／１１６）Ｃｏｌ＋大肠杆菌具有耐药性，在接合实验中，２６．２％（１６／６１）大肠杆菌能传递Ｃｏｌ质粒。将Ｃｏｌ＋菌株接种保种半固体置室温２年，１９．９％（３１／１５６）的菌株丢失Ｃｏｌ质粒。本实验同时建立一种直接覆盖指示菌──插种受检菌以检测大肠菌素的方法。通过大量菌株考核，结果与Ｍｏｎｋ法基本一致，而且比后者简单、快速。在细菌素的调查研究中具有广泛的应用价值。

大肠菌素Colicin S4的重组表达、分离纯化和性质

传统抗生素的广谱抗菌特性是诱导细菌普遍产生耐药性重要原因之一。细菌素由于具有窄谱抗菌活性而可能被开发为新一代抗菌药物。大肠菌素因被认为对人体安全而倍受关注。本研究利用PCR方法获得了大肠菌素S4及其抑制蛋白基因并将其克隆到pQE30质粒中,构建了表达质粒pQE30-Col S4。大肠菌素S4在该表达体系中以可溶形式高表达,产量达到30-50 mg/L。活性分析发现,带有6His标签的重组大肠菌素S4与天然大肠菌素S4特性一致,仍具有对大肠杆菌的选择性抗菌活性,是一种有潜力的抗菌物质。

抗耐药金黄色葡萄球菌工程多肽—一种人工构建的抗菌蛋白质分子机器

目的 将两个不同种来源 ,不同生物活性的蛋白质结构域构建成为一种具有靶向抗菌活性的工程多肽。方法 利用质粒重组 ,将金黄色葡萄球菌 Agr D信息素 (8肽 )基因连接在大肠菌素 Ia水性孔道结构域 (K5 4 4 -I6 2 6 )羧基端基因上 ,将重组的质粒转化入工程菌生产构建的工程多肽 ,离子交换柱纯化后经体外抗菌试验检测其抗菌活性。结果 构建的工程多肽具有强于青霉素类抗生素上百倍的抗耐药金黄色葡萄球菌的活性。结论 构建的工程多肽表现出了两个结构域前体都不具备的靶向抗金黄色葡萄球菌活性 。

Establishment and Application of a Real-time PCR Method for Detecting stx2 Gene in Shiga Toxin-producing Escherichia coli(STEC)

[Objective] This study aimed to establish a real-time PCR method for de- tecting stx2 gene in Shiga toxin-producing E. coli （STEC）. [Method] According to the known STEC stx2 gene sequences published in GenBank, PCR primers and probes were designed based on the conserved region to construct recombinant plasmid as a positive template, thus optimizing the reaction conditions and establishing the real- time PCR method. [Result] A standard curve was established based on the opti- mized real-time PCR system, indicting a good linear correlation between the initial template concentration and Ct value, with the correlation coefficient F^e of above 0.995. The established method had a good specificity, without non-specific amplifica- tion for 10 non-STEC intestinal bacterial strains; the detection limit of initial template was 1.0x102 copies/μI, indicating a high sensitivity; furthermore, the coefficients of variation within and among batches were lower than 1% and 5% respectively, sug- gesting a good repeatability. [Conclusion] In this study, a real-time PCR method was successfully established for detecting STEC stx2 gene, which provided technical means for rapid detection of STEC in samples.

AtTHIC, a gene involved in thiamine biosynthesis in Arabidopsis thaliana

Thiamine （vitamin B1） is an essential compound for organisms. It contains a pyrimidine ring structure and a thiazole ring structure. These two moieties of thiamine are synthesized independently and then coupled together. Here we report the molecular characterization of AtTHIC, which is involved in thiamine biosynthesis in Arabidopsis. AtTHIC is similar to Escherichia coli ThiC, which is involved in pyrimidine biosynthesis in prokaryotes. Heterologous expression of AtTHIC could functionally complement the thiC knock-out mutant of E. coll. Downregulation of AtTHIC expression by T-DNA insertion at its promoter region resulted in a drastic reduction of thiamine content in plants and the knock-down mutant thicl showed albino （white leaves） and lethal phenotypes under the normal culture conditions. The thicl mutant could be rescued by supplementation of thiamine and its defect functions could be complemented by expression ofAtTHIC cDNA. Transient expression analysis revealed that the AtTHIC protein targets plastids and chloroplasts. AtTHIC was strongly expressed in leaves, flowers and siliques and the transcription of AtTHIC was downregulated by extrinsic thiamine. In conclusion, AtTHIC is a gene involved in pyrimidine synthesis in the thiamine biosynthesis pathway of Arabidopsis, and our results provide some new clues for elucidating the pathway of thiamine biosynthesis in plants.

Construction of a novel Shigella live-vector strain co-expressing CS3 and LTB/STm of enterotoxigenic E.coli

AIM: To construct and evaluate a polyvalent recombinant vaccine strain Shigella flexneri2a T32 against enterotoxigenic E.coli/(ETEC). METHODS: By using a host-plasmid balanced lethal system based on asd gene, a polyvalent recombinant strain was constructed to highly express CS3 and regularly express fusion enterotoxin of LIB subunit and mutant ST (LTB/STm) in a vaccine strain Shigella flexneri 2a T32 with specific deletion of asd gene. Fimbria CS3 was observed by immunofluorescence and electron microscopy assay. The security of LTB/STm was examined by ileal loop assay and suckling mouse assay. To evaluate this new candidate vaccine, it was compared with a previous vaccine strain in plasmid and protein level, growth assay and immunogenicity in Balb/c mice. RESULTS: The newly constructed vaccine expressed CS3 and grew better than the previously constructed vaccine except for the lower expression of LTB/STm. Serum IgG and mucosal IgA against CS3, LTB, ST, and host lipopolysaccharide (LPS) were produced after immunization of Balb/c mice by oral route with the new strain. The titers were not significantly different from the Balb/c mice with the previous strain. CONCLUSION: This novel candidate diarrheal vaccine can effectively induce serum and mucosal antibody responses against ETEC and Shigella.

Chloromycetin resistance of clinically isolated E coli is conversed by using EGS technique to repress the Chloromycetin acetyl transferase

AIM： To explore the possibility of repression of chloromycetin （Cm） acyl transferase by using external guided sequence （EGS） in order to converse the clinical E coli isolates from Cm- resistant to Cm- sensitive. METHODS： EGS directed against chloromycetin acetyl transferase gene （cat） was cloned to vector pEGFP-C1 which contains the kanamycin （Kin） resistance gene. The recombinant plasmid pEGFP-C1＋EGScatl＋cat2 was constructed and the blank vector without EGS fragment was used as control plasmids. By using the CaCl2 transformation method, the recombinant plasmids were introduced into the clinically isolated Cm resistant but Km sensitive E coli strains. Transformants were screened on LB agar plates containing Kin. Extraction of plasmids and PCR were applied to identify the positive clones. The growth curve of EGS transformed bacteria cultured in broth with Cm resistance was determined by using spectrophotometer at A600. Drug sensitivity was tested in solid culture containing Cm by using KB method. RESULTS： Transformation studies were carried out on 16 clinically isolated Cm-resistant （250 μg/mL of Cm） E colistrains by using pEGFP-C1-EGScatlcat2 recombinant plasmid. Transformants were screened on LB-agar plates containing Km after the transformation using EGS. Of the 16 tested strains, 4 strains were transformed successfully. Transformants with EGS plasmid showed growth inhibition when grown in liquid broth culture containing 200 μg/mL of Cm. In drug sensitivity test, these strains were sensitive to Cm on LB-agar plates containing 200 μg/mL of Cm. Extraction of plasmids and PCR amplification showed the existence of EGS plasmids in these four transformed strains. These results indicated that the Cat of the four clinical isolates had been suppressed and the four strains were converted to Cm sensitive ones. CONCLUSION： The EGS directed against Cat is able to inhibit the expression of Cat, and hence convert Cm- resistant bacteria to Cm-sensitive ones. Thus, the EGS has the capability of converting the phenotype of clinical drug-resistant isolates strains to drug-sensitive ones.

Clinical features of familial adenomas polyps in Chinese and establishment of its immortal lymphocyte cell lines

AIM： To reserve the rare Chinese familial adenomas polyp （FAP） family resource and to investigate the clinical features of FAP in Chinese for its diagnosis. METHODS： Clinical features of patients with FAP were investigated. If there is any question, their medical records were verified. Blood sample was taken and lymphocyte immortal cell lines were established with modified EB-transformation methods. Congenital hypertrophy of retinal pigment epithelium （CHRPE） was checked by an experienced ophthalmologist. RESULTS： Twenty seven families including 21 classical FAP （CFAP） families, 3 attenuated FAP （AFAP） families, and 3 suspected AFAP families were investigated. A total of 116 lymphocyte immortal cell lines were established from 26 families. In all the FAP families, colorectal cancer occurred at the mean age of 42.84 years. Of the 16 families checked, 15 （93.75%） had CHRPE. The mean number of patients suffering from colorectal neoplasm was 3.14 in CFAP families and 2.0 in AFAP families （P 〈 0.01）. The mean oldest age at diagnosis of FAP was 41.75 years in CFAP families, and 58.67 years in AFAP families, respectively （P 〈 0.01）. Mean age of development of colorectal cancer was 42.23 in CFAP and 57.33 years old in AFAP （P 〈 0.01）. Mean of the earliest age at diagnosis of FAP was 29.95 years in the FAP families with a positive family history and 46.80 years in the FAP families with a negative family history （P 〈 0.01）. The ratio of extra-intestinal tumors to colorectal neoplasms was different in the two kinds of families with positive and negative family history （P 〈 0.01）. CONCLUSION： Additional use of ciclosporin will effectively improve to establish lymphocyte immortal cell lines with modified EB- transformation methods. In Chinese FAP, there was a high frequency of CHRPE, and a later age at diagnosis and a later age of development of colorectal cancer in AFAR And earlier age at diagnosis in FAP with positive family history was also found that will help to diagnose various kinds of FAP in Chinese.

Sedimentary response to volcanic activity in the Okinawa Trough since the last deglaciation

To investigate the relationship between volcanic activity and sediment record on regional and temporal scales,158 surface sediment samples were collected from the East China Sea Shelf to the northern Okinawa Trough (OT),and two cores recovered in the northern and southern OT,respectively.Mineralogy,grain-size,and geochemical analyses of those samples show that:1) volcanic glass,volcanic-type pyroxene,hypersthenes,and magnetite increase in sediment influenced by volcanic activity;2) sediment grain sizes (and also silt content) increase in ash layers;and 3) the contents of Na2O and Zr are higher,while terrigenous elements,e.g.,TFe2O3 and K2O,and biogenous compositions,e.g.,CaO and Sr,are relatively lower in ash layers than those of non-ash layers.The distribution of volcanic ash has three distinguishing characteristics:1) volcanic ash is more abundant in the northern and central OT than the southern OT;2) volcanic ash increases from continental shelf to the trough;3) the sediment during the last 12 000 a suggests stronger volcanic events than during 15 000-12 000 a.The eruptive locations,frequency,and volume of calderas are among the most important factors controlling the distributions of volcanic ash.In addition,the main Kuroshio warm current that extends northward probably impeded the diffusion of volcanic ash to the west and south in the OT.However,a southward current probably carried some volcanic ash toward southern OT.

Suppression of colorectal cancer metastasis by nigericin through inhibition of epithelial-mesenchymal transition

AIM: To evaluate the effect of nigericin on colorectal cancer and to explore its possible mechanism. METHODS: The human colorectal cancer (CRC) cell lines HT29 and SW480 were treated with nigericin or oxaliplatin under the conditions specified. Cell viability assay and invasion and metastasis assay were performed to evaluate the effect of nigericin on CRC cells. Sphereforming assay and soft agar colony-forming assay were implemented to assess the action of nigericin on the cancer stem cell properties of CRC cells undergone epithelial-mesenchymal transition (EMT). RESULTS: Compared with oxaliplatin, nigericin showed more toxicity for the HT29 cell line (IC50, 12.92 ± 0.25 μmol vs 37.68 ± 0.34 μmol). A similar result was also obtained with the SW116 cell line (IC50, 15.86 ± 0.18 μmol vs 41.02 ± 0.23 μmol). A Boyden chamber assay indicated that a significant decrease in the number of HT29 cells migrating through polyvinylidene fluoride membrane was observed in the nigericin-treated group, relative to the vehicle-treated group [11 ± 2 cells per high-power field (HPF) vs 19.33 ± 1.52 cells per HPF, P < 0.05]. Compared to the control group, the numbers of HT29 cells invading through the Matrigel-coated membrane also decreased in the nigericin-treated group (6.66 ± 1.52 cells per HPF vs 14.66 ± 1.52 cells per HPF, P < 0.05). Nigericin also reduced the proportion of CD133+ cells from 83.57% to 63.93%, relative to the control group (P < 0.05). Nigericin decreased the number of spheres relative to the control group (0.14 ± 0.01 vs 0.35 ± 0.01, P < 0.05), while oxaliplatin increased the number of spheres relative to the control group (0.75 ± 0.02 vs 0.35 ± 0.01; P < 0.05). Nigericin also showed a decreased ability to form colonies under anchorage-independent conditions in a standard soft agar assay after 14 d in culture, relative to the control group (1.66 ± 0.57 vs 7 ± 1.15, P < 0.05), whereas the colony numbers were higher in the oxaliplatin group relative to the vehicle-treated controls (14.33 ± 0.57 vs 7 ± 1.15, P < 0.05). We further detected the expression of E-cadherin and vimentin in cells treated with nigericin and oxaliplatin. The results showed that HT29 cells treated with nigericin induced an increase in E-cadherin expression and a decrease in the vimentin expression relative to vehicle controls. In contrast, oxaliplatin downregulated the expression of E-cadherin and upregulated the expression of vimentin in HT29 cells relative to vehicle controls. CONCLUSION: This study demonstrated that nigericin could partly reverse the EMT process during cell invasion and metastasis.

Soluble Expression and Rapid Quantification of GFP-hepA Fusion Protein in Recombinant Escherichia coli

To establish a rapid quantification method for heparinase I during its production in recombinant Escherichia coli, a translational fusion vector was constructed by fusing the N terminus of heparinase I to the C terminus of a green fluorescent protein mutant （GFPmutl）. As a result, not only was the functional recombinant expression of heparinase I in E. coli accomplished, but also a linear correlation was obtained between the GFP fluorescence intensity and heparinase I activity, allowing enzyme activity to be quantified rapidly during the fermentation.

Moxibustion inhibits interleukin-12 and tumor necrosis factor alpha and modulates intestinal flora in rat with ulcerative colitis

AIM: To investigate the effect of moxibustion on intestinal flora and release of interleukin-12 (IL-12) and tumor necrosis factor-α (TNF-α) from the colon in rat with ulcerative colitis (UC). METHODS: A rat model of UC was established by local stimulation of the intestine with supernatant from colonic contents harvested from human UC patients. A total of 40 male Sprague-Dawley rats were randomly divided into the following groups: normal (sham), model (UC), herb-partition moxibustion (HPM-treated), and positive control sulfasalazine (SA-treated). Rats treated with HPM received HPM at acupuncture points ST25 and RN6, once a day for 15 min, for a total of 8 d. Rats in the SA group were perfused with SA twice a day for 8 d. The colonic histopathology was observed by hematoxylin-eosin. The levels of intestinal flora, including Bifidobacterium, Lactobacillus, Escherichia coli (E. coli), and Bacteroides fragilis (B. fragilis), were tested by real-time quantitative polymerase chain reaction to detect bacterial 16S rRNA/DNA in order to determine DNA copy numbers of each specific species. Immunohistochemical assays were used to observe the expression of TNF-α and IL-12 in the rat colons. RESULTS: HPM treatment inhibited immunopathology in colonic tissues of UC rats; the general morphological score and the immunopathological score were significantly decreased in the HPM and SA groups compared with the model group [3.5 (2.0-4.0), 3.0 (1.5-3.5) vs 6.0 (5.5-7.0), P < 0.05 for the general morphological score, and 3.00 (2.00-3.50), 3.00 (2.50-3.50) vs 5.00 (4.50-5.50), P < 0.01 for the immunopathological score]. As measured by DNA copy number, we found that Bifidobacterium and Lactobacillus, which are associated with a healthy colon, were significantly higher in the HPM and SA groups than in the model group (1.395 ± 1.339, 1.461 ± 1.152 vs 0.045 ± 0.036, P < 0.01 for Bifidobacterium, and 0.395 ± 0.325, 0.851 ± 0.651 vs 0.0015 ± 0.0014, P < 0.01 for Lactobacillus). On the other hand, E. coli and B. fragilis, which are associated with an inflamed colon, were significantly lower in the HPM and SA groups than in the model group (0.244 ± 0.107, 0.628 ± 0.257 vs 1.691 ± 0.683, P < 0.01 for E. coli, and 0.351 ± 0.181, 0.416 ± 0.329 vs 1.285 ± 1.039, P < 0.01 for B. fragilis). The expression of TNF-α and IL-12 was decreased after HPM and SA treatment as compared to UC model alone (4970.81 ± 959.78, 6635.45 ± 1135.16 vs 12333.81 ± 680.79, P < 0.01 for TNF-α, and 5528.75 ± 1245.72, 7477.38 ± 1259.16 vs 12550.29 ± 1973.30, P < 0.01 for IL-12). CONCLUSION: HPM treatment can regulate intestinal flora and inhibit the expression of TNF-α and IL-12 in the colon tissues of UC rats, indicating that HPM can improve colonic immune response.

Mutants of Escherichia coli heat-labile enterotoxin and cholera toxin as mucosal adjuvants

Mucosal vaccination has been getting more and more recognition because of its compliance and low risk of spreading infectious disease by contaminated syringes used in subcutaneous immunization. However, most vaccines are unable to induce immune responses when given mucosally, and require the use of strong adjuvant for effective delivery systems. Heat-labile enterotoxin (LT) and Cholera toxin(CT) are powerful mucosal adjuvants when co-administered with soluble antigens. But high toxicity hampers their use in humans. Thanks to the fine knowledge of the structure-function relationship of LT and CT, many nontoxic or low toxic mutants have been generated, part of them retain high adjuvanticity of mucosal immunization. Among these mutants, LTS63K, LTA72R, LTR192G and CTE29H, CTE112K have been widely investigated. LTS63K and CTE112K are fully non toxic, whereas LTA72R and CTE29H are low toxic, and LTR192G is nontoxic in vitro(it remains the same toxicity as wild type LT in vivo). These mutants are extremely active as mucosal adjuvants when co-administrated with a variety of antigens in different animal models. They will be investigated more widely and deeply in the future. Some of them will be tested soon in human bodies.

Efficient and Comprehensive Utilizatio℃n of Hemicellulose in the Corn Stover

Pretreatment of the corn stover powder by dilute sulphuric acid （solid-liquid ratio 1 ： 20） at 130 for 30 min was carried out with 89.09% of the hemicellulose removed. After filtration, the xylose-rich corn stover pretreatment liquid, whose fermentable sugar was from hemicellulose hydrolysis only, consisting of 81.16% xylose and 15.27% glucose, was used to cultivate genetic recombinant Escherichia coli BL21 with human-like collagen （HLC） expression enhanced by 50.00% and 63.71% xylose consumption.

Gastric dysplasia may be an independent risk factor of an advanced colorectal neoplasm

AIM: To evaluate the relationship between gastric dysplasia and Helicobacter pylori (H pylori) and the occurrence of colorectal adenoma, and to defi ne the necessity for colonoscopy in patients with gastric dysplasia or H pylori infection.METHODS: From May 2005 to February 2008, 133 patients with established gastric dysplasia by gastroduo-denoscopy (EGD) were additionally investigated by colonoscopy. The authors compared results with those of 213 subjects who underwent both EGD and colonoscopy during the same period at the author’s Health Promotion Center as a control group. H pylori infection was evaluated in both the gastric dysplasia and control groups.RESULTS: The mean age of all 346 study subjects was 54.1 ± 10.5 years, and there were 258 (73%) men and 87 (27%) women. No signif icant difference was found between the H pylori positive and negative subjects in terms of the prevalence of colorectal adenoma and advanced colorectal adenoma (P = 0.261). Patients with gastric dysplasia showed no elevated risk of colorectal adenoma (OR = 0.910, 95% CI: 0.587-1.411, P = 0.738), but had a signif icantly higher risk of having advanced colorectal adenoma (OR = 3.382, 95% CI: 1.700-6.342, P = 0.000).CONCLUSION: The study emphasizes the need for colon surveillance in patients with gastric dysplasia, regardless of H pylori infection.

Identification of E. coli K12 chromosomal insertion sites of bacteriophage φ297

Objective:To identify the specific integration site of prophage φ297 in the host of E. coli K12 chromosome. Methods:Using molecular techniques such as Siebert PCR for walking from the int gene of prophage 297, which is similar to that of phage 933W to an unknown region in genomic DNA. A special adaptor is ligated to the ends of DNA fragments generated by digestion of genomic DNA with restriction enzymes that generates blunt ended fragments. Clone and subclone of PCR products, DNA sequencing and data analysis were used in this study. Results:The attL, attR and the core sequences were determined. The bacterial attachment site of phage φ297 was located in the yecE gene of E. coli K12. Conclusion:The phage φ297 integrates into the yecE gene of the E. coli K12 genome.

Expression of E. coli heat-labile enterotoxin B subunit in transgenic tobacco plants

Objective ： To construct plant transformation vector containing Escherichia coli heat-labile enterotoxin B subunit （LT-B） gene and generate LT-B transgenic tobacco plants. Methods： The LT-B coding sequence was amplified from pMMB68 by PCR, subcloned into middle vector pUCmT and binary vector pBI121 to obtain plant expression vector pBI-LTB, in which LT-B expression was controlled under the Cauliflower mosaic virus （CaMV） 35S promoter. The tobacco plants （Nicotiana tobacum L. Cuttivar Xanthi） were transformed by co-cultivating leaf discs method via Agrobacterium tumefaciens LBA4404 harboring the plant expression vector. The regenerated transgenic tobacco plants were selected by kanamycin and confirmed by PCR, Southern blot, Western blot and ELISA. Resuits： LT-B gene integrated in the tobacco genomic DNA and were expressed in 9 strains of transgenic tobacco plants. The yield was varied from 3. 36-10. 56 ng/mg total soluble tobacco leaf protein. Conclusion： The plant binary expression vector pBI-LTB was constructed successfully, and transgenic LT-B tobacco plants was generated, and confirmed by Southern blot. The protein LT-B expressed by engineered plants was identified by Western blot analysis and had the expected molecular weight of LT-B pentamer protein. This result is an important step close to developing an edible vaccine and supplying a mucasal immunoajuvant, which will contribute to the preven- tion of mucosaroute evading pathogen.

Primary Purification of Co-expressed Soluble and Insoluble Alpha-interferon 2b from Recombinant E. coli

Alpha-interferon 2b (IFN 2b) was produced both in soluble and insoluble forms from recombinant E. coli. The dissolution of the expressed IFN 2b in inclusion body was carried out and it was found that the optimal condition to dissolve the expressed protein was 7 mol稬1guanidinium salt solution at pH 3.0. The resultant solution was diluted 20 times using pH 6.0 buffer to refold the protein correctly. The cation exchange column was employed to purify both refolded and soluble IFN 2b. For soluble IFN sample, high IFN 2b recovery yield (92.1%) with 91.7% purity was obtained in the eluate. However, for refolded IFN sample, only 72.7% of IFN 2b was recovered with relatively low purity (56.8%) by cation exchange chromatography. Although the expression level of insoluble IFN was higher than that of co-expressed soluble IFN in this recombinant E. coli cells, the productivity of bioactive IFN 2b was higher with soluble expressed IFN after primary purification process. Soluble expression of foreign proteins in recombinant bacteria might be an alternative strategy for efficient production of heterogeneous pro-teins due to high bioactivity and simple downstream protein purification process.

Neutron （Magnetic Isotope） Catalysis for Example Isotopes 24,25,26Mg in Cells E. Coli

We offered the new theory of neutron （magnetic isotope） catalysis. For the first time it was shown that the number of neutrons in the atom, which have anomalous magnetic effect, have a great influence on the chemical properties. Our proposed theory of neutron （magnetic isotope） catalysis takes into account the influence of the magnetic field on the catalytic processes.

The Truncated Gene cfaD′ Positively Regulates CFA/Ⅰ Expression of Enterotoxigenic Escherichia coli

The gene cluster cfaABCED’ of enterotoxigenic Escherichia coli, encoding the fimbriae which is called colonization factor antigen located on a plasmid. It is positively regulated by cfaR, a member of the AraC family, and the cfaD’ gene region, which is located downstream of cfaE and is homologous to cfaR, had been described as a truncated cryptic gene. In the present study we observed that the CFA/ fimbriae subunit, cfaB, was expressed in lower amount by the cfaABCED’ clone pNTP513 in host E. coli HB101. The expression of CFA/ diminished by deletion of cfaD’ gene region from pNTP513, and was restored by acquisition of cfaD’ in trans. Furthermore, CFA/ expression by cfaD’ deletion mutant, the cfaABCE clone, was remarkably increased by the presence of cfaD’ in trans in a topoisomerase A deficient strain of E. coli DM800. These data suggest that cfaD’ region is a functional region of gene, that regulates the CFA/ expression with cfaR by unknown mechanism.