RESUMO
This review focuses on the impact of bacteriophages on the manufacture of dairy foods. Firstly, the impact of phages of lactic acid bacteria in the dairy industry, where they are considered enemies, is discussed. The sources of phage contamination in dairy plants are detailed, with special emphasis on the rise of phage infections related to the growing use of cheese whey as ingredient. Other topics include traditional and new methods of phage detection, quantification and monitoring, and strategies of phage control in dairy plants, either of physical, chemical or biological nature. Finally, the use of phages or purified phage enzymes as allies to control pathogenic bacteria in the food industry is reviewed.
Assuntos
Bacteriófagos/isolamento & purificação , Laticínios/virologia , Bacteriófagos/classificação , Bacteriófagos/genética , Laticínios/microbiologia , Indústria Alimentícia , Lactobacillales/metabolismo , Lactobacillales/virologiaRESUMO
UNLABELLED: Two bacteriophages, isolated from faeces, were assayed as biocontrol agents of pathogenic Escherichia coli during milk fermentation. Phage DT1 was tested on the strain E. coli DH5α, one enteropathogenic E. coli (EPEC) strain and one Shiga toxigenic E. coli O157:H7 (STEC) strain. Phage DT6 was tested on two STEC strains (O157:H7 and non-O157). One additional assay was performed by using a cocktail of both phages against the O157:H7 STEC strain. Streptococcus thermophilus 10-C, the strain used as lactic starter, reached 10(9) CFU ml(-1) after 4 h, while pH values fell to 4·5 after 8 h, regardless of the presence of E. coli strains and/or phages. In absence of phages, E. coli strains reached 4-6 log CFU ml(-1) at 5-6 h. Escherichia coli DH5α and O157:H7 STEC strains were rapidly and completely inactivated by phage DT1 and phage cocktail, respectively, while O157:H7 STEC was completely inactivated either by DT1 or by DT6, after 8 h. The EPEC strain was not detected at 1 h (<10 CFU ml(-1) ) but grew afterwards, though at lower rates than without phage. For non-O157:H7 STEC, reductions lower than 1 log CFU ml(-1) were observed for all sampling times. Phages DT1 and DT6, either individually or as a cocktail, effectively reduce O157:H7 STEC counts during milk fermentation, without compromising the starter culture performance. SIGNIFICANCE AND IMPACT OF THE STUDY: Coliphages DT1 and DT6, isolated from faeces and selected on the basis of their host range, showed to be valuable tools for the control of pathogenic Escherichia coli during milk fermentation, without compromising the starter culture performance. Both phages, either individually or as a cocktail, may function as an extra safety barrier beyond traditional pasteurization, effectively reducing O157:H7 Shiga toxin-producing Escherichia coli (STEC) counts during early growth, thus avoiding Shiga toxin production and accumulation.
Assuntos
Agentes de Controle Biológico , Colífagos , Escherichia coli O157/virologia , Fezes/virologia , Leite/microbiologia , Escherichia coli Shiga Toxigênica/virologia , Animais , Bovinos , Escherichia coli O157/crescimento & desenvolvimento , Fermentação , Escherichia coli Shiga Toxigênica/crescimento & desenvolvimento , Streptococcus thermophilus/crescimento & desenvolvimentoRESUMO
The effect of high pressure homogenization (HPH) with respect to a traditional heat treatment on the inactivation, growth at 8°C after treatments, and volatile profile of adventitious Leuconostoc strains isolated from Cremoso Argentino spoiled cheeses and ingredients used for their manufacture was evaluated. Most Leuconostoc strains revealed elevated resistance to HPH (eight passes, 100 MPa), especially when resuspended in skim milk. Heat treatment was more efficient than HPH in inactivating Leuconostoc cells at the three initial levels tested. The levels of alcohols and sulfur compounds increased during incubation at 8°C in HPH-treated samples, while the highest amounts of aldehydes and ketones characterized were in heated samples. Leuconostoc cells resuspended in skim milk and subjected to one single-pass HPH treatment using an industrial-scale machine showed remarkable reductions in viable cell counts only when 300 and 400 MPa were applied. However, the cell counts of treated samples rose rapidly after only 5 days of storage at 8°C. The Leuconostoc strains tested in this work were highly resistant to the inactivation treatments applied. Neither HPH nor heat treatment assured their total destruction, even though they were more sensitive to the thermal treatment. To enhance the inhibitory effect on Leuconostoc cells, HPH should be combined with a mild heat treatment, which in addition to efficient microbial inactivation, could allow maximal retention of the physicochemical properties of the product.
Assuntos
Queijo/microbiologia , Manipulação de Alimentos/métodos , Temperatura Alta , Leuconostoc/fisiologia , Pressão , Contagem de Colônia Microbiana , Contaminação de Alimentos/análise , Microbiologia de Alimentos , Conservação de Alimentos/métodos , Tecnologia de Alimentos/métodos , Humanos , Leuconostoc/crescimento & desenvolvimento , Leuconostoc/metabolismo , Viabilidade Microbiana , Fatores de TempoRESUMO
Temperate bacteriophages Ñ iLp84 and Ñ iLp1308, previously isolated from mitomycin C-induction of Lactobacillus paracasei strains 84 and CNRZ1308, respectively, were tested for their resistance to several physical and chemical treatments applied in dairy industry. Long-term survival at 4 °C, -20 °C and -80 °C, resistance to either thermal treatments of 63 °C, 72 °C and 90 °C, high pressure homogenization (HPH, 100 MPa) or classic (ethanol, sodium hypochlorite and peracetic acid) and new commercial sanitizers, namely 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), were determined. Phages were almost completely inactivated after eight months of storage at 25 °C, but viability was not affected at 4 °C, -20 °C or -80 °C. Both phages tolerated well HPH treatments. Phage iLp1308 showed higher thermal resistance than Ñ iLp84, but neither resisted 90 °C for 2 min. Best chemical inactivation was accomplished using peracetic acid or biocides A, C and E, whereas biocides B and D were completely ineffective. These results help to improve selection of chemical agents and physical treatments to effectively fight against phage infections in dairy plants.
Assuntos
Bacteriófagos/química , Bacteriófagos/efeitos dos fármacos , Desinfetantes/farmacologia , Lactobacillus/virologia , Esterilização/métodos , Bacteriófagos/crescimento & desenvolvimento , Contaminação de Alimentos/prevenção & controle , Microbiologia de Alimentos , Temperatura Alta , Pressão , Inativação de Vírus/efeitos dos fármacosRESUMO
AIMS: To isolate and characterize bacterial strains derived from Lactobacillus casei and Lactobacillus paracasei strains and resistant to phage MLC-A. METHODS AND RESULTS: Two of nine assayed strains rendered resistant mutants with recovery efficiencies of 83% (Lact. paracasei ATCC 27092) and 100% (Lact. casei ATCC 27139). DNA similarity coefficients (RAPD-PCR) confirmed that no significant genetic changes occurred while obtaining resistant mutants. Neither parent nor mutant strains spontaneously released phages. Phage-resistant mutants were tested against phages PL-1, J-1, A2 and MLC-A8. Lactobacillus casei ATCC 27092 mutants showed, overall, lower phage resistance than Lact. paracasei ATCC 27092 ones, but still higher than that of the parent strain. Lactobacillus paracasei ATCC 27092 mutants moderately adsorbed phage MLC-A only in calcium presence, although their parent strain successfully did it with or without calcium. Adsorption rates for Lact. casei ATCC 27139 and its mutants were highly influenced by calcium. Again, phage adsorption was higher on the original strain. CONCLUSIONS: Several isolates derived from two Lact. casei and Lact. paracasei strains showed resistance to phage MLC-A but also to other Lact. casei and Lact. paracasei phages. SIGNIFICANCE AND IMPACT OF THE STUDY: This study highlights isolation of spontaneous bacteriophage-resistant mutants from Lact. casei and Lact. paracasei as a good choice for use in industrial rotation schemes.
Assuntos
Bacteriófagos/fisiologia , Lacticaseibacillus casei/isolamento & purificação , Lactobacillus/isolamento & purificação , Tipagem de Bacteriófagos , Cálcio/metabolismo , DNA Bacteriano/genética , Lactobacillus/genética , Lactobacillus/virologia , Lacticaseibacillus casei/genética , Lacticaseibacillus casei/virologia , Mutação , Fenótipo , Técnica de Amplificação ao Acaso de DNA Polimórfico , Internalização do VírusRESUMO
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
Bacteriophage infection of lactic acid bacteria (LAB) constitutes one of the major problems in the dairy industry, causing economic losses and a constant risk of low quality and/or unsafe foods. The first step in the phage biology is the adsorption on the host cell surface. In a previous study, a remarkable thermal, chemical and photocatalytic resistance was demonstrated by four phages of Lactobacillus plantarum (ATCC 8014-B1, ATCC 8014-B2, FAGK1 and FAGK2). In the present work, these phages were used to characterize the adsorption process on L. plantarum ATCC 8014. Clearly, the characterization of this process could increase the possibilities of design useful strategies in order to prevent phage infections. The influence of Ca(2+), temperature, pH and physiological cell state on phage adsorption was investigated. Burst sizes of phages ATCC 8014-B1 and ATCC 8014-B2 were 60 and 83 PFU/infective centre, respectively. The four phages exhibited a high infectivity even at pH 4 and pH 11. Calcium or magnesium ions were not indispensable for cell lysis and plaque formation, and more than 99% of phage particles were adsorbed either in the presence or absence of Ca(2+), after 15 min at 37 degrees C. Phage adsorption was only partially affected at 50 degrees C, while reached its maximum between 30 and 42 degrees C. The highest adsorption values (99.9%) were observed from pH 5 to 7, after 30 min at 37 degrees C. Adsorption rates decreased after the thermal inactivation of cells, though, when 20 microg/ml of chloramphenicol was used, adsorption values were similar on treated and untreated cells. All these results showed that the adsorption process was only partially affected by a few conditions: thermally killed host cells, an incubation temperature of 50 degrees C and pH values of 9 and 10. Nevertheless, and unfortunately, those conditions are not commonly applied during fermented food manufacturing, thus restricting highly the application of strategies currently available to reduce phage infections in industrial environments. This work also contributes to increase the currently knowledge on the biological aspects of L. plantarum bacteriophages.
Assuntos
Fagos Bacilares/patogenicidade , Meio Ambiente , Microbiologia de Alimentos , Lactobacillus plantarum/virologia , Ligação Viral , Inativação de Vírus , Adsorção/efeitos dos fármacos , Fagos Bacilares/genética , Fagos Bacilares/fisiologia , Cálcio/farmacologia , Cloranfenicol , Fermentação , Manipulação de Alimentos , Concentração de Íons de Hidrogênio , Magnésio/farmacologia , Temperatura , Ligação Viral/efeitos dos fármacos , Inativação de Vírus/efeitos dos fármacosRESUMO
AIMS: To evaluate the phage diversity in the environment of a dairy industry which manufactures a product fermented with a probiotic strain of Lactobacillus paracasei. METHODS AND RESULTS: Twenty-two Lact. paracasei phages were isolated from an industrial plant that manufactures a probiotic dairy product. Among them, six phages were selected based on restriction profiles, and two phages because of their notable thermal resistance during sample processing. Their morphology, host range, calcium dependency and thermal resistance were investigated. All phages belonged to the Siphoviridae family (B1 morphotype), were specific for Lact. casei and paracasei strains showing identical host spectrum, and only one phage was independent of calcium for completing its lytic cycle. Some of the phages showed an extraordinary thermal resistance and were protected by a commercial medium and milk. CONCLUSIONS: Phage diversity in a probiotic product manufacture was generated to a similar or greater extent than during traditional yogurt or cheese making. SIGNIFICANCE AND IMPACT OF THE STUDY: This work emphasizes probiotic phage infections as a new ecological situation beyond yogurt or cheese manufactures, where the balanced coexistence between phages and strains should be directed toward a favourable state, thus achieving a successful fermentation.
Assuntos
Bacteriófagos/isolamento & purificação , Indústria de Laticínios , Lactobacillus/virologia , Apoptose/efeitos dos fármacos , Bacteriófagos/genética , Bacteriófagos/crescimento & desenvolvimento , Cálcio/farmacologia , DNA Viral/análise , Eletroforese em Gel de Ágar , Microbiologia Ambiental , Manipulação de Alimentos , Temperatura Alta , Cinética , Microscopia Eletrônica , Mapeamento por Restrição , Esterilização/métodosRESUMO
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.
Assuntos
Bacteriófagos/fisiologia , Lactobacillus delbrueckii/virologia , Lisogenia/fisiologia , Bacteriólise/fisiologia , Bacteriófagos/genética , Bacteriófagos/ultraestrutura , Cálcio/farmacologia , DNA Viral/genética , DNA Viral/isolamento & purificação , Lactobacillus delbrueckii/fisiologia , Microscopia Eletrônica , Mapeamento por Restrição , Proteínas Virais/genéticaRESUMO
AIMS: Characterization of four virulent Lactococcus lactis phages (CHD, QF9, QF12 and QP4) isolated from whey samples obtained from Argentinean cheese plants. METHODS AND RESULTS: Phages were characterized by means of electron microscopy, host range and DNA studies. The influence of Ca(2+), physiological cell state, pH and temperature on cell adsorption was also investigated. The double-stranded DNA genomes of these lactococcal phages showed distinctive restriction patterns. Using a multiplex PCR, phage QP4 was classified as a member of the P335 polythetic species while the three others belong to the 936 group. Ca(2+) was not needed for phage adsorption but indispensable to complete cell lysis by phage QF9. The lactococci phages adsorbed normally between pH 5 and pH 8, and from 0 degrees C to 40 degrees C, with the exception of phage QF12 which had an adsorption rate significantly lower at pH 8 and 0 degrees C. CONCLUSIONS: Lactococcal phages from Argentina belong to the same predominant groups of phages found in other countries and they have the same general characteristics. SIGNIFICANCE AND IMPACT OF THE STUDY: This work is the first study to characterize Argentinean L. lactis bacteriophages.