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Stabilization of Lactose Metabolism in Streptococcus lactis C2.

The integration of the lactose plasmid from lactic streptococci into the host chromosome could stabilize this trait for dairy fermentations. Sixty lactose-positive (Lac) transductants of lactose- and proteinase-negative (Lac Prt) LM0220 were induced for temperature phage by UV irradiation or mitomycin C. Four of the transductants, designated KB18, KB21, KB54, and KB58, yielded lysates demonstrating less than one Lac transductant per 0.2 ml of phage lysate. Successive transferring in the presence of acriflavine did not yield Lac segregants from KB18, KB21, KB54, or KB58, whereas Streptococcus lactis C2 (parent culture) and three other Lac transductants showed 12 to 88% conversion from Lac to Lac within 6 to 10 repetitive transfers. When grown in continuous culture, KB21 did not show any Lac variants in 168 h, while S. lactis C2 had 96% conversion from Lac to Lac in 144 h. Agarose gel electrophoresis of plasmid DNA isolated from KB18, KB21, KB54, and KB58 revealed that the lactose plasmid, pLM2103, normally present in Lac transductants, was missing. This suggested integration of the transferred lactose plasmid into the chromosome. In contrast to phage lysates induced from S. lactis C2, which exhibited an exponential decrease in the number of Lac transductants after exposure to small doses of UV irradiation, the transduction frequency for lactose metabolism was stimulated by UV irradiation of lysates from KB58. The latter indicated chromosomal linkage for lac and that integration of the lactose genes plasmid into the chromosome had occurred.

Journal Article↗

Galactose Expulsion during Lactose Metabolism in Lactococcus lactis subsp. cremoris FD1 Due to Dephosphorylation of Intracellular Galactose 6-Phosphate.

In Lactococcus lactis subsp. cremoris FD1, galactose and lactose are both transported and phosphorylated by phosphotransferase systems. Lactose 6-phosphate (lactose-6P) is hydrolyzed intracellularly to galactose-6P and glucose. Glucose enters glycolysis as glucose-6P, whereas galactose-6P is metabolized via the tagatose-6P pathway and enters glycolysis at the tagatose diphosphate and fructose diphosphate pool. Galactose would therefore be a gluconeogenic sugar in L. lactis subsp. cremoris FD1, but since fructose 1,6-diphosphatase is not present in this strain, galactose cannot serve as an essential biomass precursor (glucose-6P or fructose-6P) but only as an energy (ATP) source. Analysis of the growth energetics shows that transition from N limitation to limitation by glucose-6P or fructose-6P gives rise to a very high growth-related ATP consumption (152 mmol of ATP per g of biomass) compared with the value in cultures which are not limited by glucose-6P or fructose-6P (15 to 50 mmol of ATP per g of biomass). During lactose metabolism, the galactose flux through the tagatose-6P pathway (r(max) = 1.2 h) is lower than the glucose flux through glycolysis (r(max) = 1.5 h) and intracellular galactose-6P is dephosphorylated; this is followed by expulsion of galactose. Expulsion of a metabolizable sugar has not been reported previously, and the specific rate of galactose expulsion is up to 0.61 g of galactose g of biomass h depending on the lactose flux and the metabolic state of the bacteria. Galactose excreted during batch fermentation on lactose is reabsorbed and metabolized when lactose is depleted from the medium. In vitro incubation of galactose-6P (50 mM) and permeabilized cells (8 g/liter) gives a supernatant containing free galactose (50 mM) but no P(i) (less than 0.5 mM). No organic compound except the liberated galactose is present in sufficient concentration to bind the phosphate. Phosphate is quantitatively recovered in the supernatant as P(i) by hydrolysis with alkaline phosphatase (EC 3.1.3.1), whereas inorganic pyrophosphatase (EC 3.6.1.1) cannot hydrolyze the compound. The results indicate that the unknown phosphate-containing compound might be polyphosphate.

Journal Article↗

Molecular cloning, transcriptional analysis, and nucleotide sequence of lacR, a gene encoding the repressor of the lactose phosphotransferase system of Lactococcus lactis.

The repressor gene (lacR) of the lactose phosphotransferase system of Lactococcus lactis subsp. lactis strain MG1820 has been cloned and characterized. Transcription of lacR, into a 1.2-kilobase monocistronic messenger, is repressed approximately 5-fold during growth on lactose. Nucleotide sequence analysis of the lacR gene showed the presence of an open reading frame of 861 base pairs. The deduced amino acid sequence of LacR is homologous to three Escherichia coli regulatory proteins (DeoR, FucR, and GutR) and includes a N-terminal domain (helix-turn-helix) involved in DNA binding and a C-terminal domain that may be responsible for inducer binding. The in vivo function of LacR has been determined by introducing multiple copies of lacR into L. lactis, under control of its own or the unrelated prtP promoter. Growth rates and lactose phosphotransferase system enzyme activities were measured during growth on lactose and glucose. The presence of lacR on a multicopy plasmid resulted in the decrease of lactose phosphotransferase system activity, whereas only on lactose a decrease (25%) of growth rate was observed. No significant difference in growth rate was observed on glucose, indicating that LacR specifically represses the lactose genes of L. lactis.

Amino Acid Sequence↗

Jejunoscopic lactose provocation.

A new method for testing lactose intolerance was developed. The examination is performed by jejunoscopy using lactose dilution for provocation and sucrose for control. Macroscopic and microscopic changes in the jejunal mucosa are examined after administration of solutions. The results of this study demonstrated that lactose administration causes prompt change in the jejunal mucosa in cases of lactose intolerance but had no effect in cases of lactose tolerance. Administration of sucrose did not result in any change. This method is effective in proving lactose intolerance and may be useful to elucidate the pathogenesis of this disorder.

Adult↗

A possible explanation of the effects of dietary lactose concentration during recovery of protein depletion in growing rats.

It is generally accepted that lactose intolerance is a cause of delayed recovery from malnutrition. To discriminate the true importance of this factor, the effect of dietary lactose concentration on "catch up" growth was studied on rats fed a protein free diet from weaning to 34 days of age and then refed "ad libitum" with diets providing different levels of Protein Calories (P%: 5, 11, 17 and 25%) and Lactose (L%: 0, 1, 15, 27 and 37%) for 4 days. Weight changes, daily dietary intakes and gastrointestinal disorders (GID) were recorded. The growth rate (GR) -g/100g rat/day- showed a direct correlation with P% whereas a decrease in (GR) as well as in Protein Utilization (PU) was associated with increasing L%, the higher the P%, the greater the effect of L%. The observed effect of lactose on (PU) does not correlate with (GID). Analysis of intake data -expressed as calories/day/W0.73- showed that animals ate less diet when lactose was present resulting in energy restriction (ER) from 12 to 62%. A primary effect of food intake might be proposed as a limiting factor for (GR) and (PU) in rats recovering from depletion. The influence of P% on improved lactose tolerance must also be stressed.

Animals↗

Management of lactose maldigestion by consuming milk containing lactobacilli.

The influence of nonfermented milk containing L. acidophilus or L. bulgaricus on lactose utilization by lactose maldigesters was investigated. Nonfermented milks containing L. acidophilus or L. bulgaricus at 10(8) and 10(9) CFU/ml were prepared using 2% low-fat milk. Lactose maldigestion was monitored by measuring breath hydrogen at hourly intervals for 8 hr following consumption of 400 ml of each diet. Nonfermented milk containing L. acidophilus B at 10(8) CFU/ml were not effective in reducing breath hydrogen and symptoms. Nonfermented milk containing L. acidophilus B at 10(9) CFU/ml only slightly decreased breath hydrogen production; however, the symptoms were significantly improved. Nonfermented milks containing L. bulgaricus 449 at 10(8) and 10(9) CFU/ml were effective in reducing breath hydrogen and symptoms. The results for bulgaricus milk were all significant. In this study, L. acidophilus B and L. bulgaricus 449 were chosen because of their similar beta-galactosidase activity and bile sensitivity. L. acidophilus and L. bulgaricus are both thermophilic lactobacilli and an active transport (permease) system is found in both species for lactose transport. The major factor affecting in vivo lactose digestion in this study appears to be the bacterial cell wall/membrane structures. That the cell wall/membrane structures of L. acidophilus are different from those of L. bulgaricus can be indirectly proven by the results of sonication time for maximum beta-galactosidase activity measurement. The results of this study indicate that L. bulgaricus is usually a better choice than L. acidophilus for manufacturing nonfermented milks for lactose maldigesters.

Animals↗

In vitro and in vivo lactose and lactulose effects on colonic fermentation and portal-systemic encephalopathy parameters.

Lactose intolerance occurs in the majority of human groups, excluding people from Northern Europe. Because its effect is similar to that of lactulose, lactose seems to be an alternative treatment for patients with portal-systemic encephalopathy (PSE) and lactase deficiency. The mechanism of action of lactose is similar to that of lactulose. In vivo, lactose improves PSE parameters and causes acidic diarrhea. We performed in vitro studies in a fecal incubation system to investigate the biochemical and bacteriological effects induced by different substances customarily used for the treatment of patients with PSE (lactose, lactulose and Neomycin). In vitro experiments showed that lactose and lactulose decreased aerobic flora counts and reduced the pH of fecal incubation. Both disaccharides reduced the ammonia concentration in the incubation system.

Anti-Bacterial Agents↗

Influence of changes in lactase activity and small-intestinal mucosal growth on lactose digestion and absorption in preterm infants.

BACKGROUND: Feeding intolerance (ie, achieving and maintaining full enteral feedings) is a significant problem in preterm infants. A relation exists between feeding intolerance and incomplete lactose digestion. OBJECTIVES: We sought to identify the factors relating to lactose digestion and absorption, lactase activity, and small-intestinal mucosal growth. DESIGN: Lactose digestion and absorption, lactase-specific activity, and lumen-to-mucosa water flux as a measure of small-intestinal mucosal surface area were determined by using the triple-lumen perfusion technique on 2 occasions 3 wk apart in 10 preterm infants (x+/-SEM gestational age: 28.0+/-0.2 wk). RESULTS: Lactose digestion and absorption and lactase activity doubled between studies (P=0.035 and P=0.041, respectively). The change in digestion and absorption was related to lactase activity (P=0.034, R2=0.38). Lactase activity correlated with gestational age at birth (P=0.012, R2=0.51). The number of days of feeding explained 80% of the variability in small-intestinal mucosal surface area (P=0.001). CONCLUSIONS: To our knowledge, this is the first study to measure directly lactose digestion and absorption, lactase activity, and small-intestinal surface area in preterm infants. Changes in lactose absorption relate primarily to lactase activity rather than to mucosal growth. We showed directly a relation between enteral feeding and small-intestinal mucosal growth.

Digestion↗

Long-term acceptance of low-lactose milk.

Low-lactose milk was produced by incubating cow's milk with yeast lactase. Sixteen lactose tolerant and 15 intolerant volunteers ingested 500 ml of the product twice daily for 1 month. During the testing period all subjects received on three occasions the same volume of unmodified milk in double-blind tests. Symptoms recorded throughout the study and for an additional 15 day base-line observation period were: diarrhea, abdominal pain and distention, flatulence, heartburn, and headache. Low-lactose milk acceptance was excellent. No significant differences were found between tolerants and intolerants during the base-line period and while ingesting low-lactose milk. By contrast, unmodified milk induced severe symptoms only in the intolerants. Availability of low-lactose milk and of its by-products allows consumption of greater volumes of this highly nutritious food by subjects with lactose intolerance with none or less symptoms compared to unmodified milk.

Adolescent↗

Nutritional consequences of low dose milk supplements consumed by lactose-malabsorbing children.

The nutritional consequences of supplementary milk consumption by lactose-malabsorbing children were determined by nutrient balance studies. Twelve subjects received a marginally adequate rice and vegetable base-line diet alone and with simulated milk supplements containing either glucose or lactose during three separate balance periods. The diets were equally well accepted and tolerated. The children gained significantly more weight and had improved apparent nitrogen absorption and retention on the milk supplemented diets (P less than 0.001), and there was no difference between the effects of glucose milk and lactose milk. Fecal wet weights and energy and carbohydrate excretions were modestly increased with the lactose-containing diet, but not significantly so. It is suggested that low dose milk supplements can be well utilized when consumed by lactose malabsorbers in conjunction with other foods. Milk consumption need not be discouraged for populations among whom lactose malabsorption is widely prevalent, but milk should be provided in relatively low doses and the clinical responses to its consumption should be monitored.

Animals↗

Clinical studies with low-lactose milk.

A low-lactose milk was evaluated for taste acceptance and clinical symptomatology by means of a double-blind control study in two groups of individuals. One group consisted of nine milk intolerant individuals, while the other consisted of five milk tolerant individuals. Each week for 9 wk the participants were given a coded sample of skim milk, lactose hydrolyzed milk, skim milk plus glucose, or sweet acidophilus milk. Each participant was asked to consume four liters of milk during a week and keep a daily log of symptoms (pain, bloating, nausea, flatus, emesis, bowel frequency) along with taste acceptability. After assigning a numerical value to the intensity of symptomatology a X2 analysis was performed on the data. In the milk intolerant population lactose hydrolyzed milk produced significantly milder (p < 0.05) pain and gas symptoms than the nonhydrolyzed milks. Bowel frequency was not altered between the types of milk in both groups. The lactose hydrolyzed milk did not reduce the symptoms of lactose intolerance in the milk intolerance population to the response of the control group. Although both study populations found decreased taste acceptability to the lactose hydrolyzed milk, a taste panel assessment did not show any significant differences in the milks.

Adult↗

Prevalence of primary adult lactose malabsorption and awareness of milk intolerance in Italy.

A total of 308 healthy Italian adults (192 females, 116 males; mean age 29.2 yr) were examined using a field version of the lactose tolerance test with breath hydrogen determination. Two geographical groups were formed according to the birth places of the probands' grandparents: 208 subjects from northern Italy (mainly from the regions of Piemonte, Lombardia, and Veneto) and 100 probands from Sicily. Lactose malabsorption was diagnosed in 106 subjects in group "north" (51%) and in 71 subjects in group "Sicily" (71%). Awareness of milk intolerance was more frequent in lactose malabsorbers. The incidence of diarrhea after the test dose of lactose was significantly higher in "aware" lactose malabsorbers, when compared with persons of the same group who had not experienced milk intolerance. The significant difference in lactose malabsorption frequency between northern Italy and Sicily is further evidence of a north-south gradient of lactase gene frequencies in Europe.

Adult↗

Lactose malabsorption from yogurt, pasteurized yogurt, sweet acidophilus milk, and cultured milk in lactase-deficient individuals.

The use of fermented dairy foods is common in areas of the world where lactase deficiency is prevalent. Recently, we have shown that the digestion of lactose from yogurt is enhanced as compared to that from milk. This enhanced digestion is apparently due to inherent B-galactosidase in yogurt which is active in the gastrointestinal tract after consumption of the yogurt. Furthermore, yogurt is well tolerated by lactase-deficient subjects resulting in little or no gastrointestinal distress. Since other fermented and microbial-containing dairy foods are consumed worldwide and may also contain some "lactase" activity, we chose to evaluate the digestion of lactose from three of these products: pasteurized yogurt, cultured milk (buttermilk), and sweet acidophilus milk. Breath hydrogen techniques were used to evaluate lactose malabsorption in nine lactase-deficient subjects. The studies demonstrated that yogurt is unique among the products tested in enhancing the digestion of lactose. Furthermore, pasteurization of yogurt eliminated the enhanced digestion of lactose, reduced the inherent lactase activity of the yogurt by 10-fold and reduced cell counts by 100-fold. Interestingly, eight of nine subjects fed cultured milk experienced gastrointestinal distress, whereas all subjects fed pasteurized yogurt were symptom free, even though the amount of malabsorbed lactose was similar.

Adult↗

Effective in vivo hydrolysis of milk lactose by beta-galactosidases in the presence of solid foods.

The feasibility of enzyme replacement therapy with exogenous, food-grade, microbial enzymes at mealtime to effect intragastrointestinal hydrolysis of the lactose from 360 ml of cow's milk consumed with a solid food meal (breakfast cereals) was investigated in adult Guatemalan lactose-malabsorbers using a hydrogen breath-analysis procedure to quantify the completeness of postprandial carbohydrate absorption. Adding 2 g of a commercial preparation of beta-galactosidase from Kluyveromyces lactis at mealtime to milk taken with a refined cereal (cornflakes) and an unrefined cereal (bran) reduced the production of excess breath H2 attributable to lactose maldigestion to a level not significantly different from that achieved with lactose-prehydrolyzed milk. Sucrase, as expected, had no effect on H2 production. A beta-galactosidase from Aspergillus niger was less effective that the K. lactis enzyme for in vivo hydrolysis. Thus, exogenous betagalactosidases can eliminate lactose malabsorption in lactase-deficient individuals even in the presence of solid foods, allowing lactose intolerant persons to consume milk and dairy products without gastrointestinal discomfort.

Adult↗

Modification of sweet acidophilus milk to improve utilization by lactose-intolerant persons.

Enhanced digestion of yogurt by lactose-intolerant individuals is believed to be due to inherent beta-galactosidase (lactase) in the culture organisms that aids in the hydrolysis of ingested lactose. However, sweet acidophilus milk, which contains lactase-rich organisms, does not enhance lactose digestion. Using breath-hydrogen measurements to indicate malabsorption in 14 human subjects, we compared utilization of: milk, yogurt, heated yogurt, yogurt plus lactose, heated yogurt plus lactase, sweet acidophilus milk (SAM), and SAM made with sonicated cells. Results indicate that both the reduction of lactose during fermentation and the presence of indigenous bacterial lactase are responsible for the increased ability to tolerate lactose in yogurt. Improved utilization of SAM by sonication suggests that intracellular lactase is not available during digestion and that sonication releases the lactase activity from the cells.

Animals↗

Lactose absorption in patients with ovarian cancer.

To determine whether lactase persistence might be related to ovarian cancer risk, in 1994-1995 the authors assessed the capacity to digest lactose by measuring breath hydrogen production after oral administration of lactose in 50 women with ovarian cancer and 100 healthy controls. All of the women came from Sassari (Sardinia), Italy, an area where the population has a high frequency of lactose malabsorption. Thirty percent of cases were lactose absorbers, as compared with 15% of controls. The odds ratio for ovarian cancer among lactose absorbers was 2.51 (95% confidence interval 1.10-5.68). These results provide some support for a role of lactose ingestion and galactose cytotoxicity in the pathogenesis of ovarian cancer.

Administration, Oral↗

Use of the lactose-ethanol tolerance test in diabetes.

The standard lactose tolerance test involves measuring a patient's blood glucose after the ingestion of lactose. If the patient has lactase deficiency and is unable to hydrolyze lactose and absorb its monosaccharides, glucose and galactose, the blood glucose does not usually increase greater than 20 mg/100 ml. Since factors other than the absorption of glucose can cause an increase in the blood glucose of greater than 20 mg/100 ml in a diabetic, this test could be unreliable when it is performed on a diabetic. The present study was performed to determine whether the lactose-ethanol tolerance test could be used to diagnose lactoase deficiency in diabetics. This test involves measuring the blood galactase level, instead of the blood glucose, and the administration of ethyl alcohol to a subject prior to the test to delay the clearance of galactose from the circulation. The results indicate that the standard lactose tolerance test in which the blood glucose is measured is unreliable when performed on insulin-dependent diabetics, but that it can be reliable when performed on non-insulin-dependent diabetics. The lactose-ethanol tolerance test gave results in each type of diabetic which were qualitatively similar to those of nondiabetics. It was concluded that the latter test is a useful screening test for lactase deficiency in diabetics.

Adult↗

Two-hour lactose breath hydrogen test.

Current requirements for the lactose breath hydrogen test (LBHT) include serial expired air samplings and multiple hydrogen (H2) determinations. One hundred thirty-two consecutive LBHTs were evaluated to determine whether multiple samplings are indeed necessary for detection of lactose malabsorption. Expired air samples were collected at 0, 30, 60, 90, 120, 150, and 180 min following ingestion of lactose. Fifty-five LBHTs were positive for lactose malabsorption. All tests showed abnormally elevated breath H2 concentrations at 120 min. The mean value of the change in parts per million (delta ppm) of H2 at 120 min (51.1 +/- 4.7 SEM) was higher than at any other time point. If only the 120-min samples were examined without subtracting the initial concentrations, four of the 77 negative tests (5.2%) would have been falsely positive. Thus, the values of H2 at 0 and 120 min were sufficient to define lactose malabsorption in all cases. We conclude that just as a single blood sample now suffices for determining xylose malabsorption, so expired air sampling at only 0 and 120 min during the LBHT is a reliable method for detecting lactose malabsorption and diminishes the need for acquiring and analyzing multiple samples.

Adolescent↗