Novel electric power-driven hydrodynamic injection system for gene delivery: safety and efficacy of human factor IX delivery in rats.

The development of a safe and reproducible gene delivery system is an essential step toward the clinical application of the hydrodynamic gene delivery (HGD) method. For this purpose, we have developed a novel electric power-driven injection system called the HydroJector-EM, which can replicate various time-pressure curves preloaded into the computer program before injection. The assessment of the reproducibility and safety of gene delivery system in vitro and in vivo demonstrated the precise replication of intravascular time-pressure curves and the reproducibility of gene delivery efficiency. The highest level of luciferase expression (272 pg luciferase per mg of proteins) was achieved safely using the time-pressure curve, which reaches 30 mm Hg in 10 s among various curves tested. Using this curve, the sustained expression of a therapeutic level of human factor IX protein (>500 ng ml(-1)) was maintained for 2 months after the HGD of the pBS-HCRHP-FIXIA plasmid. Other than a transient increase in liver enzymes that recovered in a few days, no adverse events were seen in rats. These results confirm the effectiveness of the HydroJector-EM for reproducible gene delivery and demonstrate that long-term therapeutic gene expression can be achieved by automatic computer-controlled hydrodynamic injection that can be performed by anyone.

Distinguishing adrenal adenomas from non-adenomas on dynamic enhanced CT: a comparison of 5 and 10 min delays after intravenous contrast medium injection.

AIM: To evaluate the usefulness of several parameters of 5 min compared to 10 min delayed contrast-enhanced CT in distinguishing adenomas from non-adenomas. MATERIALS AND METHODS: The study population consisted of 94 patients (52 men and 42 women; mean age 62 years) with 103 adrenal lesions (75 adenomas and 28 non-adenomas). In each patient, unenhanced CT was followed by early, 5 and 10 min enhanced CT. Diagnostic parameters included delayed enhanced attenuation at 5 and 10 min, washout attenuation (WO) at 5 and 10 min, absolute percentage washout (APW) at 5 and 10 min, and relative percentage washout (RPW) at 5 and 10 min. The accuracy of each parameter for diagnosing adenomas from non-adenomas was calculated using receiver operating characteristic (ROC) analysis. RESULTS: Upon comparison between 5 and 10 min delayed contrast-enhanced CT for differentiating total adenomas or lipid-poor adenomas from non-adenomas, there was no significant difference in the area under the binomial ROC curve (Az) values of delayed enhanced attenuation (total adenomas versus non-adenomas, p = 0.164; lipid-poor adenomas versus non-adenomas, p = 0.178), WO (total adenomas versus non-adenomas, p = 0.216; lipid-poor adenomas versus non-adenomas, p = 0.230), APW (total adenomas versus non-adenomas, p = 0.401; lipid-poor adenomas versus non-adenomas, p = 0.870), or RPW (total adenomas versus non-adenomas, p = 0.160; lipid-poor adenomas versus non-adenomas, p = 0.780). CONCLUSION: Five minute contrast-enhanced CT was as useful as 10 min contrast-enhanced CT for differentiation of adrenal adenomas from non-adenomas.

Continuous ulnar nerve block at the forearm for early active mobilisation following flexor tendon reconstruction.

A 63-year-old woman had sustained a subcutaneous rupture of the flexor digitorum profundus tendon of the little finger due to osteoarthritis of the pisotriquetral joint. She underwent excision of the pisiform bone and reconstruction of the flexor digitorum profundus tendon of the little finger using an autogenous palmaris longus tendon graft. After surgery, a continuous ulnar nerve block was performed at the forearm under ultrasound and nerve stimulator guidance. During rehabilitation, she could not actively extend her little finger independently due to the block; however, she could actively extend it when the dorsum of the metacarpophalangeal joint was pressed by the occupational therapist, resulting in successful early active mobilisation. A continuous ulnar nerve block at the forearm may help to facilitate early active mobilisation after reconstructive surgery for little finger flexor tendon rupture. However, it may restrict the active extension of the little finger because the block does not spare the innervation of the intrinsic muscles responsible for little finger extension.

Functions of chondroitin sulfate and heparan sulfate in the developing brain.

Chondroitin sulfate and heparan sulfate proteoglycans are major components of the cell surface and extracellular matrix in the brain. Both chondroitin sulfate and heparan sulfate are unbranched highly sulfated polysaccharides composed of repeating disaccharide units of glucuronic acid and N-acetylgalactosamine, and glucuronic acid and N-acetylglucosamine, respectively. During their biosynthesis in the Golgi apparatus, these glycosaminoglycans are highly modified by sulfation and C5 epimerization of glucuronic acid, leading to diverse heterogeneity in structure. Their structures are strictly regulated in a cell type-specific manner during development partly by the expression control of various glycosaminoglycan-modifying enzymes. It has been considered that specific combinations of glycosaminoglycan-modifying enzymes generate specific functional microdomains in the glycosaminoglycan chains, which bind selectively with various growth factors, morphogens, axon guidance molecules and extracellular matrix proteins. Recent studies have begun to reveal that the molecular interactions mediated by such glycosaminoglycan microdomains play critical roles in the various signaling pathways essential for the development of the brain.

Multi-step lymphomagenesis deduced from DNA changes in thymic lymphomas and atrophic thymuses at various times after gamma-irradiation.

Whole-body gamma-irradiation to mice causes thymic atrophy where a population of precancerous cells with mutation can be found. Thus, clonal growth and DNA changes at Bcl11b, Ikaros, Pten, Notch1 and Myc were examined in not only thymic lymphomas but also in atrophic thymuses at various times after irradiation. Clonal expansion was detected from the distinct patterns of rearrangements at the TCRbeta receptor locus in a fraction of atrophic thymuses at as early as 30 days after irradiation. This expansion may be in part owing to the rearranged TCRbeta signaling because the transfer of bone marrow cells with the rearrangement and the wild-type locus into severe-combined immunodeficiency mice showed preferential growth of the rearranged thymocytes in atrophic thymus. Loss of heterozygosity (LOH) at Bcl11b and trisomy of Myc were found at high frequencies in both lymphomas and atrophic thymuses, and in contrast, LOH at Ikaros and Pten were rare in atrophic thymuses but prevalent in lymphomas. Notch1 activation was detected in lymphomas and in atrophic thymuses only at a late stage. Similar patterns of DNA changes were found in atrophic thymuses induced in Bcl11b(+/-) mice. These results suggest the order of genetic changes during lymphomagenesis, Bcl11b and Myc being at the early stage; whereas Ikaros, Pten and Notch1 at the late stage.

Lack of Bcl11b tumor suppressor results in vulnerability to DNA replication stress and damages.

Bcl11b/Rit1 is involved in T-cell development and undergoes chromosomal rearrangements in human T-cell leukemias. Thymocytes of Bcl11b(-/-) newborn mice exhibit apoptosis at a certain developmental stage when thymocytes re-enter into the cell-cycle. Here, we show that Bcl11b-knockdown T-cell lines, when exposed to growth stimuli, exhibited apoptosis at the S phase with concomitant decreases in a cell-cycle inhibitor, p27 and an antiapoptotic protein, Bcl-xL, owing to transcriptional repression. This repression was a likely consequence of the impairment of Sirt1, a nicotinamide adenine dinucleotide-dependent deacetylase associating with Bcl11b. Activation of the apoptotic process cleaved the mediator protein, Claspin, and inhibited phosphorylation of cell-cycle checkpoint kinase 1 (Chk1) that plays a central role in sensing and responding to incomplete replication. Bcl11b(-/-) thymocytes also failed to phosphorylate Chk1 when UV irradiated. These results implicate Bcl11b in the remedy for DNA replication stress and maintenance of genomic integrity.

Downregulation of HLA Class I molecules in the tumour is associated with a poor prognosis in patients with oesophageal squamous cell carcinoma.

As antigenic peptides in the context of human leukocyte antigen (HLA) class I molecules are recognised by cytotoxic T lymphocytes (CTL), the downregulation of HLA class I molecules is one of the reasons why tumour cells can evade CTL-mediated anti-tumour immunity. In this study, we investigated HLA class I expression in oesophageal squamous cell carcinoma (ESCC) (n=70) and in their metastatic lesions (lymph nodes (n=40) and liver (n=3)), by immunohistochemistry with anti-HLA class I monoclonal antibody (EMR8-5). As a result, the downregulation of HLA class I expression in primary lesions of ESCC was observed in 43%, and that in metastatic lymph nodes was noted in 90%. Furthermore, patients with preserved HLA class I expression in primary tumours showed a better survival in comparison to those with downregulated HLA class I molecules (P<0.01). Furthermore, multivariate analysis using Cox's proportional hazards model revealed that the downregulated expression of HLA class I in primary lesions was an independent, unfavourable prognostic factor (P<0.01). In conclusion, the downregulation of HLA class I expression frequently occurred in primary tumour and, to a greater extent, in metastatic lesions of patients with ESCC and was associated with patient survival.

Intravoxel Incoherent Motion in Normal Pituitary Gland: Initial Study with Turbo Spin-Echo Diffusion-Weighted Imaging.

BACKGROUND AND PURPOSE: DWI with conventional single-shot EPI of the pituitary gland is hampered by strong susceptibility artifacts. Our purpose was to evaluate the feasibility of intravoxel incoherent motion assessment by using DWI based on TSE of the normal anterior pituitary lobe. MATERIALS AND METHODS: The intravoxel incoherent motion parameters, including the true diffusion coefficient (D), the perfusion fraction (f), and the pseudo-diffusion coefficient (D*), were obtained with TSE-DWI in 5 brain regions (the pons, the WM and GM of the vermis, and the genu and splenium of the corpus callosum) in 8 healthy volunteers, and their agreement with those obtained with EPI-DWI was evaluated by using the intraclass correlation coefficient. The 3 intravoxel incoherent motion parameters in the anterior pituitary lobe were compared with those in the brain regions by using the Dunnett test. RESULTS: The agreement between TSE-DWI and EPI-DWI was moderate (intraclass correlation coefficient = 0.571) for D, substantial (0.699) for f', but fair (0.405) for D*. D in the anterior pituitary lobe was significantly higher than in the 5 brain regions (P < .001). The f in the anterior pituitary lobe was significantly higher than in the 5 brain regions (P < .001), except for the vermian GM. The pituitary D* was not significantly different from that in the 5 brain regions. CONCLUSIONS: Our results demonstrated the feasibility of intravoxel incoherent motion assessment of the normal anterior pituitary lobe by using TSE-DWI. High D and f values in the anterior pituitary lobe were thought to reflect its microstructural and perfusion characteristics.

Genetic risk factors in Japanese Alzheimer's disease patients: alpha1-ACT, VLDLR, and ApoE.

We studied the polymorphism of alpha1-antichymotrypsin (ACT), very low density lipoprotein receptor (VLDLR) and apolipoprotein E (ApoE) genes in 200 control subjects and 65 patients with Alzheimer's disease (AD) in Japanese. The subjects consisted of 30 patients with early onset familial Alzheimer's disease (FAD), a patient with late onset FAD, 29 patients with an early onset isolated form of AD, and 5 patients with late onset AD. ApoE genotypes were significantly different between controls and FAD (p < 0.0005) or AD (p < 0.05), and patients carrying at least one ApoE epsilon4 allele were found in 44% of FAD and 34.3% of AD; both were significantly different (p < 0.001) from the controls (12.5%). ACT genotypes and allele frequencies were not different among these groups except for genotypes between ApoE epsilon4 FAD and ApoE epsilon4 controls (p = 0.019). There was a slight but significant increase of the 5 repeat allele of VLDLR in AD (p = 0.014), but the difference was rather diminished in the presence of an ApoE epsilon4 allele. None of combinations of ACT and VLDLR genotypes in the presence or absence of an ApoE epsilon4 allele gave significant difference. Thus, we conclude that among the reported genetic risk factors, ApoE epsilon4 is the only definite risk factor for both FAD and AD, and the VLDLR polymorphism might be associated with AD cases in Japanese.

Papain digestion of the subunits of (Na,K)-ATPase.

Membrane-bound (Na,K)-ATPases were exposed to limited papain digestion. We could not find the active (Na,K)-ATPase lacking glycoprotein subunit for the enzymes from three different sources (outer medulla of dog kidney, electric organs of Narke japonica and larvae of Artemia salina). It seemed unlikely that the glycoprotein subunit was selectively removed from (Na,K)-ATPase by papain digestion.

Hereditary motor and sensory neuropathy with myelin folding and juvenile onset glaucoma.

OBJECTIVE: We describe three patients from a family with motor and sensory neuropathy accompanied by open-angle glaucoma. BACKGROUND: Autosomal recessive demyelinating hereditary motor and sensory neuropathies (HMSN) include different disorders. To our knowledge, autosomal recessive HMSN has not been associated with juvenile onset glaucoma. METHODS: Sural nerve pathology of the three patients were examined, and genetic analysis of the family was performed. RESULT: - The most prominent pathologic finding was a highly unusual myelin abnormality consisting of irregular redundant loops and folding of the myelin sheath. The family survey supports autosomal recessive inheritance. The molecular analysis failed to demonstrate either linkage of the disease to MPZ gene, PMP22 gene, Cx32 gene, orEGR2 gene. Analysis did not establish linkage of the disease to the locus of CMT4A, 4B, and 4C genes. CONCLUSION: The present cases may represent a new type of HMSN accompanied by juvenile onset glaucoma.

Large-scale preparation of human thrombin: polyethylene glycol potentiates the factor Xa-mediated activation of prothrombin.

We investigated the ability of polyethylene glycol 4000 to accelerate thrombin generation in a mixture of prothrombin and factor X at concentrations of 1-30%. In the presence of 5 mM of CaCl2, polyethylene glycol 4000 promoted prothrombin activation at concentrations above 1%. The peak of activation was seen at levels of 14 and 20% of polyethylene glycol 4000. The effect of the polyethylene glycol was remarkably dependent on its molecular weight; molecular weights greater than 2000 were required for accelerating thrombin generation. Under optimal conditions, polyethylene glycol 4000, in the presence of CaCl2, promoted conversion of all of the prothrombin into thrombin and its derivatives. We conclude that polyethylene glycol 4000, at concentrations ranging from 14 to 20%, effectively accelerates thrombin generation in the presence of 5 mM of CaCl2. This new method for preparing thrombin is based on the use of polyethylene glycol 4000 and CaCl2 and is applicable to the manufacture of thrombin.

Familial Alzheimer's disease genes in Japanese.

More than 40 missense mutations and a splice-site mutation in the presenilin 1 (PS-1) gene, two missense mutations of presenilin 2 (PS-2), and more than three missense mutations of amyloid precursor protein (APP) cosegregate with early onset familial Alzheimer's disease (FAD). In order to determine the incidence of mutations of these genes in Japanese patients, we screened 25 early onset FAD families, one late-onset FAD case, 33 early onset AD cases and five late-onset AD cases for mutations in the coding regions of the genes using SSCP analysis. Four different missense mutations of the PS-1 gene, including a novel mutation, Glu273Ala, were identified in five early onset FAD families and one missense mutation of PS-1 in one isolated AD patient. While no missense mutations of PS-2 were detected, four silent nucleotide substitutions were observed. Our data indicate that PS-1 mutations account for 20.0% of early onset FAD cases in Japan. Since mutations in PS-2 and APP genes were not found in the remaining cases, which could be explained only partially by apolipoprotein E epsilon4, important FAD genes or risk-factor genes remain to be identified.

Proboscis amputation facilitates the study of mosquito (Diptera: Culicidae) attractants, repellents, and host preference.

Proboscis amputation has facilitated the study of mosquito behavior. Using humans as a host is very important in the study of mosquito attractants, repellents, and host preference. However, mosquito bites cause potential medical problems because of hypersensitivity and perhaps secondary bacterial infection, even using laboratory mosquitoes. Moreover, once a normal female mosquito bites and feeds on human blood, it cannot be used in subsequent probing tests. These problems were resolved by proboscis amputation. Variation of attraction among humans was examined effectively without bites using proboscis-amputated Aedes albopictus Skuse. Proboscis-amputated and normal mosquitoes also showed equal repellency against 1% L-lactic acid. Although the mosquitoes lacked the tip of the labium and some sensilla, they alighted on human forearms in the same way as normal mosquitoes. Because proboscis-amputated mosquitoes continued to probe avidly, they could be used repeatedly, thereby reducing the number of mosquitoes required for experimentation. The use of proboscis-amputated mosquitoes would promote various studies of mosquito attraction or repellency with no risk of hypersensitivity and secondary bacterial infection by mosquito bites.

L-lactic acid as a mosquito (Diptera: Culicidae) repellent on human and mouse skin.

The attraction of Aedes albopictus (Skuse) to hands and forearms of human subjects treated with several concentrations of L-LA solution were studied in a test chamber containing proboscis-amputated mosquitoes. Fewer mosquitoes alighted on L-LA treated human skin than on water-treated control skin. Similar results were found using normal mosquitoes following L-LA and water treatment of mouse skin. The relative repellent effects of L-LA varied with concentration. The minimum repellent concentration was lower than previously reported for human skin. The number of alightments decreased at increasing concentrations of L-LA, demonstrating the absolute repellency of L-LA. Unlike previous reports suggesting that L-LA attracted mosquitoes, our studies using human and mouse skin showed that L-LA exhibited both relative and absolute repellency.

Cyclical transmission of Plasmodium berghei (Coccidiida: plasmodiidae) by Anopheles omorii (Diptera: Culicidae).

Anopheles omorii, a tree-hole breeding anopheline collected in Japan, transmitted a rodent malaria, Plasmodium berghei ANKA strain, in the laboratory. Mice with 0.2-15% parasitemia, 0.01-1.5% gametocytemia, and 0.001-0.5% exflagellations were used as infective hosts. Oocyst numbers ranged from 2 to 171 (mean 44) on the midgut 8-16 d after the blood meal. Several to hundreds of sporozoites were detected in the salivary glands 14 d after feeding. The mosquitoes were infective to mice from 13 to 40 d after feeding. An. omorii occurs naturally at temperatures less than 25 degrees C and therefore is a suitable laboratory vector for the cyclic transmission of P. berghei, a malaria parasite that completes sporogony at 18-21 degrees C.

Production of hydrogen sulfide from tetrathionate by the iron-oxidizing bacterium Thiobacillus ferrooxidans NASF-1.

When incubated under anaerobic conditions, five strains of Thiobacillus ferrooxidans tested produced hydrogen sulfide (H2S) from elemental sulfur at pH 1.5. However, among the strains, T. ferrooxidans NASF-1 and AP19-3 were able to use both elemental sulfur and tetrathionate as electron acceptors for H2S production at pH 1.5. The mechanism of H2S production from tetrathionate was studied with intact cells of strain NASF-1. Strain NASF-1 was unable to use dithionate, trithionate, or pentathionate as an electron acceptor. After 12 h of incubation under anaerobic conditions at 30 degrees C, 1.3 micromol of tetrathionate in the reaction mixture was decomposed, and 0.78 micromol of H2S and 0.6 micromol of trithionate were produced. Thiosulfate and sulfite were not detected in the reaction mixture. From these results, we propose that H2S is produced at pH 1.5 from tetrathionate by T. ferrooxidans NASF-1, via the following two-step reaction, in which AH2 represents an unknown electron donor in NASF-1 cells. Namely, tetrathionate is decomposed by tetrathionate-decomposing enzyme to give trithionate and elemental sulfur (S4O6(2-)-->S3O6(2-) + S(o), Eq. 1), and the elemental sulfur thus produced is reduced by sulfur reductase using electrons from AH2 to give H2S (S(o) + AH2-->H2S + A, Eq. 2). The optimum pH and temperature for H2S production from tetrathionate under argon gas were 1.5 and 30 degrees C, respectively. Under argon gas, the H2S production from tetrathionate stopped after 1 d of incubation, producing a total of 2.5 micromol of H2S/5 mg protein. In contrast, under H2 conditions, H2S production continued for 6 d, producing a total of 10.0 micromol of H2S/5 mg protein. These results suggest that electrons from H2 were used to reduce elemental sulfur produced as an intermediate to give H2S. Potassium cyanide at 0.5 mM slightly inhibited H2S production from tetrathionate, but increased that from elemental sulfur 3-fold. 2,4-Dinitrophenol at 0.05 mM, carbonylcyanide-m-chlorophenyl- hydrazone at 0.01 mM, mercury chloride at 0.05 mM, and sodium selenate at 1.0 mM almost completely inhibited H2S production from tetrathionate, but not from elemental sulfur.

Isolation and some properties of Thiobacillus ferrooxidans strains with differing levels of mercury resistance from natural environments.

Fifty iron-oxidizing bacteria isolated from natural environments were screened for resistance to mercuric ions (Hg2+). Thiobacillus ferrooxidans Funis 2-1, the strain found to show the greatest resistance to Hg2+ among the fifty isolates, gave a cell yield of 7.0 x 10(7) cells/ml after 8 d cultivation in an Fe2+-medium (pH 2.5) containing 0.7 microM Hg2+. Funis 2-1 volatilized 80% of the total mercury added to the medium over 8 d of cultivation. T. ferrooxidans AP19-3, more sensitive to Hg2+ than Funis 2-1, could not grow in an Fe2+-medium (pH 2.5) containing 0.7 microM Hg2+ even over a 28 d cultivation period. When resting cells of strains Funis 2-1 and AP19-3 were incubated for 3 h in a salt solution containing 0.7 microM Hg2+ (pH 3.0), 14.3% and 7.9% of the total mercury added to the reaction mixtures respectively, were volatilized. The activity of the mercuric reductase from Funis 2-1 was only 2.8 times higher than that of the enzyme from AP19-3. Since the markedly higher mercury resistance of Funis 2-1 compared with that of AP19-3 cannot be explained only by the level of the mercuric reductase activity, the levels of mercury resistance of iron oxidase and cytochrome c oxidase were studied. The 1 microM mercuric ions inhibited the 35% of iron-oxidizing activity from AP19-3. In contrast, the same concentration of Hg2+ did not inhibit the activity of iron oxidase from Funis 2-1. In the case of the cytochrome c oxidases purified from both strains, the 0.2 microM Hg2+ inhibited approximately 40% of cytochrome c oxidizing activity from AP19-3, on the contrary, the activity of the enzyme from Funis 2-1 was activated 1.8- and 1.2-fold, respectively, in the presence of 0.08 and 0.2 microM Hg2+. Since cytochrome c oxidase is one of the most important components of the iron-oxidizing system, these results indicate that both the existence of cytochrome c oxidase resistant to Hg2+ as well as that of mercuric reductase in the cells is responsible for the more rapid growth of Funis 2-1 than that of in an Fe2+-medium containing 0.7 microM Hg2+.

Purification and some properties of sulfur reductase from the iron-oxidizing bacterium Thiobacillus ferrooxidans NASF-1.

Thiobacillus ferrooxidans strain NASF-1 grown aerobically in an Fe2+ (3%)-medium produces hydrogen sulfide (H2S) from elemental sulfur under anaerobic conditions with argon gas at pH 7.5. Sulfur reductase, which catalyzes the reduction of elemental sulfur (S0) with NAD(P)H as an electron donor to produce hydrogen sulfide (H2S) under anaerobic conditions, was purified 69-fold after 35-65% ammonium sulfate precipitation and Q-Sepharose FF, Phenyl-Toyopearl 650 ML, and Blue Sepharose FF column chromatography, with a specific activity of 57.6 U (mg protein)(-1). The purified enzyme was quite labile under aerobic conditions, but comparatively stable in the presence of sodium hydrosulfite and under anaerobic conditions, especially under hydrogen gas conditions. The purified enzyme showed both sulfur reductase and hydrogenase activities. Both activities had an optimum pH of 9.0. Sulfur reductase has an apparent molecular weight of 120,000 Da, and is composed of three different subunits (M(r) 54,000 Da (alpha), 36,000 Da (beta), and 35,000 Da (gamma)), as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This is the first report on the purification of sulfur reductase from a mesophilic and obligate chemolithotrophic iron-oxidizing bacterium.

Isolation of iron-oxidizing bacteria from corroded concretes of sewage treatment plants.

Thirty-six strains of iron-oxidizing bacteria were isolated from corroded concrete samples obtained at eight sewage treatment plants in Japan. All of the strains isolated grew autotrophically in ferrous sulfate (3.0%), elemental sulfur (1.0%) and FeS (1.0%) media (pH 1.5). Washed intact cells of the 36 isolates had activities to oxidize both ferrous iron and elemental sulfur. Strain SNA-5, a representative of the isolated strains, was a gram-negative, rod-shaped bacterium (0.5-0.6x0.9-1.5 microm). The mean G+C content of its DNA was 55.9 mol%. The pH and temperature optima for growth were 1.5 and 30 degrees C, and the bacterium had activity to assimilate 14CO2 into the cells when ferrous iron or elemental sulfur was used as a sole source of energy. These results suggest that SNA-5 is Thiobacillus ferrooxidans strain. The pHs and numbers of iron-oxidizing bacteria in corroded concrete samples obtained by boring to depths of 0-1, 1-3, and 3-5 cm below the concrete surface were respectively 1.4, 1.7, and 2.0, and 1.2 x 10(8), 5 x 10(7), and 5 x 10(6) cells/g concrete. The degree of corrosion in the sample obtained nearest to the surface was more severe than in the deeper samples. The findings indicated that the levels of acidification and corrosion of the concrete structure corresponded with the number of iron-oxidizing bacteria in a concrete sample. Sulfuric acid produced by the chemolithoautotrophic sulfur-oxidizing bacterium Thiobacillus thiooxidansis known to induce concrete corrosion. Since not only T. thiooxidans but also T. ferrooxidans can oxidize reduced sulfur compounds and produce sulfuric acid, the results strongly suggest that T. ferrooxidans as well as T. thiooxidans is involved in concrete corrosion.