RESUMO
AIMS: The aim of this work was to study the adsorption step of two new temperate bacteriophages (Cb1/204 and Cb1/342) of Lactobacillus delbrueckii and to isolate phage-resistant derivatives with interesting technological properties. METHODS AND RESULTS: The effect of divalent cations, pH, temperature and cell viability on adsorption step was analysed. The Ca2+ presence was necessary for the phage Cb1/342 but not for the phage Cb1/204. Both phages showed to be stable at pH values between 3 and 8. Their adsorption rates decreased considerably at pH 8 but remained high at acid pH values. The optimum temperatures for the adsorption step were between 30 and 40°C. For the phage Cb1/342, nonviable cells adsorbed a lower quantity of phage particles in comparison with the viable ones, a fact that could be linked to disorganization of phage receptor sites and/or to the physiological cellular state. The isolation of phage-resistant derivatives with good technological properties from the sensitive strains and their relationship with the cell heterogeneity of the strains were also made. CONCLUSIONS: Characterization of the adsorption step for the first temperate Lact. delbrueckii phages isolated in Argentina was made, and phage-resistant derivatives of their host strains were obtained.
Assuntos
Bacteriófagos/fisiologia , Lactobacillus delbrueckii/virologia , Adsorção/efeitos dos fármacos , Bacteriófagos/isolamento & purificação , Cálcio/farmacologia , Concentração de Íons de Hidrogênio , Viabilidade Microbiana , TemperaturaRESUMO
The aim of this work was to study the efficiency of diverse chemical and thermal treatments usually used in dairy industries to control the number of virulent and temperate Lactobacillus delbrueckii bacteriophages. Two temperate (Cb1/204 and Cb1/342) and three virulent (BYM, YAB and Ib3) phages were studied. The thermal treatments applied were: 63 degrees C for 30 min (low temperature--long time, LTLT), 72 degrees C for 15 s (high temperature--short time, HTST), 82 degrees C for 5 min (milk destined to yogurt elaboration) and 90 degrees C for 15 min (FIL-IDF). The chemical agents studied were: sodium hypochlorite, ethanol, isopropanol, peracetic acid, biocides A (quaternary ammonium chloride), B (hydrogen peroxide, peracetic acid and peroctanoic acid), C (alkaline chloride foam), D (p-toluensulfonchloroamide, sodium salt) and E (ethoxylated nonylphenol and phosphoric acid). The kinetics of inactivation were drew and T(99) (time necessary to eliminate the 99% of phage particles) calculated. Results obtained showed that temperate phages revealed lower resistance than the virulent ones to the treatment temperatures. Biocides A, C, E and peracetic acid showed a notable efficiency to inactivate high concentrations of temperate and virulent L. delbrueckii phages. Biocide B evidenced, in general, a good capacity to eliminate the phage particles. Particularly for this biocide virulent phage Ib3 showed the highest resistance in comparison to the rest of temperate and virulent ones. On the contrary, biocide D and isopropanol presented a very low capacity to inactivate all phages studied. The efficiency of ethanol and hypochlorite was variable depending to the phages considered. These results allow a better knowledge and give useful information to outline more effective treatments to reduce the phage infections in dairy plants.
Assuntos
Fagos Bacilares/fisiologia , Laticínios/microbiologia , Desinfetantes/farmacologia , Temperatura Alta , Lactobacillus delbrueckii/virologia , Fagos Bacilares/efeitos dos fármacos , Qualidade de Produtos para o Consumidor , Laticínios/virologia , Microbiologia de Alimentos , Lactobacillus delbrueckii/crescimento & desenvolvimento , Lactobacillus delbrueckii/patogenicidade , Fatores de Tempo , Virulência , Inativação de VírusRESUMO
The bacteriophages Cb1/204 and Cb1/342 were obtained by induction from the commercial strain Lactobacillus delbrueckii subsp. lactis Cb1, and propagated on Lactobacillus delbrueckii subsp. lactis 204 (Lb.l 204) and Lactobacillus delbrueckii subsp. bulgaricus 342 (Lb.b 342), respectively. By cross sensitivity, it was possible to detect a delay in the lysis of Lb.l 204 with Cb1/342 phage, while the adsorption rate was high (99.5%). Modified and unmodified phages were isolated using phage Cb1/342 and strain Lb.l 204. The EOP (Efficiency of Plaquing) values for the four phages (Cb1/204, Cb1/342, Cb1/342modified and Cb1/342unmodified) suggested that an R/M system modified the original temperate phage, and the BglII-DNA restriction patterns of these phages might point out the presence of a Type II R/M system. Also, the existence of a Type I R/M system was demonstrated by PCR and nucleotide sequence, being the percentages of alignment homology with Type I R/M systems reported previously higher than 95%. In this study it was possible to demonstrate that the native phage resistant mechanisms and the occurrence of prophages in commercial host strains, contribute strongly to diversify the phage population in a factory environment.
Assuntos
Bacteriófagos/fisiologia , Enzimas de Restrição-Modificação do DNA/metabolismo , DNA Bacteriano/genética , Lactobacillus delbrueckii/virologia , Sequência de Bases , Enzimas de Restrição-Modificação do DNA/genética , Regulação Bacteriana da Expressão Gênica/fisiologia , Dados de Sequência MolecularRESUMO
AIMS: Frequency of lysogeny in Lactobacillus delbrueckii strains (from commercial and natural starters) and preliminary characterization of temperate bacteriophages isolated from them. METHODS AND RESULTS: Induction of strains (a total of 16) was made using mitomycin C (MC) (0.5 mug ml(-1)). For 37% of the MC-treated supernatants, it was possible to detect phage particles or presence of killing activity, but only two active bacteriophages were isolated. The two temperate phages isolated were prolate-headed phages which belonged to group c of Lact. delbrueckii bacteriophages classification. Different DNA restriction patterns were obtained for each phage, while the structural protein profiles and packaging sites were identical. Distinctive one-step growth curves were exhibited by each phage. An influence of calcium ions was observed for their lysis in broth but not on the adsorption levels. CONCLUSIONS: Our study showed that lysogeny is also present in Lact. delbrueckii strains, including commercial strains. SIGNIFICANCE AND IMPACT OF THE STUDY: Commercial strains could be lysogenic and this fact has a great practical importance since they could contribute to the dissemination of active-phage particles in industrial environments.