Lactobacillus algidus

Lactobacillus algidus is a Gram-positive, anaerobic, non-motile, non-spore-forming, rod shaped bacteria found on vacuum packaged refrigerated beef. [1] Furthermore, next generation amplicon sequencing studies have found that L. algidus is also very abundant in fresh meats as well. [2] L. algidus was first isolated by Kato et al. on five specimens of beef that were stored at 2°C for at least 3 weeks, collected from different meat shops. [1] The bacterium was named such that: al´gi.dus. L. adj. algidus cold, referring to the ability to grow at low temperature. L. algidus, relative to other meat-associated lactic acid bacteria (LAB), is rarely recovered from samples and research on the species have been sparse due to its characteristically poor growth on media that is used to culture food spoilage bacteria. [3]

Kato et. al. first characterized the bacteria (named strain M6A9) with SDS-PAGE whole-cell protein patterns and 16S rDNA sequencing that showed distinctive patterns that differentiated L.algidus from several other psychrophilic lactic acid bacteria in the genus Lactobacillus, Carnobacterium and Brochothrix, with the phylogenetically closest strain being Lactobacillus mali (93% sequence similarity) of the Lactobacillus casei/Pediococcus group. [1] L. algidus is unique and differentiated from Carnobacterium species such as C. divergens that have also been isolated frequently from vacuum-packaged refrigerated meat. Carnobacterium species are able to grow at 30 °C, have different carbohydrate fermentation patterns, showcase nitrate reduction and production of arginine decarboxylase, obtain ammonia from arginine, acetoin and hydrogen sulfide. Furthermore, the bacteria are distinctly differentiated from Brochothrix species, one of the classical meat organisms, with different carbohydrate fermentation patterns and lack of catalase production. Most importantly, the bacteria could be distinguished from mDAP-positive or psychrophilic Lactobacillus species in growth characteristic (able to grow in much colder temperatures), "motility, catalase production, gas production from glucose and gluconate, types of lactic acid isomer [products], carbohydrate fermentation patterns, and/or G¬C content of DNA." [1]

L. algidus is considered a potential spoilage bacterium for refrigerated, packaged meats, as meats associated with the bacterium developed sour and intense odors. However, much of its characteristics and role in the spoilage process are poorly understood, such as its ability to produce biogenic amines known to negatively affect safety of meat. [3]

Scientific Classification [1][4]

Domain: Bacteria

Phylum: Firmicutes

Class: Bacilli

Order: Lactobacillales

Family: Lactobacillaceae

Genus: Lactobacillus

Species: L. algidus

Genome & Genomic Comparisons

The G¬C contents of genomic DNA extracted from L. algidus strains M6A9T, M6A19 and M6A40 were 36±5, 37±1 and 36±4 mol%, respectively. [1] Phylogenetic analysis showed that the strain M6A9 was most closely related to the members of the L. casei/Pediococcus group of Lactobacillus, specifically L. mali with the sequence similarity of 93±0%. [1] The genome of L. algidus strain CMTALT10, characterized in 2018 using illumina MiSeq, had a size of 1,624,384 bps with 1,582 coding regions. [5]

Morphology

The L. algidus cells showed uniform characteristics: they were “facultatively anaerobic, psychrophilic, Gram-positive, non-spore-forming, nonmotile, lactic acid-homofermentative rods”. [1] The L. algidus cells/colonies appear singly and in pairs on agar media and in very long chains in broth media. [1] Colonies on MRS agar are circular, convex, smooth, cream–white and 1 to 1.5 mm in diameter after 48 h incubation at 20 °C. The cellular morphology seemed to change slightly based on medium: from being short, almost cocci-like rods in MRS broth to definite rods on MRS agar. [1]

Strain M6A9T, a representative of the strains studied, was found to possess oleic acid as a major cellular fatty acid. Presence of oleic acid was previously used to differentiate Carnobacterium (known to possess oleic acids) from Lactobacillus. However, few Lactobacillus has shown to possess the acids in large amounts as well. [1]

This atypical species was also meso-diaminopimelic acid (mDAP) positive, with mDAPs commonly found on peptidoglycan walls of Gram-negative bacteria and less often found on Gram-negative bacteria. [1][6]

Culture Growth

L. algidus strains are often cultured using a protocol for psychrotrophic LAB that includes a nutrient-rich growth medium, most commonly the de Man Rogosa Sharpe (MRS) medium, with an anaerobic incubation condition with temperature of 25 °C or below. [3] L. algidus was first cultured in BL agar that was enhanced by the addition of 5% (v/v) horse blood. The optimum temperature for growth was 15–25 °C and no growth was observed at 30 °C. [1] This inability to grow at 30°C has been a major obstacle in research. [3] Growth was observed even at temperatures as low as 2°C capable of surviving in refrigerated meat. Anaerobic incubation in BL and MRS agars yielded larger colonies, indicating the anaerobic nature of the bacteria. Additionally, strains did not resist heating at 70°C for 10min. [1]

Recently, researchers have raised concerns that the use of MRS medium fails to accurately simulate L. algidus growth conditions in meat. Observations have shown that L. algidus strains frequently lose their viability after one or two subcultures in MRS media. It is unclear whether their poor growth is due to oxidative stress, or the insufficient growth factors of the MRS medium that are needed for strains’ growth. [3] However, studies have found that MRS agar supplemented with complex substances such as peptone, meat and yeast extract, or liver digest succeeded in supporting the growth of L. algidus, and even allowed the recovery of new bacterial colonies isolates from meat. [3]

Biochemical Properties

Kato et. al. showed that L. algidus produced no gas in Briggs liver broth that is based on glucose or gluconate, thus indicating a homofermentative mechanism with lactic acid as the sole product of glucose fermentation. Catalase, oxidase, nitrate, or hydrogen sulfide are not produced. [1] Supplementation of MRS broth with catalase allowed the bacteria to grow in aerated cultures indicating that the strains produce hydrogen peroxide when grown aerobically. [3]

Homofermentative LAB like L. algidus ferment glucose to produce energy and releases lactic acid as the primary by-product.[7] Since L. algidus is facultatively anaerobic, even in aerobic conditions, the bacteria will ferment and undergo cellular respiration at the same time. [1] During fermentation, the bacteria will convert glucose to two molecules of lactate and simultaneously use this reaction to perform substrate-level phosphorylation to make two molecules of ATP [7]:

    glucose + 2 ADP + 2 Pi → 2 lactate + 2 ATP 

References

[1] Kato, Y., Sakala, R. M., Hayashidani, H., Kiuchi, A., Kaneuchi, C., and Ogawa, M. (2000) Lactobacillus algidus sp. nov., a psychrophilic lactic acid bacterium isolated from vacuum-packaged refrigerated beef. Int. J. Syst. Evol. Microbiol. 50, 1143–1149.

[2] Delhalle, L., Korsak, N., Taminiau, B., Nezer, C., Burteau, S., Delcenserie, V., Poullet, J. B., and Daube, G. (2016) Exploring the Bacterial Diversity of Belgian Steak Tartare Using Metagenetics and Quantitative Real-Time PCR Analysis. J. Food Prot. 79, 220–229.

[3] Säde, E., Johansson, P., Heinonen, T., Hultman, J., and Björkroth, J. (2020) Growth and metabolic characteristics of fastidious meat-derived Lactobacillus algidus strains. Int. J. Food Microbiol. 313, 108379.

[4] Hammes, W. P., and Hertel, C. (2009) Genus I. Lactobacillus. Bergeys Man. Syst. Bacteriol. 3, 465–511.

[5] Poirier, S., Coeuret, G., Champomier-Vergès, M.-C., and Chaillou, S. (2018) Draft Genome Sequences of Nine Strains of Brochothrix thermosphacta, Carnobacterium divergens, Lactobacillus algidus, Lactobacillus fuchuensis, Lactococcus piscium, Leuconostoc gelidum subsp. gasicomitatum, Pseudomonas lundensis, and Weissella viridescens, a Collection of Psychrotrophic Species Involved in Meat and Seafood Spoilage. Genome Announc. 6.

[6] Uehara, A., Fujimoto, Y., Kawasaki, A., Kusumoto, S., Fukase, K., and Takada, H. (2006) Meso-diaminopimelic acid and meso-lanthionine, amino acids specific to bacterial peptidoglycans, activate human epithelial cells through NOD1. J. Immunol. Baltim. Md 1950 177, 1796–1804.

[7] Lactic Acid Bacteria. http://textbookofbacteriology.net/lactics_2.html