Lactobacillus Rhamnosus GG Cells Research Articles

Effect of air freezing, spray freezing, and pressure shift freezing on membrane integrity and viability of Lactobacillus rhamnosus GG

The objective of this work was to assess and compare the impact of three different freezing methods on the physiology of probiotic bacteria based on evaluation of vitality, membrane integrity and special metabolic properties. Lactobacillus rhamnosus GG (LGG) was used as a model strain and was analysed with the plate enumeration method and flowcytometric analyses (FCM) before and after treatment in phosphate buffer saline (PBS) and 20% (w/v) skim milk (RSM), respectively. To get insight into the induced changes in esterase activity and membrane integrity combined staining with carboxyfluorescein diacetate (cFDA) and propidium iodide (PI) was applied. Furthermore the ability of the cells to extract intracellular accumulated cF in presence of glucose was investigated as an indicator for ATP driven membrane transport processes. The freezing methods applied were static air freezing (AF) at −30 °C for 3 h, high pressure shift freezing (PSF) at −21 °C/210 MPa and spray freezing (SF) at −30 °C. Spray freezing was accomplished by spraying the bacteria suspension in a cold air stream using a two fluid atomizer. The atomizing gas was air. After each treatment all samples were stored at −20 °C for one day before they were analysed. Minimum losses in viability occurred when spray freezing LGG in presence of 20% (w/v) RSM whereas air freezing in PBS resulted in the lowest survival rates. For all treatments higher survival rates were achieved when 20% (w/v) RSM was used as protecting media. Comparing the results of live cell count with the FCM data gave evidence for a fraction of viable but not culturable cells. LGG cells frozen by PSF showed viability comparable to AF samples, but physiological characteristics of pressure treated bacteria.

Effects of Lactobacillus rhamnosus GG addition in ice cream

A 24 full factorial experimental design was applied to verify the effects of Lactobacillus rhamnosus GG (LGG) addition in retail‐manufactured ice cream stored at two different freezing temperatures (−16°C and −28°C) and containing two different levels of sugar (15–22%) and fat (5–10%). In addition to microbial counts, the pH, acidity, viscosity of the mixes and functional properties of the ice creams were evaluated. Both fresh and frozen‐thawed LGG cells underwent preliminary resistance tests to bile, antibiotics and acidity. The LGG strain proved to be highly resistant to most of the stress factors. When the micro‐organism was added to ice cream mixes in a quantity of 108 cfu/g, it did not change the overrun, firmness or melting behaviour of the finished product. Regardless of formulation, no count decay of LGG cells was observed in ice cream stored for up to 1 year.

Stability and activity of specific antibodies against Streptococcus mutans and Streptococcus sobrinus in bovine milk fermented with Lactobacillus rhamnosus strain GG or treated at ultra-high temperature.

Passive local immunization against dental caries is a promising approach to its prevention, as clinical evidence of active oral or nasal immunization is still limited and controversial. By means of systemic immunization of pregnant cows with a multivalent vaccine, high titres of IgG antibodies against human cariogenic bacteria, Streptococcus mutans and Streptococcus sobrinus, were produced in bovine colostrum. The purified immune product (IP) of this preparation has a number of anticariogenic properties, such as inhibition of streptococcal adherence to saliva-coated hydroxyapatite and inhibition of glucosyltransferase enzymes. This study investigated whether IP antibodies remained active and functional when added to ultra-high temperature (UHT)-treated milk or to Lactobacillus rhamnosus GG (LGG)-fermented milk stored for an extended time. LGG was chosen because of its widely known health benefits in humans and animals. A commercial UHT toddler's milk was supplemented with IP and stored for 2 months at 5, 21 and 30 degrees C. The antistreptococcal titres in UHT milk did not decline at any temperature during storage, and UHT-IP inhibited the adherence of S. mutans for up to 2 months. This was not the case with UHT toddler's milk without IgG antibodies. Milk was fermented with live LGG cells in the presence or absence of 5% IP. The antistreptococcal titres declined to about 30% of the original titres after storage. Fresh milk alone slightly enhanced streptococcal adhesion but fresh milk with IP inhibited the adherence of S. mutans by over 50%. LGG-positive fermented milk without antibodies also inhibited (P < 0.05) the adhesion by about 40%. In both LGG-fermented and UHT immune milk, the activity of antibodies against cariogenic streptococci was maintained during the expected shelf-life of these products. From the anticariogenic point of view it may be beneficial to add bovine-specific antibodies against mutans streptococci to probiotic LGG-containing milk products.