Lactococcus lactis

Description and significance
"Lactococcus Lactis" is a non pathogenic,Gram-positive bacteria known as one of the most important microbes in dairy food production. Lactococci lactis, which is closely related to the strptococcus genus, has two subspecies, Lactococcus lactis subsp. and Lactococcus cremoris subsp. They are used to make soft and hard cheese respectively. L.Lactis group in pairs or short chains with a length of about 1.2µm by 1.5µm. Lactococci are immotile, nonsporelating, and are usually spherical or ovoid cells. L. Lactis is the starter culture for the production of fermented dairy products such as milk, cheese, yogurt, etc. Lactococcus is considered a opurtunistic pathogen, because of its ability to convert almost all it's sugar into lactic acid. Lactic acid produced by the the microbe curdles the milk which seperates into curds inturn are used to produce cheese and whey.In nature, L. lactis is found to be inactive on plant surfaces and on to multiply in the gastrointestinal tract after being swallowed by an animal.

Genome structure
L.lactis genome has been sequenced to investigate its essential role in dairy fermentation. Unexpected features of the analyzed genome have been disclosed such as genes that suggest the bacterium can perform aerobic respiration and can undergo horizontal gene transmission by the process of transformation. The continued research unveils the characteristics of L.lactis that can be manipulated, in particular those that can improve the flavor, texture, and preservation of 10 million tons of cheese produced annually. Knowledge of the genome sequence will also facilitate current and future work that aims to exploit Lactococcus lactis for a variety of medical and health maintenance applications. L. lactis is a circular chromosome, its genome contains 2,365,589 base pairs. Protein-coding genes represent 86% of the genome, stable RNA 1.4%, and noncoding regions 12.6%. These percentages are very similar to other bacteria genome. Biological functions have been assigned to 64% of the genome, 20.1% have similar genome to other unknown proteins, however, 15% of the genome is unidentifiable. This 15% is hypothesized to contain the traits specific to L.lactis.

Cell structure and metabolism
"Lactococcus lactis" functions through both aerobic and anaerobic metabolic pathways. The primitive source of energy is produced anaerobicly, which results in the accumulation of lactic acid. The deprivation of oxygen leads the glycolysis process to breakdown carbohydrates into pryvate which then convert into lactic acid. This process is only possible through the production of the lactate dehydrogenase enzyme and NAD. Lactate is transported to the median which causes the efflux of protons resulting in the appropriate membrane potential for energy production. The lactic acid produced is used for food production.Aerobic respiration is also used as a supplemental metabolic process. When oxygen and a heme source is present new traits are observed such as increased growth index, resistance to oxidative and acid stress, and long-termed endurance at low temperatures. Along with heme source, the presence of membrane bound NADH dehydrogenase oxidizes NADH and increases the cell growth and production of proteins and vitamins.

Ecology
Lactococcus lactis is an oportunistic bacteria, in nature, can be found on wild plants and consequently in the gastrointesinal tract of animals, such as cows, and are ultimately inoculated in milk. A study has been done showing the survival rate of ingested L.lactis. Data has shown that only 2% of all the ingested bacteria is found in the remaining feces if consumed within three days.


 * Note : Work on this section is still in progress***

Pathology
Lactococcus lactis is a non-pathogenic bacteria.

Application to Biotechnology
Extensive research has been attributed to the sequencing of Lactococcus lactis' genome due to its major role in the production of dairy products and preservation. Manufactures use the discovered properties of L.lactis to increase food preservation, distinguish flavor and aroma. L.lactis contains a bacteriocin, a natural antimocrobial agent that fights against a wide range of Gram-positive bacteria, such as food-bourne pathogens. Uses of nisin to control spoilage lactic acid bacteria have been identified in beer, wine, alcohol production, and high acid foods such as salad dressings. Production of highly purified nisin preparations has led to interest in the use of nisin for human ulcer therapy and mastitis control in cattle. A recently discovered application of Lactococcus lactis is in the development of vaccine delivery systems. L.lactis can be genetically engineered to generate proteins from pathogenic species on their cell surfaces. Mucoasal administration of the modified strain will induce an immune response to the cloned protein and provide immunity to the pathogen. Mucosal immunity is a main concern of the public health since it is the primary way of pathogenic entry. In underdeveloped countries, where diseases spread rapidly, mucosal immunity can facilitate the distribution of vaccines since it less cost effective and easily administered. This approach theoretically can be applied to any pathogen that enters a human or mammal through a mucoasal surface, however, it is most commonly used to provide immunity to Streptococcus pyogenes, the pathogenic agent of strep throat.

Economical Importance
L.Lactis holds great value to the fermented dairy product industry. The bacteria is found in milk and its main function is to produce lactic acid. The bacterium can be a single strain starter culture or a mixed strain culture with other lactic acid bacteria. It is a key starter culture in the production of cheeses such as cheddar, colby, cottage cheese, cream cheese, etc. as well as other dairy products like cultured butter, buttermilk, and sour cream. The cheese producing industry has contributed a great deal to the economy. The cheese industry is of such great importance in Wisconsin that it has nominated Lactococcus lactis as it's state microbe. Wisconsin is the leading cheese producing state of the US,in one year they produce 2.5 billion lbs of cheese and 90% p of their milk is used to producing cheese. Wisconsin's cheese producing revenue totals up to $18 billion.

Current Research
Immunogenicity and protective efficacy of orally administered recombinant Lactococcus lactis expressing surface-bound HIV Env The use of L.lactis in successful vaccine delivery has encouraged many researchers to further investigate the various pathogens this approach can be appllied to. This study applies this theory to the human immunodeficiency virus (HIV). Researchers created a recombinant vector using L.lactis, the newly engineered L.lactis expressed the Env gene of HIV, specifically, the V2-V4 loop of HIV Env. Surface expression of Env was achieved by fusing the secreting signal and cell wall anchor to the N- and C-terminal ends of the V2-V4 loop. Mice were then fed the vector containing HIV, along with a control group who were fed a vector with a HIV Env–expressing vaccinia virus. Results showed that the HIV vaccinated mice obtained a viral load 350 fold less than the control group. This outcome has encouraged the further development of a L. lactis based HIV vaccine.

Innate inflammatory responses to the Gram-positive bacterium Lactococcus lactis The impressive use of L.lactis as a useful immunity tool has promoted further research to support the potential applications of the bacteria. In this study L.lactis' adjuvant characteristics are comparatively investigated with other bacteria. The study contained comparative data on the proinflammatory effects of L. lactis strain NZ9000, a non-pathogenic bacterium, with E. coli strain DH5α and Salmonella typhi strain Ty21a, both non-pathogenic strains of pathogenic bacteria. Results showed that when L.lactis, E.coli, and S.typhi were co-incubated with B10R murine macrophages, in vitro, they all expressed pro-inflammatory properties. However, L.lactis expressed lower levels of chemokine mRNA expression. Leukocyte recruitment was also compared, in vivo, between all three bacteriums. L. lactis, E. coli and S. typhi showed similar levels of leukocyte recruitment into murine air-pouches, these recruited cells displayed a specific activation status according to the bacterial stimuli. The results demonstate L. lactis' ability to induce chemokine expression both in vitro and in vivo, and displays similar potency as S. typhi, an established live vaccine.

A new plasmid vector for DNA delivery using lactococci Continuous studies are done trying to improve the use of L.lactis as a DNA carrier for such uses as mucosal vaccines. In this study researchers strive to develop a new type of plasmid that can be used with L.lactis in antigen transfer. The new plasmid,pValac, was created by fusing a prokaryotic and eukaryotic region together. The gfp ORF was cloned into the pValac and was analysed by transfection in PK15 cells. Pvalac:gfp's potential was tested by combining it with Lactococcus lactis inlA+ strains and attempting to insert it into Caco-2 cells. Results showed that after transfection with pValac:gfp, GFP expression was observed in PK15 cells. L. lactis inlA+ were able to invade Caco-2 cells and delivered a functional expression cassette (pCMV:gfp) into epithelial cells.