Adherence and internalisation of Lactococcus lactis M4 towards human colorectal cancer cell line, Caco-2

Aims@#Lactococcus lactis is a non-colonizing, generally-regarded as safe (GRAS) lactic acid bacteria that has been frequently studied as a potential vector for bactofection. To mediate bactofection, a series of interaction between the bacteria and the host cell needs to occur. This study aims to investigate the in vitro bacterial-cell interaction between a locally-isolated L. lactis M4 strain with human colorectal cancer line, Caco-2.@*Methodology and results@#Bacterial interaction was evaluated via adherence and internalisation assays. A 250:1 ratio of bacteria to cancer cell was selected as the optimum multiplicity of infection for all assays. After 2 h, L. lactis M4 was able to adhere to and internalise into Caco-2 cells at comparable rates to commercial strains L. lactis NZ9000 and MG1363. @*Conclusion, significance and impact of study@#Findings from this study showed that this strain has similar interaction properties with the commercial strains and would make a promising candidate for future bactofection studies and development of bacteria-mediated DNA vaccination against various diseases.

Evaluation of lactase activity in probiotic cultures containing inulin rich plant extracts as prebiotic sources

@#Aims: The present study deals with the isolation and identification of lactase producing probiotic strains from camel and sheep milk, determination of the enzyme activity by β-galactosidase assay (Miller Assay) in the presence of garlic, peas, onion and leeks extracts containing inulin as a prebiotic component. Methodology and results: The two isolates were screened for lactase producing ability to degrade lactose on MRS agar at 37 °C. These were identified as Lactococcus lactis from camel (Marecha) milk and Lactobacillus casei from sheep (Kajli) milk through morphological and biochemical tests using MRS medium. The optimized pH and temperature of both strains were 6 and 35 °C, respectively. Among the three concentrations used (0.2%, 0.4%, 0.8%), the optimal concentration of inulin rich onion and leeks extracts was 0.8% for maximum growth of L. casei and of the peas extract for L. lactis growth. 0.2% garlic extract was more effective prebiotic source for L. lactis growth. 0.8% commercial inulin used as a positive control was less effective as compared to plant extracts used in the study. With o-nitrophenyl-β-Dgalactoside) used as a substrate in the enzyme assay, maximum lactase activity obtained with 0.8% concentration of garlic extract is 7.10 Miller Units as compared to the peas extract with 6.17 Miller Units from L. lactis. Lactobacillus casei has produced more lactase, 6.85 Miller units with onion extract than with leeks extract, 6.43 Miller Units. Pure commercial inulin used as a control has given maximum enzyme activity as 9.14 Miller Units at 0.2% concentration. Conclusion, significance and impact of the study: It is concluded that the extracted prebiotic may enhance lactase activity of the probiotics to supplement the development of food products for lactose intolerant patients.

Protective effect of staphylococcal nuclease on 2, 4, 6-trinitro-benzene sulfonic acid-induced colitis in mice / 中国药科大学学报

@#To explore the improving effect and mechanism of staphylococcal nuclease(SNase)-mediated degradation of neutrophil extracellular traps(NETs)on 2, 4, 6-trinitro-benzene sulfonic acid(TNBS)-induced colitis in mice. The model of colitis in female BALB/c mice was established by intrarectal injection of 2. 5% TNBS solution, and SNase loaded by Lactococcus lactis(L. lactis)were orally administrated for 6 days. To investigate the effect of SNase-mediated degradation of neutrophil extracellular traps on colitis in mice, the experiment was divided into control group, TNBS model group, NZ900 group and L. lactis pCYT: SNase group. The daily body weight, stool consistency and bleeding of mice were observed. The pathological condition of HE in colon group was detected. The activity of MPO and the mRNA expression level of inflammatory cytokines in each group were measured, and the concentration of inflammatory factors in serum was detected. The expression of NETs level marker citH3 in colon tissue was determined by immunohistochemistry. The results showed that SNase loaded by lactis acid bacteria could alleviate the weight loss, disease activity index score, colonic length and pathological damage induced by TNBS in mice, and reduce the levels of inflammatory cytokines in serum and colonic tissue, inhibit the activity of MPO and the expression of Ly6G and citH3 in colon tissue. The preliminary mechanism showed that SNase could down-regulate the expression of inflammatory cytokines and reduce the content of NETs markers to alleviate colitis in mice.

Construction and verification of Lactococcus lactis NZ9000 genome-scale metabolic model / 生物工程学报

With the advent of the post-genomic era, metabolic engineering of microorganisms plays an increasingly important role in industrial production. The genome-scale metabolic model (GSMM) integrates all known metabolic information in the organism to provide an optimal platform for global understanding of the metabolic state of the organism and rational guidance for metabolic engineering. As a model strain, Lactococcus lactis NZ9000 plays an important role in industrial fermentation, but there is still no specific genome-scale metabolic model for it. Based on genomic function annotation and comparative genomics, we constructed the first genome-scale metabolic model iWK557 of L. lactis NZ9000, which contains 557 genes, 668 metabolites, and 840 reactions, and further verified at both qualitative and quantitative levels, to provide a good tool for rationally guiding metabolic engineering.

Ação in vivo de Lactococcus lactis subsp. lactis com potencial probiótico na estabilização de células cancerígenas no epitélio colorretal / In vivo action of Lactococcus lactis subsp. lactis with probiotic potential in the stabilization of cancer cells in the colorectal epithelium

Na última década, a ciência contribuiu significativamente para inúmeros avanços em relação ao tratamento e prevenção do câncer colorretal (CCR), porém, a prevalência global e a taxa de mortalidade permanecem elevadas. Há relatos sobre efeitos benéficos de espécies de Bifidobacterium e Lactobacillus com potencial probiótico na prevenção de CCR. No entanto, a bactéria probiótica Lactococcus lactis subps. lactis é comumente utilizada para fins industriais, não havendo comprovações in vivo sobre seu potencial anticarcinogênico. Visto o interesse emergente dos efeitos benéficos dos probióticos a fim de prevenir ou tratar o CCR, o presente estudo objetivou explorar os efeitos de L. lactis subsp. lactis sobre o CCR. Ratos Wistar receberam doses subcutâneas de 1,2 dimetilhidrazina (DMH) e suspensão de L. lactis subsp. lactis por via oral. Após 20 semanas, os tecidos intestinais foram analisados e de acordo com o resultado, o isolado demonstrou potencial anticarcinogênico contra CCR.

Risk Assessment of Lactococcus lactis W58 Used as "Other Substances"

The Norwegian Scientific Committee for Food Safety (Vitenskapskomiteen for mattrygghet, VKM) has, at the request of the Norwegian Food Safety Authority (Mattilsynet; NFSA), assessed the risk of "other substances" in food supplements sold in Norway. These risk assessments will provide NFSA with the scientific basis while regulating the addition of “other substances” to food supplements and other foods. "Other substances" are described in the food supplement directive 2002/46/EC as substances other than vitamins or minerals that have a nutritional and/or physiological effect. It is added mainly to food supplements, but also to other foods. VKM has not in this series of risk assessments of "other substances" evaluated any claimed beneficial effects from these substances, only possible adverse effects. The present report is a risk assessment of Lactococcus lactis W58, and it is based on previous risk assessments and articles retrieved from a literature search. The risk of L. lactis W58 was assessed for the general population. However, in previous assessments of “probiotics” published by VKM, concerns have been identified for specific groups. Therefore, the risk was assessed for the age group with immature gastro-intestinal microbiota (age group 0-36 months), population with mature gastro-intestinal microbiota (>3 years) and vulnerable groups independent of age. VKM has also assessed the risk of L. lactis W58 in food supplements independent of the dose and have assessed exposure in general terms. Other sources of L. lactis W58, such as foods, have not been included in the present risk assessment. VKM concludes that it is unlikely that L. lactis W58 causes adverse health effects in the general healthy population with mature gastro-intestinal tract. However, no data on long-term adverse effects on infants and young children were identified. As evidence is accruing that the early microbial composition of the neonatal gut is important for the development of the gut microbiota and the immune system of the growing child, it is not possible to exclude that a daily supply of a single particular bacterial strain over a prolonged period of time to an immature gastro-intestinal tract may have long-term, although still unknown, adverse effects on that development.

Effects of heme oxygenase-1 recombinant Lactococcus lactis on the intestinal barrier of hemorrhagic shock rats

This study aimed to investigate the effects of heme oxygenase-1 recombinant Lactococcus lactis (LL-HO-1) on the intestinal barrier of rats with hemorrhagic shock. One hundred Sprague-Dawley male rats (280–320 g) were randomly divided into healthy control group (N group) and hemorrhagic shock group (H group). Each group was subdivided into HO1t, HO2t, HO3t, PBS and LL groups in which rats were intragastrically injected with LL-HO-1 once, twice and three times, PBS and L. lactis (LL), respectively. The mortality, intestinal myeloperoxidase (MPO) activity, intestinal contents of TNF-α, IL-10 and HO-1, and intestinal Chiu's score were determined. Results showed that in N group, the HO-1 content increased after LL-HO-1 treatment, and significant difference was observed in HO1t group and HO2t group (P<0.05). In H groups, MPO activity and Chiu's score decreased, but IL-10 content increased in LL-HO-1-treated groups when compared with PBS and LL groups (P<0.05). When compared with N group, the MPO activity reduced dramatically in LL-HO-1-treated groups. Thus, in healthy rats (N group), intragastrical LL-HO-1 treatment may increase the intestinal HO-1 expression, but has no influence on the intestinal barrier. In hemorrhagic shock rats, LL-HO-1 may significantly protect the intestinal barrier, and repeating the intragastrical LL-HO-1 treatments twice has the most obvious protection.

A Case of Septic Shock Following Catheter-related Infection Caused by Lactococcus lactis subsp. lactis in an Adult

Lactococcus lactis is a gram-positive cocci used extensively in the dairy industry, but considered an unusual pathogen in humans. Among its five subspecies, L. lactis subsp. lactis in particular has rarely been reported as a pathogen. We report a case of septic shock caused by L. lactis subsp. lactis in an adult patient. A 64-yr-old male patient was admitted to outpatient clinics, with chief complaints of fever and chills for one week after convalescent hospital admission. He had severe ileus requiring surgery. He had a peripherally inserted central catheter from convalescent hospital, which was immediately removed. From two sets of blood and catheter tip cultures, we identified L. lactis subsp. lactis using the Vitek 2 system (bioMerieux Inc., USA), and confirmed this result by 16S rRNA sequencing. The patient was empirically treated with ciprofloxacin, and he recovered and was discharged.

Metabolitos de bacterias lácticas antagónicos al crecimiento de microorganismos cariogénicos / Antagonistic activity of lactic acid bacteria metabolites toward cariogenic microorganisms growth

Aislar, purificar y conservar cepas de Streptococcus spp. y lactobacillus spp. de la cavidad bucal y enfrentarlas "in vitro" a bacterias lácticas. Materiales y métodos: se seleccionaron individuos con caries y se recolectaron muestras de saliva. Para recuperar Streptococcus spp. se empleó el medio Mitis Salivarius (Difco, Detroit, MI, Estados Unidos) y para Lactobacillus spp. se usó Rogosa (Blokar Diagnostics, Beauvais, Francia). Como cepas productoras de bacteriocinas se utilizaron 7 cepas de Lactococcus lactis subsp. lactis, 2 de Leuconostoc mesenteroides y 1 de Lactococcus lactis subsp. diacetylactis. La actividad antagónica de las bacterias lácticas al crecimiento in vitro de bacterias cariogénicas se determinó con el método de difusión en agar. Resultado: el desarrollo y la multiplicación de las cepas de Streptococcus spp. de origen bucal ensayadas se vieron afectados por la presencia de metabolitos generados por las cepas de Lactococcus lactis subsp. lactis. Conclusión: el crecimiento de las cepas de Streptococcus subsp. fue inhibido por efecto de L. lactis subsp. lactis...

Bioencapsulation of probiotic Lactococcus lactis subsp. lactis on Artemia franciscana nauplii: Effects of encapsulation media on Nauplii survival and probiotic recovery

Aims: This study aimed to investigate the suitability and efficacy of various encapsulation media in bioencapsulating the probiotic Lactococcus lactis subsp. lactis in Artemia franciscana nauplii. The impact of the encapsulation media on nauplii survival and probiotic recovery was also determined. Methodology and results: Various encapsulation media (sodium alginate, palm oil, starch, gum Arabic and gelatin) were prepared by dissolving the respective media in artificial sea water. Each media was prepared in four different concentrations: 0.25, 0.5, 1.0 and 2.0 g/L. To determine the suitability of encapsulation media on the survivability of A. franciscana, survival rate (SR) of Artemia nauplii was determined after 8 hours post-encapsulation. Instar II stage Artemia nauplii at 1 nauplii per mL was used for each replicate. The result revealed that A. franciscana reached 100 % SR in the encapsulation media at ≤ 0.5 g/L. All media enabled > 23 % recovery of L. lactis subsp. lactis from encapsulated A. franciscana, which is similar (p > 0.05) to the recovery of free-cells (non-encapsulated) of L. lactis subsp. lactis. Noticeably in sodium alginate (E1) treatment, the total counts of L. lactis subsp. lactis in bioencapsulated A. franciscana were the highest among others, accounting for 2.44 × 107 CFU/mL per A. franciscana tissue homogenate. Conclusion, significance and impact of study: Artemia nauplii bioencapsulated with L. lactis subsp. lactis using 0.5 g/L sodium alginate as encapsulation medium has the highest SR for nauplii and bioencapsulation efficiency, respectively. This result provides a basic guideline for Artemia bioencapsulation in fish/shrimp larval culture.

Polymicrobial Peritonitis with Lactococcus lactis in a Peritoneal Dialysis Patient / 전남의대학술지

Lactococcus lactis (L. lactis) is an important gram-positive bacterium in dairy products. It is a rare cause of opportunistic infections with only four cases of Lactococcus peritoneal dialysis (PD) peritonitis reported in the literature. In Korea, L. lactis infection was first reported in a liver abscess patient in 2010; however, PD peritonitis with Lactococcus has not been reported in Korea. Recently, we experienced a case of Lactococcus-associated polymicrobial PD peritonitis. The patient was initially managed with broad-coverage antibiotics; however, owing to a poor response, the PD catheter was removed and the patient was switched to hemodialysis. We discuss this case and review the literature.

Polymicrobial Peritonitis with Lactococcus lactis in a Peritoneal Dialysis Patient / 전남의대학술지

Lactococcus lactis (L. lactis) is an important gram-positive bacterium in dairy products. It is a rare cause of opportunistic infections with only four cases of Lactococcus peritoneal dialysis (PD) peritonitis reported in the literature. In Korea, L. lactis infection was first reported in a liver abscess patient in 2010; however, PD peritonitis with Lactococcus has not been reported in Korea. Recently, we experienced a case of Lactococcus-associated polymicrobial PD peritonitis. The patient was initially managed with broad-coverage antibiotics; however, owing to a poor response, the PD catheter was removed and the patient was switched to hemodialysis. We discuss this case and review the literature.

Obtención de microcápsulas de tamaño controlado conteniendo Lactococcus lactis, utilizando mezclas de gelanos de alto y bajo acilo / Obtaining size-controlled microcapsules by ionic gelation with high and low acyl gellans containing Lactococcus lactis

Control the diameter of microcapsules obtained with functional biopolymer is a crucial parameter in the success of food applications, since it affects the protection of microencapsulated microorganism and also in the texture of the final product. The aim of this study was to assess the obtaining of controlled size microcapsules containing Lactococcus lactis, using mixtures of high acyl gellan (HA) and low acyl gellan (LA). A concentration of 0.2% (w/w) gellan was employed using a simple design, generating the following mixtures: 100HA/0.0LA, 0.0HA/100LA, 25HA/75LA, 50HA/50LA and 75HA/25LA. The diameter of the microcapsules, efficiency of microencapsulation and viability of the microencapsulated microorganism were studied in function of the speed of agitation (400-800 rpm) and surfactant concentration (sorbitan monooleate) (0.0-0.2%)v/v. The results indicated that mixtures with concentration equal or greater than 50% of HA gellan are not efficient for obtaining microcapsules, only the LA gellan and the mixture 25HA/75LA gave acceptable results. The viability of the microorganism and the efficiency of microencapsulation were descending function of the stirring speed and surfactant concentration. The microcapsules obtained had diameters not greater than 80 µm when the highest concentrations of surfactant (0.2% v/v) and stirring speed (800 rpm) were used, suggesting that the ionic gelation can be used to obtain microcapsules of controlled size (15-75 µm) containing Lactococcus lactis with high viability (83.32%) and high efficiency of microencapsulation (82.4%), which makes it feasible for use in food applications.

Controlar el diámetro de microcápsulas obtenidas con biopolímeros funcionales es un parámetro crucial en el éxito de aplicaciones alimentarias, ya que influye en la protección del microorganismo microencapsulado y también en la textura del producto final. El objetivo de este trabajo fue evaluar la obtención de microcápsulas de tamaño controlado conteniendo Lactococcus lactis, utilizando mezclas de gelana de alto (HA) y bajo acilo (LA). Se empleó una concentración de gelana de 0.2% p/p usando un diseño de mezclas simple, generando las siguientes mezclas, 100HA/0.0LA, 0.0HA/100LA, 25HA/75LA, 50HA/50LA, 75HA/25LA. El diámetro de las microcápsulas, la eficiencia de microencapsulación y la viabilidad del microorganismo microencapsulado fueron estudiadas en función de la velocidad de agitación (400-800 rpm) y concentración de surfactante (sorbitan monooleate) (0.0-0.2%)v/v. Los resultaron indicaron que las mezclas con concentración igual o superior al 50% de gelana de HA, no son eficientes para obtener microcápsulas; solamente dieron resultados aceptables la gelana de LA y la mezcla 25HA/75LA. La viabilidad del microorganismo y la eficiencia de microencapsulación variaron en función descendente de la velocidad de agitación y concentración de surfactante. Las microcápsulas obtenidas no presentaron diámetros superiores a 80 µm cuando se emplearon las mayores concentraciones de surfactante (0.2%) y velocidad de agitación (800 rpm), sugiriendo que la gelación iónica puede ser utilizada para obtener microcápsulas de tamaño controlado (15-75 µm) conteniendo Lactococcus lactis con alta viabilidad (83.32%) y eficiencia de microencapsulación (82.4%), cuando se utiliza la mezcla 25HA/75LA a 800 rpm y 0.2% v/v de surfactante, lo cual la hace factible para su uso en aplicaciones alimentarias.

Disponibilidad de fósforo soluble en ensilado experimental fermentado con una cepa de Lactococcus lactis subsp. lactis con alta actividad de fitasa / Soluble phosphorous availability in experimental silage fermented with a Lactococcus lactis subsp. lactis strain with high phytase activity

El fitato es la forma principal de almacenamiento de fósforo en semillas y granos, pero el fósforo unido a fitato resulta inaccesible para los peces. El objetivo del presente trabajo fue estudiar la eficacia de Lactococcus lactis subsp. lactis Tw34 para liberar fósforo de fitato en un alimento experimental que contenía harina de cereal. La bacteria inoculante del ensilado biológico se seleccionó por su alta actividad de fitasa y las mezclas se prepararon con productos de descarte de merluza (Merluccius hubbsi). Las harinas de girasol, maíz, trigo y cebada se mezclaron con desechos picados de merluza hasta alcanzar el 25% y se inocularon con la bacteria citada. Después de 7 días de fermentación a 18 ºC los mejores resultados se obtuvieron cuando se utilizó harina de cebada. La cepa fue capaz de reducir el valor del pH a 4,4 y aumentar cuatro veces la concentración inicial de fósforo libre. Los resultados sugieren que la fitasa de Lactococcus lactis subsp. lactis Tw34 puede aumentar la accesibilidad del fósforo en dietas basadas en harina de cereales, evitando los efectos negativos del ácido fítico en la biodisponibilidad de minerales y proteínas, y disminuyendo la liberación de residuos de fósforo.

Phytate is the main phosphorous storage form in grains and seeds, but phytate linked phosphorous is inaccessible for fishes. The purpose of this wok was to study the efficacy of Lactococcus lactis subsp. lactis Tw34 for liberating phosphorous in an experimental feed which contained cereal flour. The inoculating bacterium of the biological silage was selected due to its high phytase activity, and the mixtures were prepared with waste hake products (Merluccius hubbsi). Sunflower, corn, wheat and barley flours were mixed with waste ground hake products up to 25% and inoculated with the bacterium mentioned. After 7 days fermentation at 18 oC, the best results were obtained when barley flour was used. The strain was able to reduce the pH value to 4.4 and increase four times the initial concentration of free phosphorous. The results suggest that Lactococcus lactis subsp. lactis Tw34 can increase phosphorous accessibility in cereal flour based diets, avoiding the negative effects of phytic acid on the bioavailability of minerals and proteins, and decreasing the liberation of phosphorous residues.

Antimicrobial activity of the Nisin Z producer Lactococcus lactis subsp. lactis Lc08 against Listeria monocytogenes in skim milk / Atividade antimicrobiana de Lactococcus lactis subsp. lactics Lc08 produtor de Nisina Z contra Listeria monocytogenes em leite desnatado

The presented study aimed to verify the effect of different pH values, enzyme solutions and heat treatments on the antimicrobial activity of the bacteriocinogenic strain Lactococcus lactis subsp. lactis Lc08 and to test their antimicrobial activity against Listeria monocytogenes in reconstituted skim milk at refrigeration temperatures. This strain was previously described as a nisin Z producer and capable of inhibiting L. monocytogenes growth in in vitro tests. The antimicrobial activity of the bacteriocin cell-free supernatant of Lc08 was sensitive to enzyme treatments (except papain). The pH values and heating (65ºC for 30min, 75ºC for 15s) had no apparent effect on the antimicrobial activity of the bacteriocin produced by Lc08. Only treatment at autoclave conditions result in loss of their antimicrobial activity. Lc08 presented antimicrobial activity against L. monocytogenes in the milk system after 12h at 25ºC. No effect was found at 7ºC. The results show the application viability of the Lc08 in food systems as a biopreservative against L. monocytogenes.

O presente estudo teve como objetivo verificar o efeito de diferentes valores de pH, soluções enzimáticas e tratamentos térmicos na atividade antimicrobiana da cepa bacteriocinogênica Lactococcus lactis subsp. lactis Lc08 e testar sua atividade antagonista contra Listeria monocytogenes em leite desnatado reconstituído em diferentes temperaturas de estocagem. Essa cepa já foi descrita como produtora de nisina Z e capaz de inibir o desenvolvimento de L. monocytogenes em testes in vitro. A atividade antimicrobiana do sobrenadante de Lc08 contendo a bacteriocina produzida e livre de células foi sensível ao tratamento pelas enzimas testadas (exceto papaína). A aplicação de diferentes valores de pH e o tratamento térmico (65ºC por 30 min, 75ºC por 15s) não influenciaram na atividade antimicrobiana da bacteriocina produzida por Lc08. Apenas o tratamento em autoclave resultou em perda da sua capacidade em inibir o desenvolvimento de L. monocytogenes. A cepa Lc08 apresentou atividade antagonista contra L. monocytogenes em leite após período de estocagem de 12h a 25ºC. Não foi observado efeito a 7ºC. Os resultados mostram a viabilidade de aplicação da cultura Lc08 ou de sua bacteriocina em produtos lácteos como bioconservador contra L. monocytogenes.

An evaluation and partial characterization of a bacteriocin produced by Lactococcus lactis subsp lactis ST1 isolated from goat milk

A bacteriocin-like inhibitory substance producing Lactococcus lactis subsp lactis strain, ST1, isolated from goat milk of Iranian origin and with broad spectrum of activity and desirable technical properties was used for evaluating some futures of bacteriocin inhibitory activity. Cell growth and bacteriocin production studies were carried out in MRS medium incubated statically under uncontrolled pH condition. The antibacterial activity presented a primary metabolite pattern and showed a rapid decrease at the stationary phase. Microaerobiosis and capnophily growth conditions resulted in higher bacteriocin production while aerobiosis showed negative effect on both cell growth and bacteriocin production. Bacteriocin production, on the other hand, was favored in MRS broth (pH; 6.5) inoculated with 0.1 ml l-1 fresh culture when incubation was carried out at 30 °C. This indicated that the conditions resulted in higher levels of growth were frequently favoring bacteriocin production by ST1 as well. Decrease in activity, at the stationary growth phase, was much pronounced in favored growth condition. Nutrient depletion, deferent effect of low pH on bacteriocin production and/or protein degradation seemed more responsible for this phenomenon. The study also provided further data on new method for bacteriocin release from the cell wall of producer. It was clearly shown that both heating and ultrasound shock for 5 min at pH 2 could increase bacteriocin activity significantly. The release was more pronounced in the presence of 0.5% Tween80.

Increase of stress resistance in Lactococcus lactis via a novel food-grade vector expressing a shsp gene from Streptococcus thermophilus

The effects of the expression of a small heat shock protein (shsp) gene from Streptococcus thermophilus on stress resistance in Lactococcus lactis under different environmental stresses were investigated in this study. pMG36e-shsp, an expression vector, was first constructed by inserting a shsp open reading frame (ORF) cloned from S. thermophilus strain St-QC into pMG36e. Then, a food-grade expression vector, pMG-shsp, was generated by deleting the erythromycin resistance gene from pMG36e-shsp. The transformation rate of pMG-shsp was comparable to that of pMG36e-shsp when each of these two vectors was introduced into L. lactis. These results demonstrated that the shsp ORF could successfully used as a food-grade selection marker in both pMG-shsp and pMG36e-shsp. Furthermore, the growth characteristics were almost the same between L. lactis ML23 transformants harboring pMG36e or pMG-shsp. The survival rate of L. lactis ML23 expressing the shsp ORF were increased to 0.032%, 0.006%, 0.0027%, 0.03%, and 0.16% under the following environmental stresses: heat, acid, ethanol, bile salt and H2O2, respectively. These results indicated that the expression of the shsp gene in the food-grade vector pMG-shsp conferred resistance to environmental stresses without affecting the growth characteristics of L. lactis ML23.

Viabilidade de Staphylococcus aureus FRI S-6 e produção de SEB em queijo elaborado com adição de Lactobacillus rhamnosus e Lactococcus lactis / Staphylococcus aureus FRI S-6 viability and its SEB production in cheese produced with Lactobacillus rhamnosus and Lactococcus lactis

Staphylococcus aureus foi inoculado em queijos produzidos de forma estéril em laboratório, juntamente com Lactobacillus rhamnosus e Lactococcus lactis, isolados de queijo de coalho artesanal e identificados por PCR-ARDRA16S-23S. L. lactis foi capaz de reduzir a contagem de S. aureus no primeiro dia após produção (P<0,05) dos queijos de 3,3x10(7)UFC/g para 1,0x10(7)UFC/g. L. rhamnosus não impediu o crescimento de S. aureus. A presença das cepas acidoláticas, principalmente L. lactis, mostrou ainda potencial de inibição da produção de enterotoxina estafilocócica do tipo B, sendo que a concentração de enterotoxinas no 15° dia foi inferior ao limite de detecção pelo kit comercial utilizado. Concluiu-se que a presença das bactérias acidoláticas estudadas pode contribuir para a melhoria da qualidade sanitária de queijos artesanais.

Staphylococcus aureus was inoculated in cheese elaborated with Lactobacillus rhamnosus and Lactococcus lactis, produced in a sterile lab, previously isolated from Brazilian coalho artisanal cheese and identified by PCR-ARDRA16S-23S. L. lactis was able to reduce the staphylococcal count on the first day after cheese production (P<0.05) from 3.33x10(7)CFU/g to 1.0x10(7)CFU/g. L. rhamnosus did not inhibit S. aureus growth. The presence of lactic acid bacteria (LAB), mainly L. lactis, also showed evidences of inhibition of the enterotoxin B production, and the staphylococcal enterotoxins concentration on the 15th day was lower than the commercial kit threshold. Hence, the studied LAB may contribute to the sanitary quality of artisanal cheeses.

Physiochemical parameters optimization for enhanced nisin production by Lactococcus lactis (MTCC 440)

The influence of various physiochemical parameters on the growth of Lactococcus lactis sub sp. lactis MTCC 440 was studied at shake flask level for 20 h. Media optimization (MRS broth) was studied to achieve enhanced growth of the organism and also nisin production. Bioassay of nisin was done with agar diffusion method using Streptococcus agalactae NCIM 2401 as indicator strain. MRS broth (6 percent, w/v) with 0.15ìg/ml of nisin supplemented with 0.5 percent (v/v) skimmed milk was found to be the best for nisin production as well as for growth of L lactis. The production of nisin was strongly influenced by the presence of skimmed milk and nisin in MRS broth. The production of nisin was affected by the physical parameters and maximum nisin production was at 30(0)C while the optimal temperature for biomass production was 37(0)C.

Utilización de iidrocoloides en bebida láctea tipo kumis / Use of ydrocollooids in dairy beverages kums type

El objetivo del presente trabajo es evaluar la mezcla más adecuada de los estabilizantes goma guar, goma xantan y carragenina iota en la bebida láctea tipo kumis. Se lleva a cabo un estudio preliminar que define la mezcla óptima de estos estabilizantes, utilizando el nivel máximo de 0,5%, aprobado por el Ministerio de la Protección Social de la República de Colombia, la variable respuesta medida es viscosidad. Definida la mejor mezcla, es utilizada en tres concentraciones (T1: 0,08%; T2: 0,1% y T3: 0,12%), y comparada con un control (T4: sin adición de estabilizante) en el producto lácteo tipo kumis. La mezcla óptima encontrada fue de 93% goma xantan y 7% goma guar, y los parámetros empleados para su evaluación son fisicoquímicos reométricos y sensoriales. Los resultados fisicoquímicos corresponden a los estándares aceptados para esta bebida. En lo referente a las pruebas de viscosidad y deformación, el tratamiento que muestra los mejores resultados es la concentración 0,12%; que corresponde al tratamiento T3. El análisis sensorial indica que T3 es el tratamiento más aceptado, junto con el control. Según los resultados, el nivel óptimo de concentración de la mezcla de hidrocoloides es 0,12%; con diferencia significativa (p<0,05) frente a los otros tratamientos.

The aim of this study was to evaluate the most appropriate mix of stabilizers: guar gum, xanthan gum and iota carrageenan in dairy beverages kumis type. A preliminary study defining the optimal mix between these stabilizers was carried, using the maximum level of 0.5% approved by the Colombian Ministry of Health. Viscosity was used as response variable. Once the best mix was selected it was used in three concentrations (T1: 0.08%, T2: 0.1% and T3: 0.12%), and compared to a control (T4: no added stabilizer) in the milk product type kumis. The parameters used for the assessment of the optimal mix of 93% xanthan gum and 7% guar gum were physicochemical, viscosity and sensory. Physicochemical results fitted to the accepted standards for such milk drinks. In terms of viscosity and strain tests, the treatment showing the best results was T3. Sensory analysis indicated that T3 is the more accepted treatment with the control. Based on the results obtained, the optimum concentration level of the hydrocolloids mixture was 0.12% of the mixture with a p <0.05.