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Biochemical Tests for Bacteria
Submitted by: danr November 06, 2022

Biochemical Tests Table for Bacteria

Explanation of fields used in the table

TRADITIONAL BIOCHEMICAL TESTS
Shorthand Longhand Significance
Egg yolk Egg-yolk reaction Test for lecithinase activity (Kushner1957).
Haemolysis Haemolysis of red blood cells A trait possessed by some pathogenic bacteria.
Methyl red Methyl red test Test of the acidity (pH <4.5) of the bacteria's environment after fermentation of glucose (microbiologyinfo.com).
NO3 Reduction Nitrate reduction Test for the ability of a bacterium to use nitrate as an electron acceptor (usually instead of oxygen).
NO3 → NO2 Nitrate reduction to nitrite Specified as reduction of nitrate to nitrite (see above). Does not imply further reduction to nitrogen.
NO2 Reduction Nitrite reduction Bacterium ability to use nitrite as an electron acceptor, usually producing nitrogen gas.
ONPG O-Nitrophenyl-β-D-galactopyranoside Test for β-galactosidase activity.
VP Voges-Proskauer test A test for the production of acetoin (acetylmethyl carbinol) or diacetyl.
SUBSTRATE HYDROLYSIS, DIGESTION OR DEGRADATION
Aesculin Aesculin hydrolysis Aesculin, or 6-O-β-glucoside-7-hydroxycoumarin, is an uncommon natural compound combining glucose and the aglycone, aesculetin, which can combine with Fe3+ to give a dark colouration. The test can also serve to indicate the production of β-glucosidase. About 50% of the most widespread gut bacteria are able to hydrolyse this compound.
Urea Urea hydrolysis Urease catalyses the decomposition of urea to ammonia and CO2. For some bacteria, urea hydrolysis plays a role in their pathogenicity (Mobley1989). Of the common commensals, a number of Firmicutes and Proteobacteria produce urease; Actinobacteria are fewer and Bacteroides quite rare.
Gelatin Gelatin digestion Gelatin digestion tests for production of gelatinases - a protease - by bacteria (delaCruz2012). Liquifaction allows fine charcoal particles to migrate through the medium, indicating a positive result. Of the most dominant gut bacteria, mostly Bacteroides species produce gelatinase, while Firmicutes (e.g. Ruminococcus torques, Clostridioides difficile) species are the next most prevalent.
Starch Starch hydrolysis Tests for the presence of exozymic α-amylase and oligo-1,6-glucosidase, which break down amylose and amylopectin starch components to readily useable glucose. Many gut bacteria, including Prevotella, Bacteroides, Bacillus and Clostridium species can hydrolyse starch.
Casein Casein digestion A casein digestion assay tests for the production of caseinase (a peptidase), which breaks down casein protein to amino acids and peptides. The dominant gut bacteria that utilise casein are from the Firmicutes (e.g. Bacillus badius DSM 23T, Paraclostridium bifermentans and Priestia flexa).
Tyrosine Tyrosine digestion Tyrosine can be degraded by some gut bacteria to produce tyramine via decarboxylation. About a dozen common gut bacteria (all aerobic or facultative anaerobes) are capable of utilising this substrate, including Bacillus badius, Comamonas kerstersii, and Proteus mirabilis.
Arginine Arginine hydrolysis Typically a test for the presence of arginine dihydrolase, which breaks down arginine to ornithine and ammonia by decarboxylative-deamination sequence. Common bacteria producing ADH include Streptococcus parasanguinis, Citrobacter koseri, Lactococcus lactis subsp. lactis and Slackia exigua.
Cellulose Cellulose hydrolysis A small handful of common gut bacteria can produce cellulases that break down cellulose.  These include Cellulosilyticum lentocellum, Bacteroides cellulosilyticus, Ruminococcus albus and Lachnoclostridium phytofermentans.
Dextrin Dextrin hydrolysis Wheat dextrins are comprised of human digestion-resistant polyglucose fragments and are a good measure of a bacterium's ability to ferment this dietary fibre in the gut (Noack2013).
DNA DNA degradation Bacteria can release extracellular DNase enzymes in order to degrade DNA of host defences (Buchanan2006), destroy competing bacterial secretions or consume DNA as a nutrient (Finkel2001). Only minor colonisers from Proteobacteria (e.g. Proteus vulgaris and Stenotrophomonas maltophilia) and Firmicutes (e.g. Eubacterium limosum and Mammaliicoccus sciuri) tend to produce these enzymes.
Meat Meat digestion Traditionally, this assay was used to determine the protealytic capability a bacterium might have. Given that high consumption of red meat can lead to adverse health and gut composition issues (AlbrachtSchulte2021), recent work has found the byproduct, N-glycolylneuraminic acid, to be a potential factor involved in various disease states (NIH2019).
Milk Milk curdle A traditional test for presence of the protease, caseinase. Bacteria that produce curdling are used for manufacturing cheese, yoghurt and other fermented dairy products. These bacteria can often colonise the gut because they are consumed with the food, often on a continual basis. Numerous Bacteroides (e.g. Bacteroides thetaiotaomicron), Bifidobacteria (e.g. Bifidobacterium pseudocatenulatum) and Firmicutes (e.g. Lactobacillus kefiranofaciens and Ruminococcus lactaris) can curdle milk.
Chitin Chitin digestion Chitinase is produced by a number of Bacillus and Serratia species (Rathore2015), some of which colonise the gut on occasion (e.g. Serratia liquefaciens, Bacillus licheniformis and Serratia marcescens). Clostridium tertium and several other related organisms can also digest this abundant structural polymer.
Alginate Alginate digestion A test for alginate lyases, often produced by environmental bacteria associated with the degradation of brown algae. Apart from Bacteroides intestinalis (strain DSM 17393; Mathieu2018), very few gut bacteria of signficance are able to digest this substance, however.
Agar Agar digestion Agar differs from alginate in that the former is produced by red algae, whereas the latter by brown algae. Whereas starch is made up of amylose and amylopectin, agar is a mixture of agarose and agaropectin; the jelly-like component is the linear chained agarose, which itself is made up repeating units of agarobiose - a disaccharide of β-D-galactose and 3,6-anhydro-L-galactopyranose. The enzyme β-agarase breaks down agar.
Pectin Pectin digestion Pectin - a polyuronide - is formed as a copolymer of uronic acids (e.g. glucoronic, mannuronic and galacturonic acids) and monosaccharides. Some important colonic bacteria, such as Bacillus subtilis, Bacteroides galacturonicus, Bacillus licheniformis, Bacteroides caecimuris, and Bacteroides cellulosilyticus, can attack pectin.
Hipp. Hippurate hydrolysis A test for the presence of hippuricase. Hippurate is a byproduct of microbial gut metabolism and is also made in the liver and kidneys. A number of important gut commensals, such as Lacticaseibacillus rhamnosus, Enterococcus faecalis and Finegoldia magna are able to decompose this compound.
Mucin Mucin digestion Mucin degradation is of particular interest because the intestinal tract produces this O-glycoprotein to create a protective barrier between the lumen and the epithelial layer. Both beneficial and pathogenic bacteria take advantage of this layer and use it as a food source, a method of immobilisation or their own protection. Important mucin degraders include Bacteroides fragilis, Bacteroides thetaiotaomicron, Akkermansia muciniphila, Bifidobacterium bifidum and Eggerthella lenta.
Xanthine Xanthine hydrolysis Xanthine is a purine found in many tissues of the body. Few common gut microbes (e.g. Megamonas hypermegale) are reported to digest this substance.
Tweens Tween digestion Tweens are non-ionic surfactants that combine a hydrophilic polyethoxylated sorbitan with a single ester of laurate (Tween 20), palmitate (Tween 40), stearate (Tween 60) or oleate (Tween 80). An organism that produces a positive result can hydrolyse the ester with an ester-lipase, but also can survive the detergent effects of the Tween. Gemella morbillorum, Serratia liquefaciens and Stenotrophomonas maltophilia are examples of minor gut colonisers that can hydrolyse Tween 80.

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