No abstract

This review highlights the main strategies available to control phage infection during large-scale milk fermentation by lactic acid bacteria. The topics that are emphasized include the factors influencing bacterial activities, the sources of phage contamination, the methods available to detect and quantify phages, as well as practical solutions to limit phage dispersion through an adapted factory design, the control of air flow, the use of adequate sanitizers, the restricted used of recycled products, and the selection and growth of bacterial cultures.

Aim:  To investigate the influence of several environmental factors on the viability and cell‐adsorption for two Lactobacillus casei/paracasei bacteriophages (PL‐1 and J‐1). Methods and Results:  Both phages showed a remarkably high specificity of species, sharing similar host spectra. Two phages and four sensitive strains were used to conform five phage/strain systems. Each showed a particular behaviour (burst size: ranging from 32 to 160 PFU/infective centre; burst time: 120–240 min and latent time: 5–90 min). For both phages, the viability was not significantly affected from pH 4 to 11 (room temperature) and from pH 5 to 10 (37°C). Adsorption rates were not influenced by calcium ions, but decreased after the thermal inactivation of cells. Adsorption rates were high between 0 and 50°C with maximum values at 30°C and pH 6. System PL‐1/Lact. paracasei A showed noticeable differences in comparison with the others, being times required to reach 90% of adsorption of 4 h and lower than 45 min, respectively. Conclusions:  The data obtained in this work demonstrated that environmental parameters can influence the viability and cell adsorption rates of Lact. casei/paracasei phages. The extent of this influence was phage dependent. Significance and Impact of the Study:  This work contributes to the enlargement of the currently scarce knowledge of phages of probiotic bacteria.

Aims: The survival of two collection Lactobacillus casei and L. paracasei bacteriophages when subjected to thermal and chemical treatments was investigated. Methods and Results: Thermal resistance was evaluated by heating phage suspensions at 63, 72 and 90°C in three different media [Tris-magnesium gelatin (TMG) buffer: 10 mmol l )1 Tris-Cl, 10 mmol l )1 MgSO 4 and 0AE1% w/v gelatin; Man Rogosa Sharpe (MRS) broth and reconstituted nonfat dry skim milk (RSM)]. A marked heat sensitivity was evident in both phages, as 15 min at 72°C was enough to completely inactivate (6 log 10 reduction) them. No clear influence was demonstrated by the suspension media. The phages also showed similar resistance to biocides. Peracetic acid and sodium hypochlorite (800 ppm) were the most effective ones, destroying the phages within 5 min. Concentrations of 75 and 100% ethanol were not suitable to inactivate phage particles even after 45 min. Isopropanol did not show an effect on phage viability. Conclusions: The data obtained in this work are important to design more effective control procedures in order to inactivate phages in dairy plants and laboratories. Significance and Impact of the Study: This work will contribute to enhance the background knowledge about phages of probiotic bacteria. 1

No abstract

A new virulent bacteriophage (MLC-A) was recently isolated in Argentina from a probiotic dairy product containing a strain of Lactobacillus paracasei. Observation of the lysate with an electron microscope revealed bacteriophage particles with an icosahedral capsid of 57 +/- 2 nm; with a collar and a noncontractile tail of 156 +/- 3 nm terminating with a baseplate to which a tail fiber was attached. Therefore, phage MLC-A belongs to the Siphoviridae family. This phage was able to survive the pasteurization process and was resistant to alcohols and sodium hypochlorite (400 mg/kg). Only peracetic acid could inactivate high-titer suspensions of phages in a short time. The maximum rates of phage adsorption to its host cells were obtained at 30 degrees C with a pH between 5 and 7, and in the presence of calcium or magnesium ions. The host range of phage MLC-A encompassed L. paracasei and Lactobacillus casei strains, but it was not able to infect Lactobacillus rhamnosus or Lactobacillus gasseri strains. One-step growth kinetics of its lytic development revealed latent and burst periods of 30 and 135 min, respectively, with a burst size of about 69 +/- 4 plaque-forming units per infected cell. Phage MLC-A had a distinctive restriction profile when compared with the 2 well-studied Lactobacillus phages, PL-1 and J-1. The genome size of the MLC-A phage was estimated to be approximately 37 kb. This study presents the description of the first phage specific for L. paracasei isolated in Argentina. The isolation of phage MLC-A indicates that, beside lactic acid bacteria starters, probiotic cultures can also be sensitive to virulent phages in industrial processes.

The effect of several biocides and thermal treatments on the viability of four Lactobacillus helveticus phages was investigated. Times to achieve 99% inactivation of phages at 63 degrees C and 72 degrees C in three suspension media were calculated. The three suspension media were tris magnesium gelatin buffer (10 mM Tris-HCl, 10 mM MgSO4, and 0.1% wt/vol gelatin), reconstituted skim milk sterile reconstituted commercial nonfat dry skim milk, and Man Rogosa Sharpe broth. The thermal resistance depended on the phage considered, but a treatment of 5 min at 90 degrees C produced a total inactivation of high titer suspensions of all phages studied. The results obtained for the three tested media did not allow us to establish a clear difference among them, since some phages were more heat resistant in Man Rogosa Sharpe broth and others in tris magnesium gelatin buffer. From the investigation on biocides, we established that sodium hypochlorite at a concentration of 100 ppm was very effective in inactivating phages. The suitability of ethanol 75%, commonly used to disinfect utensils and laboratory equipment, was confirmed. Isopropanol turned out to be, in general, less effective than ethanol at the assayed concentrations. In contrast, peracetic acid (0.15%) was found to be an effective biocide for the complete inactivation of all phages studied after 5 min of exposure. The results allowed us to establish a basis for adopting the most effective thermal and chemical treatments for inactivating phages in dairy plant and laboratory environments.

No abstract