Distributional uptake of gamma-globulin in small intestine of neonatal calves.

Newborn calves deprived of colostrum were used to determine distribution of uptake of gamma-globulin labeled with iodine-125 in small intestine. Ten calves less than 12.5 h of age (average 7 h) were anesthetized, and intestines were exteriorized through an abdominal incision. Intestine was ligated into 10-cm segments, 70 cm apart, beginning at the ileocecal junction and progressing anteriorally, then each segment injected with 100 mg (appoximately 1 microCi) labeled gamma-globulin in 5 ml electrolyte solution, and incubated for 1.5 h. One additional segment was formed adjacent to segments 1, 5, and 10 to assess uptake after .5 h incubation with [iodine-125] gamma-globulin. After prescribed gamma-globulin exposure, segments were excised; then volume of lumen contents, segment weight, and tissue activity were measured. The location of each segment was the percentage of distance from cecum to abomasum. Uptake was milligrams gamma-globulin per gram of segment tissue. Distribution of gamma-globulin uptake after 1.5 h incubation was a cubic function of segment position. Uptake was greatest in a region 15% of the cecum-abomasum distance and declined progressively toward the abomasum. After .5-h incubation with gamma-globulin, regression of uptake on segment position was a quadratic function with greatest uptake at 30% of cecum-abomasum distance. Uptake was greater in segments exposed to gamma-globulin for 1.5 h than .5 h.     

Endogenous production of immunoglobulin IgG1 in newborn calves

There is a decrease in the specific activity of labeled IgG1 of serum over 3 wk following the feeding of iodine-125 labeled immunoglobulin IgG1 in colostrum to calves at birth. This decrease indicated the appearance of new IgG1 from some source. To determine if this new IgG1 came from endogenous production in the calf or from continued small amount of intestinal absorption from milk, labeled IgG1 was added to normal milk and fed to calves of various ages up to 3 wk after an initial feeding of colostrum at birth. Labeled IgG1 was also added to colostrum fed to calves at birth, and the calves were maintained on a normal milk diet or fed a synthetic milk diet. Determination of iodine-125 in the serum protein fractions of these calves indicated that there was no apparent intestinal absorption of labeled IgG1 from the milk in the period from 2 days to 3 wk. Furthermore, comparable decreases occurred in the specific activity of labeled IgG1 in serum in the calves fed the labeled IgG1 in colostrum at birth and subsequently maintained either on a diet including milk or on the synthetic milk diet devoid of IgG1. The results support the conclusion that the origin of new IgG1 in the calf after about 36 h and up to about 3 wk of age arises from endogenous production at a rate of about 1 g of IgG1 per day.     

Effect of orally administered duodenal fluid on serum proteins in neonatal calves.

This study was to determine if orally administered duodenal fluid, as a source of intestinal microorganisms, would influence absorption of gamma-globulin of colostrum in newborn calves. Duodenal fluid was obtained 2 h postfeeding from a cannulated milk-fed calf. Twenty-seven male and female Holstein calves obtained within 6 h of birth were assigned randomly to one of three treatments: 1) colostrum alone, 2) 200 ml duodenal fluid immediately followed by colostrum, and 3) 200 ml duodenal fluid followed 3 h later by colostrum. Pooled colostrum was fed at 10% of body weight in two daily feedings. Total protein, albumin, alpha-globulin, beta-globulin, and gamma-globulin of blood serum were determined prior to colostrum consumption, and again 24 and 48 h after administration of duodenal fluid. Total proteins, beta-globulin, and gamma-globulin of serum increased with age in all calves. Inocula interference with absorption was indicated by depressed values of serum in calves of treatment 3 as compared to treatment 1 calves for protein (5.28 to 6.28 g/dl), beta-globulin (.67 to .87 g/dl), and gamma-globulin (.76 to 1.33 g/dl). Protein was also lower in calves of treatment 3 (6.05 g/dl) compared to uninoculated calves (6.28 g/dl). Malabsorption of colostral proteins may be related to early establishment of large numbers of intestinal microorganisms in the newborn calf.     

4株乳酸菌对模拟胃肠环境的耐受性及生长特性研究

对4株来源于自然发酵肉制品中的乳酸菌进行耐酸耐胆盐性能的测定,选取在pH3.0和含0.3 g/100 mL胆盐的MRS培养基中存活率均超过50%的2株菌（M12、M23）,进一步研究其在模拟胃肠液中的耐受性及MRS培养基中的生长特性。结果表明,这2株乳酸菌对人工模拟胃肠道环境有较好的耐受性,经过人工模拟胃液处理2 h后,存活率均＞50%,经人工模拟肠液处理10 h后仍可存活;这2株菌具有较强的增殖及产酸能力,24 h内均已进入稳定生长期,随着菌体数目的增加,培养基pH值快速降低,在肉制品的发酵温度范围内均能较好地生长。因此,这2株菌可作为潜在的益生菌菌株应用于肉制品的发酵。     

Production of free conjugated linoleic acid by Lactobacillus acidophilus and Lactobacillus casei of human intestinal origin

A gas chromatographic procedure was used for analysis of conjugated linoleic acid (CLA) isomers cis-9, trans-11-octadecadienoic; trans-10, cis-12 octadecadienoic; and trans-9, trans-11-octadecadienoic (c9t11, t10c12, t9t11) produced by lactobacilli. Four different cultures, two strains each of Lactobacillus acidophilus and Lactobacillus casei were tested for their ability to produce CLA from free linoleic acid in MRS broth supplemented with linoleic acid. Different concentrations of linoleic acid (0, 0.05, 0.1, 0.2 and 0.5 mg/ml) were added to MRS broth, inoculated with the lactobacilli, and incubated at 37 degrees C. Viable counts and amounts of individual isomers of CLA (c9t11, t10c12, t9t11) were measured at 0, 24, 48, and 72 h. All the cultures were able to produce free CLA in media supplemented with linoleic acid. Maximum production of CLA (80.14 to 131.63 microg/ml) was observed at 24 h of incubation in broth containing 0.02% of free linoleic acid. No significant (P > 0.05) increases in total CLA levels were observed after 24 h of incubation. The ability of the cultures to produce CLA in skim milk supplemented with 0.02% free linoleic acid also was studied. In this medium, the total amounts of free CLA after 24 h of incubation ranged from 54.31 to 116.53 microg/ml. The use of lactic acid bacteria able to form free CLA in cultured dairy products may have potential health or nutritional benefits. Free CLA in the products likely would be more readily available for absorption from the digestive tract than if it were incorporated into the cells of the starter culture.     

Novel quantitative assays for estimating the antimicrobial activity of fresh garlic juice

Novel agar diffusion and broth dilution assays were developed for quantitatively estimating the antimicrobial activity of fresh garlic juice. Bacteria found to be inhibited by garlic juice in agar diffusion assay included two gram-positive and five gram-negative species. Leuconostoc mesenteroides was not inhibited. Escherichia coli B-103 (HB101, with pJH101, ampicillin resistant, 100 microg ml(-1)) was inhibited and chosen as the standard culture for quantitative assays. The agar diffusion assay was based on the slope ratio method, where the slope of dose response for garlic juice was divided by the slope of dose response for methylmethane thiosulfonate (MMTSO2). Juice from fresh garlic varied in activity between 1.76 and 2.31 microg of MMTSO2 per mg of garlic juice. The activity of juice decreased during 11 months of storage of garlic cloves at 5 degrees C from 2.31 to less than 0.1 microg of MMTSO2 per mg of juice. The broth dilution assay also used the E. coli B-103 culture, which permitted selective enumeration of this bacterium when 100 microg ml(-1) of ampicillin was incorporated into the enumerating agar. Selective enumeration was essential since the garlic juice was not sterile and, thus, contained natural flora. Growth of E. coli was unaffected by 0.1%, delayed by 0.25%, and completely inhibited at 0.5 and 2% garlic juice in broth during 24 h of incubation at 37 micro C. The minimum inhibition concentration of garlic juice by broth dilution assay was, thus, estimated to be 0.5%, which is equivalent to 3.46 microg of MMTSO2 per mg of garlic juice by the agar diffusion assay.     

Evaluation of culture enrichment procedures for use with Salmonella detection immunoassay

To design efficient culture strategies for use with immunoassays to detect Salmonella in food, the growth of these organisms was investigated according to the Bacteriological Analytical Manual (BAM) and enrichment-immunoassay (EI) culture procedures. The cultures were further evaluated using a commercial enzyme-linked immunosorbent assay (ELISA) kit. The BAM procedure includes pre-enrichment in nutrient broth (NB) for 16 h followed by selective enrichment in either Rappaport-Vassiliadis (RV) or tetrathionate brilliant green (TBG) broth for 16 h. The EI procedure includes pre-enrichment in NB for 4 h, selective enrichment in RV for 16 h and post-enrichment in NB for 4 h. The effects of different incubation times for pre- and post-enrichment, and different culture media for selective enrichment (TBG and RV) and post-enrichment in NB and Brain Heart Infusion broth (BHI) on the growth of the bacteria and ELISA titers in the EI procedure were also investigated. Salmonella enteritidis and S. typhimurium inoculated at different initial concentrations between 0.1 and 35 CFU/ml grew to similar concentrations of 10(7) to 10(8) colony forming unit (CFU)/ml in pure culture and generally 2 to 4 fold lower concentrations (P<0.05) in mixed culture using spiked chicken rinse. In the BAM procedure, the concentration of Salmonella cultured in RV was higher (P<0.01) than that in TBG. The cultures in TBG showed positive results for ELISA, but those in RV were generally negative. In the EI procedure, the ELISA titers from cultures post-enriched in NB or BHI were higher (P<0.01) when TBG, as compared to RV, was used for selective enrichment. Post-enrichment in BHI yielded higher numbers of Salmonella and higher ELISA titers than those in NB (P<0.05) for post-enrichment. This study demonstrated that in both culture procedures small numbers of Salmonella could be increased to at least 10(7) CFU/ml which is detectable by most ELISAs, and that the type of the culture media used may have a significant impact on ELISA results.     

辣椒酱发酵菌肠膜明串珠菌C27高密度培养条件优化

为了实现辣椒酱发酵肠膜明串珠菌（Leuconostoc mesenteroides）C27的高密度培养，以MRS肉汤培养基为基础，L. mesenteroides C27的菌体密度为评价指标，采用单因素试验和响应面法对培养基中的碳源、氮源、生长因子进行优化，同时采用响应面法对培养条件进行优化。结果表明，最佳培养基配方为蔗糖21 g/L，酵母浸粉22 g/L，土豆汁14 g/L；最佳培养条件为发酵温度38.4 ℃、初始pH值6.2，接种量2.4%。在此优化条件下培养24 h，L. mesenteroides C27的菌体密度（OD600 nm值）达1.034，活菌数为1.30×109 CFU/mL。     

Antagonistic effect of Enterococcus faecium J96 against human and poultry pathogenic Salmonella spp.

Production of antagonistic compounds was studied in a strain of Enterococcus faecium isolated from the intestinal tract of a free-ranging chicken. Production of lactic acid and a bacteriocin was observed in cultures of this bacterium, alone and in mixed culture fermentations with pathogenic Salmonella serotypes (i.e., Gallinarum, Pullorum, Enteritidis, and Typhimurium). Growth inhibition of these avian and human pathogens was observed after 4 h of incubation at 37 degrees C in CAm broth, a medium developed according to the nutrients present in chicken food. The antibacterial action was due to the combined effect of lactic acid and bacteriocin. Accumulation of these metabolites caused both a bacteriostatic and a bactericidal action against the gram-negative bacteria assayed.     

双歧杆菌RH 菌株不同处理物修复肠道菌群平衡失调的研究

为探讨双歧杆菌RH 菌株对肠道菌群平衡失调的修复功能,本研究通过动物实验,考察双歧杆菌活菌(LB)、菌体破碎物(CE)及发酵上清液(FS)3 种处理物灌胃由抗生素造成的菌群失调模型小鼠后对其肠道内双歧杆菌、乳杆菌、产气荚膜梭菌、肠球菌、肠杆菌和拟杆菌6 大类主要菌群的影响。结果显示：3 种处理物均可使小鼠肠道内双歧杆菌和乳杆菌数量极显著增加(P ＜ 0.01),产气荚膜梭菌和肠球菌数量显著降低(P ＜ 0.05 和P ＜ 0.01),并能使肠杆菌和拟杆菌数量恢复至正常水平,说明该双歧杆菌LB、CE 和FS 均可起到修复、调节小鼠肠道微生态平衡的作用。     

Impact of intestinal microbiota and gastrointestinal conditions on the in vitro survival and growth of Bacillus cereus

Ingestion of B. cereus can result in diarrhea, if these bacteria survive gastrointestinal passage and achieve growth and enterotoxin production in the small intestine. The gastrointestinal survival of vegetative cells and spores of the diarrheal food poisoning strain B. cereus NVH 1230-88 was investigated during in vitro batch experiments simulating the stomach, duodenum and ileum using simulation media and competing intestinal microbiota. All spores and approx. 30% of the vegetative B. cereus cells survived the 2 h incubation in gastric medium with pH 4.0. Sterile intestinal medium induced germination of spores and enabled outgrowth of vegetative cells to approx. 7 log CFU/mL. The behavior of B. cereus in the intestinal environment with competing intestinal bacteria was determined by their relative concentrations. Besides the numbers of intestinal bacteria, the nutrition and composition of the intestinal community were also very important for the growth inhibition of B. cereus.     

仔猪复合微生态制剂的液体发酵工艺优化

对仔猪复合微生态制剂液体发酵产酶工艺进行了优化。采用响应面分析法得到最佳培养条件为:pH6.0、发酵温度30℃、培养时间65h、接种量10%,在此培养条件下,菌体生物量理论值为2.95×1012个/ml;对最佳培养条件进行验证性试验,重复3次得到菌体生物量平均值为3.04×1012个/ml。在最优混合培养条件下研究了发酵液中的纤维素酶、淀粉酶、糖化酶的酶活性,3种酶的最大酶活力分别为1 085、492和1 192U/ml。     

应用PCR检测食品中沙门氏菌前的增菌程序的优化

为了探讨更为简便、快速的增菌程序,对4 株食品中来源沙门氏菌进行增菌,比较振荡培养与静止培养,MM 增菌液和 BPW 增菌液,以及以BPW 为增菌液培养6、8、24h 的增菌效果。结果显示对于4 株食品中来源的沙门氏菌增菌6h 和8h 后,振荡培养相对于静止培养并不能够提高沙门氏菌的增菌效果(p ＞ 0.05),但增菌24h 后振荡培养能够极显著的提高增菌效果(p ＜ 0.01);在增菌6h 和8h 后,使用MM增菌液相对于BPW增菌液并不能提高增菌效果(p ＞ 0.05),但增菌24h 后,使用MM增菌液能够极显著的提高增菌效果(p ＜ 0.01);与使用BPW增菌液增菌8h 比增菌6h 的增菌效果达到极显著效应(p ＜ 0.01),增菌24h 与增菌8h 相比,增菌效果极显著(p ＜ 0.01)。结果表明：应用PCR 检测食品中的沙门氏菌前对于沙门氏菌增菌的前8h,使用振荡培养和选择性培养液MM培养液增菌并不能提高增菌效果;使用BPW 增菌液增菌8h 与增菌6h 相比增菌效果显著。由此可见在对食品中的沙门氏菌增菌时使用BPW 液37℃静止培养增菌8h,是一种优化的增菌程序。     

Pediocin PA-1 production during repeated-cycle batch culture of immobilized Pediococcus acidilactici UL5 cells

Pediocin PA-1 production by Pediococcus acidilactici UL5 cells immobilized in kappa-carrageenan/locust bean gum gel beads was studied during repeated-cycle batch (RCB) culture with pH control in Man Rogosa and Sharpe (MRS) broth supplemented with 1% glucose and whey permeate (SWP) medium. The pediocin PA-1 production by free P. acidilactici cells pH-controlled batch culture has reached 2048 and 4096 AU ml(-1) after 11 and 12 h of incubation, with volumetric productivities of 187 and 342 AU ml(-1) h(-1) in SWP and MRS media, respectively. In RCB culture, immobilized cells reached a maximum concentration of 7.3+/-0.2 x 10(10) and 4.3+/-0.9 x 10(10) cfu g(-1) of beads in MRS and SWP media, respectively. The maximum pediocin PA-1 activity obtained during RCB fermentation was 4096 AU ml(-1); it was attained after only 0.75 and 2 h of incubation in MRS and SWP media, respectively. The corresponding volumetric productivities were 5461 and 2048 AU ml(-1) h(-1). Pediocin PA-1 production in the RCB culture was highly stable over 12 fermentation cycles carried out over 3 d in SWP media.     

Production and Turnover of IgG1 and IgG2 Immunoglobulins in the Bovine around Parturition

Production rates (entry rate into blood plasma) and other metabolic parameters for the IgG1 and IgG2 subclasses of immunoglobulins in mammary secretions (ratio of about 7 to 1) were determined in cows around the time of parturition by both single-injection and continuous-infusion isotope-dilution techniques. Four cows were given a single dose of 150 to 200 μCi of iodine-125 labeled IgG1 and 100 to 250 μCi of iodine-131 labeled IgG2 at 2 to 1 wk before parturition. Four cows, including two of the above, were infused continuously with constant amounts of the labeled immunoglobulins starting at 11 to 4 days before parturition. All cows were maintained until 4 to 6 days after parturition for monitoring the specific activities of iodine-125 labeled IgG1 and iodine-131 labeled IgG2 in the plasma and mammary secretions. Maximum entry rates of IgG1 and IgG2 were between 3 and 1 day prepartum with means of 125 and 60 g/500 kg body weight per day. The exchangeable pool means for IgG1 and IgG2 were 619 and 643 g/500 kg body weight, and both immunoglobulins were divided almost equally between the intra- and extravascular pools. A greatly increased production and a shortened half-life or greater turnover for plasma IgG1 occurs around the time of parturition which can account for the large accumulation of IgG1 in colostrum.     

Evaluation of survival and release of encapsulated bacteria in ex vivo porcine gastrointestinal contents using a green fluorescent protein gene-labelled E. coli

A gfp-labelled E. coli strain K 12 was used to study the survival and release of calcium alginate encapsulated bacteria in ex vivo porcine intestinal contents. Encapsulated bacteria survived better in ex vivo porcine gastric conditions compared to non-encapsulated free bacterial cells. There was 2 log decrease in viable cells of encapsulated E. coli GFP⁺ after 3 h incubation in gastric contents compared to 4 log decrease in the free non-encapsulated cells. There was a complete release of encapsulated bacteria (E. coli GFP⁺) within 1 h of incubation in small intestinal contents at 37°C, while it took 8 h to nearly completely release the encapsulated bacteria in colon content under similar conditions. There was only a partial release of encapsulated bacteria incubated in duodenal content even after 10 h of incubation. Using gfp labelled E. coli, the efficacy of alginate capsule in protecting the bacterial cells from gastric environment and releasing the encapsulated bacteria in the intestine (desired site) was demonstrated.     

Effects of essential oils of oregano and nutmeg on growth and survival of Yersinia enterocolitica and Listeria monocytogenes in barbecued chicken

The in vitro effects of plant essential oils (EOs) against pathogenic bacteria are well known, yet few studies have addressed the effects of these compounds against pathogens associated with ready-to-cook foods. Experiments were conducted to determine the effectiveness of oregano and nutmeg EOs on the growth and survival of Yersinia enterocolitica and Listeria monocytogenes in broth culture and in Iranian barbecued chicken. Ready-to-cook Iranian barbecued chicken was prepared according to the common practice with 1, 2, and 3 microl/g of oregano and nutmeg EOs. The test and control (without EOs) samples were inoculated with Y. enterocolitica and L. monocytogenes to a final concentration of 6 to 7 log CFU/g and stored at 3, 8, and 20 degrees C. Microorganisms were counted just before and at 24, 48, and 72 h after storage based on growth on Yersinia selective agar supplemented with cefsulodine, igrasan, and novobiocin and on Listeria selective agar supplemented with nalidixic acid and acriflavin. In the broth culture system, the nutmeg EO had a greater effect on L. monocytogenes (MIC = 0.20 nicrol/ml) than did the oregano EO (MIC = 0.26 microl/ml). However, the oregano EO had a greater effect on Y. enterocolitica (MIC = 0.16 microl/ml) than did the nutmeg EO (MIC = 0.25 microl/ml). In ready-to-cook Iranian barbecued chicken, the log CFU per gram of both bacteria after up to 72 h of incubation was not decreased significantly by various combinations of oregano and nutmeg EOs (1, 2, and 3 microl/g) and storage temperatures (3, 8, and 20 degrees C) when compared with control samples (without EOs). Although examination of spices in culture media can yield accurate microbiological data, without complementary tests in foods these data are of limited value for assessing food safety.     

A conductance method for the identification of Escherichia coli O157:H7 using bacteriophage AR1.

The feasibility of using a specific phage (AR1) in conjunction with a conductance method for the identification of Escherichia coli O157:H7 was evaluated. The multiplication of strains of E. coli O157:H7 was inhibited by AR1; therefore, a time point (detection time, DT) at which an accelerating change in conductance in the culture broth was not obtained. Bacterial strains were subcultured on sorbitol-MacConkey agar and incubated at 35 degrees C for 24 h, and the ability of the bacteria to ferment sorbitol was recorded. An aliquot of 0.5 ml of the bacterial suspension (10(7) CFU/ml) and 0.5 ml of the phage suspension (10(8) PFU/ml) were added to the conductance tube of a Malthus analyzer containing 5 ml of culture broth. The tubes were incubated at 35 degrees C, and conductance changes in the tubes were continuously monitored at 6-min intervals for 24 h by the instrument. A positive reaction was defined as an E. coli strain that could not utilize sorbitol and caused no conductance change (i.e., no DT) within an incubation period of 24 h. Of the 41 strains of E. coli O157:H7 tested, all produced positive reactions. When a total of 155 strains of non-O157:H7 E. coli were tested, 14 did not have a DT within 24 h. However, among these 14 strains, 13 were sorbitol fermenters, and the remaining one was a nonfermenter. Therefore, by definition, only one strain produced a false-positive reaction. The sensitivity and specificity of the present method were 100% (41 of 41) and 99.4% (154 of 155), respectively. The present method incorporating conductimetric measurement and phage AR1 for the identification of E. coli O157:H7 was simple and capable of automation.     

一株产酸凝结芽孢杆菌的筛选及耐受特性研究

从散养鸡肠道内容物分离到5株溶钙圈较大的芽孢菌,经高效液相色谱（HPLC）测定筛选到一株产酸性能较好的菌株DPLY-7,综合菌落形态、生理生化及特异性序列分析等手段,确定其分类归属,并对其耐受特性进行了研究。结果表明,菌株DPLY-7被鉴定为凝结芽孢杆菌（Bacillus coagulans）。在37 ℃培养10 h发酵液pH最低为5.04,其乳酸、乙酸含量分别可达2.60 mg/mL、1.24 mg/mL。菌株DPLY-7在pH值2.5环境中的存活率为79.76%,胆盐浓度0.5%环境中存活率为42.86%,可以耐受人工胃肠液及85 ℃水浴处理,是一株耐受性能优越的益生菌。     

Antimicrobial effect of lactic acid producing bacteria culture condensate mixture (LCCM) against Salmonella enteritidis

The antimicrobial effects of a lactic acid producing bacteria culture condensate mixture (LCCM) were assessed against Salmonella enteritidis. In the presence of LCCM, bacterial growth was assessed in vitro by the measurement of optical density (OD) and viable bacterial counting. At concentrations of 1.25 and 2.5% LCCM, OD values were significantly lower than that of the control broth, and at concentrations of 5 and 10% LCCM, OD values did not increase for the entire period of experiment. At 8 h after incubation, the viable bacterial numbers in 5% and 10% LCCM-containing broths were remarkably lower than that in the control broth. This antimicrobial ability of the LCCM was fundamentally attributed to causing cell death rather than inhibiting growth. Even when the pH of LCCM-containing broth was adjusted to 7.2, the number of viable bacteria was significantly lower in the broths containing LCCM over 2.5% than that in control broth at 8 h after incubation. However, the OD value of each culture in the presence of each concentration of the LCCM increased over 1.0 at the completion of the experiment. The in vivo antimicrobial effects of the LCCM against S. enteritidis were also assessed. In S. enteritidis-infected mice, the LCCM decreased both the viable bacteria found in the feces and the mortality rate of the mice. These findings showed that the LCCM might have an antimicrobial ability against S. enteritidis.