Functional analysis of the biosynthetic gene cluster required for immunity and secretion of a novel Lactococcus-specific bacteriocin, lactococcin Z.

AIMS: Characterization of the biosynthesis (secretion and immunity) of lactococcin Z. METHODS AND RESULTS: Lactococcin Z is produced by Lactococcus lactisQU 7. DNA sequence analysis revealed that the lactococcin Z gene cluster (c. 5·1 kb) includes four genes encoding putative biosynthetic proteins, LczB (self-immunity protein), LczC (an ABC transporter) and LczD (a transport accessory protein), besides the previously identified LczA. LczB showed 25·5% identity to LciA, the lactococcin A immunity protein, while LczC and LczD had 93·7 and 95·3% identities, respectively, to corresponding proteins of lactococcin A. Heterologous expression of various combinations of the four genes indicated that lczB confers self-immunity against lactococcin Z, and that the four genes are necessary to produce lactococcin Z. However, LczB and LciA showed no cross-immunity to lactococcins A and Z respectively. CONCLUSIONS: The results verified that LczB is the lactococcin Z immunity protein, and LczC is responsible for lactococcin Z secretion in a manner dependent on LczD expression. SIGNIFICANCE AND IMPACT OF THE STUDY: The biosynthesis (secretion and immunity) of a new Lactococcus-specific bacteriocin, lactococcin Z, was characterized. Moreover, the results suggested that lactococcin Z has different immunity and action mechanisms from other Lactococcus-specific bacteriocins.

Identification and characterization of novel multiple bacteriocins produced by Lactobacillus sakei D98.

AIM: To characterize novel multiple bacteriocins produced by Lactobacillus sakei D98. METHODS AND RESULTS: Lactobacillus sakei D98 isolated from Shubo (rice malt) produced at least three bacteriocins. Using three purification steps, three novel antimicrobial peptides termed sakacin D98a, sakacin D98b and sakacin D98c were purified from the culture supernatant. Amino acid and DNA sequencing analysis revealed that the sakacins D98a, D98b and D98c are novel class IIa-like or class IId bacteriocins. In particular, sakacin D98b has a variant pediocin-box sequence, YANGVXC (with Ala instead of Gly), and a different location for the disulfide bridge (Cys(11) and Cys(18)) from that found in other class IIa bacteriocins. CONCLUSIONS: Three novel bacteriocins were identified from Lactobacillus sakei D98. Their antimicrobial spectra and intensities indicate that these sakacins would have different modes of action. In addition, sakacin D98b showed low inhibitory activity against Listeria, probably due to the differences in amino acids and position of the disulfide bridge compared with the other class IIa bacteriocins. SIGNIFICANCE AND IMPACT OF STUDY: Sakacins D98a and D98c are novel bacteriocins belonging to class IId bacteriocins. On the other hand, sakacin D98b, a class IIa-like bacteriocin, has a unique internal structure and activity spectrum.

Search for the Θ+ pentaquark via the π(-)p→K(-)X reaction at 1.92 GeV/c.

The Θ(+) pentaquark baryon was searched for via the π(-)p→K(-)X reaction with a missing mass resolution of 1.4 MeV/c(2) (FWHM) at the Japan Proton Accelerator Research Complex (J-PARC). π(-) meson beams were incident on the liquid hydrogen target with a beam momentum of 1.92 GeV/c. No peak structure corresponding to the Θ(+) mass was observed. The upper limit of the production cross section averaged over the scattering angle of 2° to 15° in the laboratory frame is obtained to be 0.26 µb/sr in the mass region of 1.51-1.55 GeV/c(2). The upper limit of the Θ(+) decay width is obtained to be 0.72 and 3.1 MeV for J(Θ)(P)=1/2(+) and J(Θ)(P)=1/2(-), respectively, using the effective Lagrangian approach.

Stereotactic radiotherapy for hepatocellular carcinoma induced by hepatitis C and the relationships of changes in carbohydrate antigen 19-9 with AFP and PIVKA-II.

PURPOSE: Assessing the therapeutic effects of stereotactic body radiotherapy (SBRT) for hepatocellular carcinoma (HCC) takes time. Purpose of our study was to explore the relationships of changes in carbohydrate antigen 19-9 (CA 19-9) with those in the existing markers alpha-fetoprotein (AFP) and protein induced by vitamin K absence or antagonist-II (PIVKA-II). PATIENTS AND METHODS: The subjects were 16 patients who underwent SBRT for solitary HCC ≤3cm induced by hepatitis C between June 2016 and July 2019. Observation periods ranged from 8-43 (median: 28) months, ages from 59-85 (median: 65) years. RESULTS: Changes in CA 19-9 levels after SBRT were categorised into three patterns: 1) a transient elevation followed by a decline (75%); 2) a transient decline followed by an elevation (18.8%); and 3) no change (6.3%). Among patients showing a transient CA 19-9 elevation followed by a decline, which was the most frequent pattern, 75% showed these changes in synchronisation with AFP and preceded the changes in PIVKA-II, while in the other 25%, CA 19-9 changes were in synchronisation with PIVKA-II and preceded those in AFP. At the time of recurrence, 62.5% showed a continuous CA 19-9 elevation, either in synchronisation with other markers or by itself. CONCLUSIONS: This is the first investigation of changes in CA 19-9 levels after SBRT for HCC induced by hepatitis C. Characteristic changes in CA 19-9, AFP, and PIVKA-II levels were observed as responses after treatment. As for its correlations with tumour markers, the acute responses of PIVKA-II tended to be slower than those of CA 19-9 and AFP. Although the sample size was small, our findings raise the possibility that measuring these 3 biomarkers after SBRT may be useful for monitoring patients for HCC recurrence.

Bombyx mori Ras proteins BmRas1, BmRas2 and BmRas3 are neither farnesylated nor palmitoylated but are geranylgeranylated.

The lipid modifications which occur on Bombyx mori Ras proteins BmRas1, BmRas2 and BmRas3 were studied by metabolic labelling in an insect cell-free protein synthesis system and in a baculovirus expression system, using specific inhibitors of protein prenylation and protein palmitoylation. In addition, the subcellular localization of BmRas proteins was examined using EGFP fusion proteins of constitutively active forms of BmRas proteins transiently expressed in Sf9 cells. As a result, it was revealed that the three B. mori Ras proteins BmRas1, BmRas2 and BmRas3 are neither farnesylated nor palmitoylated but are geranylgeranylated for localization to the plasma membrane of insect cells. Thus, the mechanism of membrane binding of insect Ras proteins is quite different from that reported for mammalian Ras proteins.

Oral administration of Bifidobacterium longum prevents gut-derived Pseudomonas aeruginosa sepsis in mice.

AIMS: The aim of the study was to evaluate the efficacy of probiotics on gut-derived sepsis caused by Pseudomonas aeruginosa in immunocompromised mice. METHODS AND RESULTS: After oral inoculation of P. aeruginosa, mice were treated with cyclophosphamide to induce leucopenia and translocation of the intestinal P. aeruginosa into blood, thereby producing gut-derived sepsis. In this model, administration of 1 x 10(9) CFU of Bifidobacterium longum strain BB536 for 10 days significantly (P < 0.01) increased the survival rate compared with groups of mice administered either with Bifidobacterium breve strain ATCC 15700 or excipients contained in the probiotic bacterial powder. Administration of B. longum significantly decreased viable counts of P. aeruginosa in the liver and blood compared with other groups. Culture of intestinal contents revealed a significantly lower viable count of P. aeruginosa in the jejunum of B. longum-treated mice compared with other groups of mice. Furthermore, in vitro data demonstrated that B. longum possessed apparently higher adherent activity to Caco-2 cell monolayers and significantly suppressed the adherence of P. aeruginosa to the monolayers of cells compared with other groups. CONCLUSION: Oral administration of B. longum protects mice against gut-derived sepsis caused by P. aeruginosa, and the effect may be due to interference of P. aeruginosa adherence to intestinal epithelial cells. SIGNIFICANCE AND IMPACT OF THIS STUDY: This study demonstrated that oral administration of B. longum BB536 is effective to protect against opportunistic infection with drug-resistant bacteria such as P. aeruginosa. The results suggest that probiotics may play an important role even in the immunocompromised patients.

Metronomic chemotherapy using weekly low-dosage CPT-11 and UFT as postoperative adjuvant therapy in colorectal cancer at high risk to recurrence.

This study was designed to evaluate the antitumor efficacy and feasibility of postoperative adjuvant metronomic chemotherapy using weekly low-dosage CPT-11 and UFT in colorectal cancer at high risk to recurrence. A total of 49 patients (24 stage IIIb and 25 distant metastasis) who underwent a R0 operation were enrolled in this prospective study. Forty mg/m2 of CPT-11 were administered on day 1, day 8, and on day 15 in 28-day cycles. A dosage of 335 mg/m2/day of UFT was given perorally on daily schedule. Cycles were repeated for 6 months, and were followed by UFT alone for further 6 months. One or more adverse effects were seen in 43 of the 49 patients. However, most of these effects were mild at grade 1 or 2: with only nausea in 3 patients, vomiting in 2, leucopenia in 2 and neutropenia in 2 at grade 3. The overall survival rates were favorable both in the stage IIIb group (5-year: 73%) and in the distant metastases group (5-year: 62%). Postoperative adjuvant metronomic chemotherapy using weekly low-dosage CPT-11 and UFT might be safe and feasible and prolong survival time in colorectal cancer at high risk to recurrence.

Patient outcomes of whole brain radiotherapy for brain metastases versus leptomeningeal metastases: A retrospective study.

AIM: Important factors typically associated with prognosis in brain metastases include Karnofsky performance status (KPS), extracranial or cerebellar localization and combination chemotherapy. However, few studies investigated the prognostic role of leptomeningeal metastases (LM) following whole brain radiotherapy (WBRT). On the basis of our experience suggesting better survival of asymptomatic patients with LM than those with brain metastases, we herein evaluated LM as a prognostic factor after WBRT. METHODS: Medical records of 206 patients (median age, 65 years) who received WBRT in 2007-2015 were retrospectively reviewed. The two most common cancers were of lung, breast origin in 78.5%, 10%, patients, respectively. Patients received parallel-opposed WBRT, with a dose of 20-40 Gy. Additional doses of 9-12 Gy were used in patients who were operated on or had single metastases. Overall survival (OS) was determined, and clinical parameters including age, KPS, symptoms, radiation dose, dose per fraction, type of metastasis, extracranial metastases, primary status and surgery plus WBRT were assessed as prognostic factors. RESULTS: The median survival was 6 months (range, 1-100), and 1- and 2-year survival rates were 28% and 17%, respectively. In univariate analysis, improved survival was associated with KPS of ≥70, absence of symptoms, radiation dose of ≥37.5 Gy, favorable primary lesion, LM, and surgery plus WBRT. Multivariate analysis revealed that these factors with the exception of radiation dose was significant prognostic factors for OS. CONCLUSION: We found that LM were independent prognostic factors for good clinical outcomes.

Coiling is Not Essential to Anhydrobiosis by Aphelenchus avenae on Agar Amended with Sucrose.

The roles of preconditioning and coiling upon entrance into anhydrobiosis by Aphelenchus avenae were tested via video-assisted analysis at 25(2) degrees C. Fourth-stage juveniles or young adults of A. avenae were individually placed on 5% agar containing 0.8 M sucrose. Nematodes became quiescent within 3 hr, then gradually resumed a low level of activity and assumed a coiled posture. High desiccation survival rate was recorded when nematodes were incubated on agar for more than 6 hr; the survival rates were 0%, 3%, 73%, and 92% for 0, 2, 6, and 12 hr on agar, respectively. All nematodes placed on agar for 24 hr or more revived after rehydration following desiccation. Once nematodes were on agar for a sufficient time, no difference in desiccation survival was observed between nematodes taking a coiled posture and those uncoiled artificially. Based on these results, exposure to osmotic stress for 6 hr can prepare A. aveae physiologically for anhydrobiosis, but coiling does not appear to be a physiological requirement for desiccation in survival.

Molecular mechanism by which acyclic retinoid induces nuclear localization of transglutaminase 2 in human hepatocellular carcinoma cells.

Nuclear accumulation of transglutaminase 2 (TG2) is an important step in TG2-dependent cell death. However, the underlying molecular mechanisms for nuclear translocation of TG2 are still poorly understood. In this study, we demonstrated that acyclic retinoid (ACR) induced nuclear accumulation of TG2 in JHH-7 cells, a hepatocellular carcinoma (HCC) leading to their apoptosis. We further demonstrated molecular mechanism in nuclear-cytoplasmic trafficking of TG2 and an effect of ACR on it. We identified a novel 14-amino acid nuclear localization signal (NLS) (466)AEKEETGMAMRIRV(479) in the 'C' domain and a leucine-rich nuclear export signal (NES) (657)LHMGLHKL(664) in the 'D' domain that allowed TG2 to shuttle between the nuclear and cytosolic milieu. Increased nuclear import of GAPDH myc-HIS fused with the identified NLS was observed, confirming its nuclear import ability. Leptomycin B, an inhibitor of exportin-1 as well as point mutation of all leucine residues to glutamine residues in the NES of TG2 demolished its nuclear export. TG2 formed a trimeric complex with importin-α and importin-ß independently from transamidase activity which strongly suggested the involvement of a NLS-based translocation of TG2 to the nucleus. ACR accelerated the formation of the trimeric complex and that may be at least in part responsible for enhanced nuclear localization of TG2 in HCC cells treated with ACR.

Probiotics and safety.

Bacterial species that have traditionally been regarded as safe are used in probiotics; the main strains used include lactic acid bacteria and bifidobacteria that inhabit the intestinal tracts of humans and animals. However, reports of frequent isolation of bacteria used in probiotics from infection sources in recent years have raised much debate over the safety of probiotics. This article describes the status quo of isolation of probiotic bacteria from infections and reviews each of the factors that have to be addressed in assessing the safety of probiotics, namely pathogenicity, infectivity, toxicity, and intrinsic properties of the bacteria. Monoassociation with Bifidobacterium longum in gnotobiotic mice as a method to assess safety with respect to infection, and translocation and immune responses as a result of the monoassociation are also described.

BAYw6228 suppresses accumulation of macrophages in balloon-induced intimal thickening of rabbit carotid artery.

To determine the in vivo activity of BAYw6228 (BAYw), a newly developed 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase inhibitor, we examined its effect on balloon catheterization-induced carotid arterial intimal thickening in Japanese white rabbits. In all rabbits weighing approximately 3.5 kg, a balloon catheter was inserted from a peripheral branch of the left external carotid artery to the aorta in order to denude the endothelium of the left common carotid artery. At day 12, they were divided into control and BAYw groups. The latter were subcutaneously injected with BAYw, 1 mg/kg/day, for the following 17 days. At day 15, the second balloon catheterization was performed in the same left common carotid artery in both groups. After another two weeks, the left common carotid artery was removed and stained with Elastica-Masson (EM) and anti-rabbit macrophage antibody (RAM 11). With RAM 11 staining, the number of positive cells in the intimal thickening was markedly decreased in the BAYw group compared with the control group (P < 0.01). Compared with the area ratio of the thickened intima/media layer of the control group, that of the BAYw group was significantly decreased to 45% (P < 0.05). These results indicate that BAYw prevents accumulation of macrophages and consequently contributes to the inhibition of intimal thickening induced by balloon catheterization.

Suppression of neointimal thickening by a newly developed HMG-CoA reductase inhibitor, BAYw6228, and its inhibitory effect on vascular smooth muscle cell growth.

1. The aim of this study was to determine whether BAYw6228 (BAYw), a newly developed 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase inhibitor, could suppress an atherogenic process such as intimal thickening by a mechanism other than lowering the level of serum cholesterol. 2. First, we evaluated the in vitro effect of BAYw on the proliferation of vascular smooth muscle cells (SMC) from various species: Sprague-Dawley (SD) rats. New Zealand (NZ) white rabbits, intimal cells from Watanabe hereditary hyperlipidemic (WHHL) rabbit and SMC from the new-born human aorta. The increasing rate of total protein content of these cells was inhibited by the addition of BAYw in a dose-dependent fashion. In the presence of 2% foetal calf serum (FCS), the value of IC50 was 1.0 microM in SD rats, 2.1 microM in NZ white rabbits, and 0.3 microM in WHHL rabbits. With human SMC, the value was 0.02 microM in the presence of 10% FCS and 0.2 microM with a mixture of growth factors. 3. Based on these above in vitro findings, we next examined the in vivo effect of the agent to determine whether it could suppress rabbit intimal thickening induced by balloon catheterization. A balloon catheter was inserted from a peripheral branch of the left external carotid artery to the aorta to denude the endothelium of the left common carotid artery in Japanese white rabbits. After 12 days they were divided into control and BAYw groups. The former were subcutaneously injected with saline and the latter with BAYw 1 mg kg-1 day-1. Two days after the beginning of treatment, a second balloon injury was performed to the previously injured left common carotid artery in both groups. After another two weeks, the left common carotid artery was removed and variously stained. Although the total serum cholesterol in the BAYw group was significantly lower than in the control (P < 0.05), the difference was not enough to affect intimal thickening. In addition, the BAYw group had a smaller intima/media ratio than the control group, decreasing to 45% of control (P < 0.05). By anti-alpha smooth muscle actin antibody staining, these intimal thickening areas were entirely occupied by SMCs, and their amount was attenuated by BAYw. By anti-rabbit macrophage antibody (RAM 11) staining, the number of positive cells in the intimal thickening was markedly decreased in the BAYw group compared to control (P < 0.01). 4. These results indicate that BAYw has an inhibitory effect on intimal thickening by attenuating intimal SMC proliferation and infiltration of macrophages, suggesting that BAYw could be effective in the prevention of the progression of atherosclerotic plaque-like restenosis after angioplasty.

Ulcerative colitis and colorectal cancer: a follow-up study in Fukuoka, Japan.

BACKGROUND: Our goal was to study the higher death rate and the causes of such deaths among ulcerative colitis (UC) patients in the Japanese population, and to compare our findings in such cases with those for Crohn's disease (CD). METHODS: In all, 174 UC (male/female: 54/120) and 66 CD (34/32) patients who were registered for the research promotion programme in Fukuoka prefecture (1971-1981) were traced up to the end of 1994. The standardized mortality ratios (SMR) were calculated based on the death rates of the Japanese population by age, sex and calendar year. RESULTS: The overall follow-up rate was 96.7%. Among the UC patients, the SMR for all causes were 0.84 (95% CI: 0.11-4.31) for men; 1.05 (95% CI: 0.08-4.69) for women; and 0.94 (95% CI :0.09-4.50) for both sexes combined. When excluding deaths due to colorectal cancer, the SMR for the same groups were 0.43, 0.94 and 0.67, respectively. The SMR for both sexes were 1.82 (95% CI: 0.17-5.96) for malignant neoplasms and 9.93 (95% CI: 4.67-17.3) for colorectal cancer. Patients who died from colorectal cancer showed onset at a younger age (mean: 25.5 years) as well as a longer disease course of UC (mean: 17.0 years). Regarding the CD patients, the SMR for all causes were 1.75 (95% CI: 0.15-5.75) for both sexes. Most deaths were caused by gastrointestinal complications. CONCLUSIONS: An excess mortality from colorectal cancers was indicated in the UC patients, especially in males. The overall SMR in male UC patients decreased by 50% when the deaths from colorectal cancer were excluded. The excess mortality in those with CD over UC patients was attributed to gastrointestinal complications rather than malignant diseases. Some carcinogenic factors therefore seem most likely to exist in the pathogenesis of UC.

Glutamine and arginine metabolism in tumor-bearing rats receiving total parenteral nutrition.

Arginine supplementation increases glutamine levels in muscle and plasma. Since glutamine production is increased in catabolic states, these observations prompted us to investigate whether the flux of arginine to glutamine was increased in tumor-bearing (TB) rats, and we measured the synthesis rate of glutamine from arginine in control versus TB rats receiving standard total parenteral nutrition (TPN) solution. Male Donryu rats (N = 36; body weight, 200 to 225 g) were divided into two groups, control and TB rats. Yoshida sarcoma cells (1 x 10(6)) were inoculated into the back of the rats (n = 18) subcutaneously on day 0. The rats were given free access to water and rat chow. On day 5, all animals, including non-TB rats (n = 18), were catheterized at the jugular vein and TPN was begun. On day 10, TPN solution containing either U-14C-glutamine (2.0 microCi/h) or U-14C-arginine (2.0 microCi/h) was infused as a 6-hour constant infusion. At the end of the isotope infusion, plasma was collected to determine the glutamine production rate in rats receiving U-14C-glutamine, and the ratio of specific activity of glutamine to specific activity of arginine was measured in rats receiving U-14C-arginine. Only 2 g tumor caused a decrease in glutamine levels and an increase in glutamine and arginine production. The low flux rate of arginine to glutamine was observed in control rats (Arg to Gln, 41.0 +/- 11.9 mumol/kg/h). On the other hand, TB caused a significant increase in Arg to Gln compared with the control (213.3 +/- 66.1 mumol/kg/h, P < .01 v control). An increase in the flux rate of Arg to Gln was associated with an enhancement in the ratio of specific activity of ornithine to specific activity of arginine in TB rats (control 51.5% +/- 10.9% v 77.4% +/- 8.9%, P < .05). We conclude that (1) glutamine and arginine metabolism is altered with very small tumors, (2) although the flux of Arg to Gln was increased in TB and rats, the small increase in Arg to Gln cannot explain the observed large increase in Gln production.

Potentiometric butyrylcholine sensor for organophosphate pesticides.

A highly sensitive potentiometric butyrylcholine (BuCh) sensor based on plasticized poly(vinyl chloride) membrane was fabricated using tetrakis (3,5-bis[2-methoxy-hexafluoro-methyl] phenyl) borate (HFPB) as a cation exchanger. The sensor showed a Nernstian response from 10(-1) M to 10(-6) M for BuCh. The detection limit of the sensor was 1.8 x 10(-7) M. The potential stability, lifetime and detection limit of the BuCh-sensor were improved in comparison with corresponding parameters of BuCh-sensors using prepared other cation exchangers. This improvement was proved to be due to higher hydrophobicity of the cation exchanger, HFPB. Enzyme activity of butyrylcholinesterase (BuChE) was determined by the BuCh-sensor fabricated. Since the selectivity coefficient of the BuCh-sensor for BuCh against a product of the enzyme reaction, choline, was as low as 6.3 x 10(-3), the sensor was not affected by interference from choline even when the enzyme reaction proceeded up to about 80%. Michaelis parameters for the BuChE reaction were obtained by the sensor. Determinations of organophosphate pesticides, 2,2-dichlorovinyldimethylphosphate and o-(4-bromo-2-chlorophenyl)o-ethyl S-propylphosphothiolate) were conducted by measuring inhibition of enzyme activity. The enzyme reaction rate was related to the concentration of pesticides. Pesticides were successfully determined between micromole and sub-nanomole levels by the BuCh-sensor.

Steroidal glycosides from the bulbs of Lilium dauricum.

The bulbs of Lilium dauricum yielded 11 compounds, including six new steroidal glycosides. The structures have been determined by spectral analysis and hydrolysis to be (25R,26R)-26-methoxyspirost-5-en-3 beta-ol 3-O-alpha-L-rhamnopyranosyl-(1----2)-O-[alpha-L-arabinopyranosyl-( 1----3)]- beta-D-glucopyranoside, (25R,26R)-26-methoxyspirost-5-en-3 beta-ol 3-O-alpha-L-rhamnopyranosyl-(1----2)-O-[beta-D-glucopyranosyl-(1----4)]- beta-D-glucopyranoside, (25R)-spirost-5-en-3 beta-ol (diosgenin) 3-O-alpha-L-rhamnopyranosyl-(1----2)-O-[alpha-L-arabinopyranosyl- (1----3)]-beta-D-glucopyranoside, (25R)-3 beta,17 alpha-dihydroxy-5 alpha-spirostan-6-one 3-O-alpha-L-rhamnopyranosyl-(1----2)-beta-D-glucopyranoside, (25R)-3 beta, 17 alpha-dihydroxy-5 alpha-spirostan-6-one 3-O-alpha-L-rhamnopyranosyl-(1----2)-O-[alpha-L- arabinopyranosyl-(1----3)]-beta-D-glucopyranoside and (20R,22R)-3 beta,20,22-trihydroxy-5 alpha-cholestan-6-one (tenuifoliol) 3-O-alpha-L-rhamnopyranosyl-(1----2)-beta-D-glucopyranoside. The absolute configurations of C-20 and C-22 of tenuifoliol were further confirmed by detailed analysis of the NOE difference spectrum of the corresponding isopropylidene derivative. Several known compounds were also isolated and identified.

Identification of bifidobacteria from dairy products and evaluation of a microplate hybridization method.

Sixteen strains of Bifidobacterium isolated from 15 dairy products such as yogurt, cultured milk, butter and cheese were characterized on the basis of phenotypic characteristics and DNA similarities were examined by a microplate hybridization method. Three of the strains were identified as Bifidodobacterium longum, one strain was identified as Bifidobacterium bifidm, and one strain was assigned to the species Bifidobacterium breve on the basis of phenotypic characteristics, and this identification was confirmed by the analysis of DNA similarities. The remaining 11 strains could not be identified by examining their phenotypic characteristics and, contrary to the product label information, these strains were identified as Bifudidobacterium animalis on the basis of DNA similarities. The applicability of the colorimetric hybridization method in micro dilution wells to genetic identification of Bifidobacterium species was also studied.

Glutamine supplementation in cancer patients.

OBJECTIVES: Three series of studies investigated whether 1) glutamine deficiency occurs in tumor-bearing rats, 2) glutamine supplementation improves protein metabolism during chemotherapy in tumor-bearing rats, and 3) oral glutamine supplement improves systemic immune and gut-barrier function in patients with esophageal cancer receiving radiochemotherapy. METHODS: In the animal studies, AH109A hepatoma cells or Yoshida sarcoma cells were inoculated into male Donryu rats to induce tumors. Glutamine production was measured by U-14C-glutamine infusion and the conversion of arginine to glutamine was measured by infusion of U-14C-arginine. The effect of glutamine on protein metabolism was investigated by 1-14C-leucine infusion. In the clinical study, 13 patients with esophageal cancer were randomized into two groups, control and glutamine supplemented (30 g/d), for 4 wk. RESULTS: Glutamine levels in plasma and skeletal muscle were decreased in tumor-bearing rats, although glutamine production and the conversion of arginine to glutamine were increased. Glutamine-supplemented total parenteral nutrition reduced whole-body protein breakdown rate during chemotherapy in tumor-bearing rats. Oral supplementation of glutamine to the patients with esophageal cancer enhanced lymphocyte mitogenic function and reduced permeability of the gut during radiochemotherapy. CONCLUSIONS: Glutamine depletion in host tissues occurs in tumor-bearing rats. Glutamine supplementation can attenuate loss of protein in the muscle in tumor-bearing animals and protect immune and gut-barrier function during radiochemotherapy in patients with advanced cancer.

Effect of glutamine supplementation on protein metabolism and glutathione in tumor-bearing rats.

BACKGROUND: Since tumor-bearing rats are deficient in glutamine, we investigated whether (1) glutamine and glutathione deficiency occur in tumor-bearing rats, (2) glutamine supplementation caused an increase of glutathione levels in host tissues and tumor, (3) glutamine enhances protein synthesis in host tissues, and (4) glutamine stimulated the tumor to synthesize protein and DNA. METHODS: Male Donryu rats were randomized into four groups: (1) non-tumor-bearing rat (NTB) + standard total parenteral nutrition (STPN); (2) NTB + glutamine-supplemented TPN (GTPN); (3) tumor-bearing rat (TB) + STPN; (4) TB + GTPN. On day 0 AH109A rat hepatoma cells were subcutaneously injected into the backs of rats to induce tumor. The animals were maintained on TPN for 6 days from day 10 through day 15. On day 15, 1-14C-leucine was given by a 5-hour continuous infusion (2.0 microCi/h per rat) to determine the fractional synthesis rate and endogenous leucine production. The levels of glutamine and glutathione were measured by HPLC. the tumor DNA synthesis was estimated by bromodeoxyuridine labeling index. RESULTS: Tumor development led to a significant weight loss, but this weight loss was significantly lessened by glutamine supplementation because of an increase in muscle protein synthesis. Glutamine did not enhance tumor weight, protein, and DNA synthesis in the tumor. Tumor development caused a significant reduction of glutathione in the muscle, jejunum, and liver, but supplemented glutamine increased the levels of glutathione in the jejunum. CONCLUSION: Glutamine supplementation is beneficial in preventing deficiencies of glutamine and glutathione and in improving protein metabolism in tumor-bearing rats.