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Propionibacterium acnes (P. acnes) is a common component of the bacterial flora of the

human skin. It is a predominant bacterium located on the surface and within the deeper recesses of pilosebaceous follicles at the sebum-rich sites of human skin (Gribbon et al.,

1993; Gribbon et al., 1994). P. acnes prefers moist, oily areas of the skin. The

concentration of P. acnes is similar between patients with acne and controls (Dessinioti et al., 2009).

2.2.1 Taxonomic classification

Higher order taxa of the P. acnes is Actinobacteria, Actinobacteridae; Actinomycetales,

Propionibacteriaceae and Propionibacterium. It is known by an alternative

nomenclature such as Corynebacterium acnes, Bacillus acnes (Henyl and Forlifer, 1994)

2.2.2 Genomic structure

The genome of P. acnes has been sequenced in its entirety. This bacterium consists of a

single circular chromosome of 2.56026 base pairs (Bruggemann et al., 2004). Its DNA

contains 2351 putative genes coding for 2297 known protein products and constituting a 60 % G-C (guanine-cytosine) content.

2.2.3 Cellar and colonial morphology

The observation of colonial and bacterial cell morphology of P. acnes were reported

similarly (Poppert et al., 2009; Perry and Lambert, 2006; Esteban et al., 1996). Colonial

morphology of P. acnes is described as opaque, circular, convex colonies of 1 mm or

less. Upon longer incubation, the colonies could change from initial small white colonies to larger and darker yellow or pink colour colonies (Anderson et al., 1999). The surface

of the colonies varied from 1.5 to 4.0 mm in diameter, and the colour of colonies was pink after 4 - 5 days of anaerobic incubation (Douglase and Gunter, 1946).

(a) (b)

Figure 2.4 Negative stains of four days old slant cultures incubated (a) anaerobically and (b) aerobically (Douglas and Gunter, 1946)

P. acnes is non-spore-forming, non-motile, anaerobic, gram-positive and pleomorphic

rod. The bacterium has homogenous cell morphology. The organism ranges from small plump rods to ellipsoids which tend to occur in pairs (Figure 2.4 a).

The size of the individual cells in Gram stains is approximately 0.4 - 0.5 by 0.8 - 0.9 microns (Douglas and Gunter, 1946). P. acnes grows better under anaerobic condition

(Collins et al., 2004) and it can also grow poorly under aerobic condition (Charles et al.,

1979). Cells from aerobic broth culture appear to be longer and swollen, and may appear to have rudimentary branching (Figure 2.5 b). Aerobic cultured cells of P. acnes are

more pleomorphic than those under anaerobic condition (Douglas and Gunter, 1946).

2.2.4 Biofilm formation

P. acnes cells residing within the skin surface follicles are capable growing as biofilm

(Figure 2.5). Biofilm formation of P. acnes is probably responsible for strengthening

the resistance to common antimicrobial agents (Coenye et al., 2007)

Figure 2.5 Visualization of early stages (24 h) of biofilm formation of P. acnes LMG

16711, using SYTO9 and epifluorescence microscopy (magnification: 500×) (Coenye

2.2.4 Biochemical characteristic of P. acnes

P. acnes normally produce acids. The spot test for indole production for P. acnes was

positive (Collins et al., 2004). In the study of Charles et al. (1979), catalase tests, Gram’s

stain, gelatine hydrolysis and indole formation were carried out routinely for checking the purity of the culture.

2.2.5 Metabolism of P. acnes

P. acnes is usually regarded as a sticky anaerobe; it can tolerate oxygen up to 100 %

saturation but grows at reduced rate (Perry and Lambert, 2006). Hence, the effects of oxygen on the in vitro propagation of P. acnes were investigated under defined micro

aerophilic conditions (Gribbon et al., 1994). The metabolic pathways are different

under both aerobic and anaerobic conditions. There is more than one mechanism involved in stimulating the aerobic growth of P. acnes shown in Figure 2.7 (Evans et al., 1979; Bruggemann et al., 2004).

When P. acnes grows under anaerobic condition, propionate, acetate and carbon

dioxide are produced at a ratio of 2:1:1 from lactate or glucose through the Krebs Cycle, Embden-Meyerh of Pathway and the pentose phosphate pathway (Ye et al.,

1999), deriving energy with the help of enzymes such as nitrate reductase, dimethyl sulfoxide reductase and fumarate reductase (Bruggemann et al., 2004).

Figure 2.6 Metabolic pathways of Propionibacterium acnes growing in oxygen (Ye et al., 1999)

The P. acnes is capable of growing in 100 % air at reduced rate; the biomass of the

bacterium is decreased between 0 to 20 % air-saturation and no further changes at higher air saturations for P. acnes (Cove et al., 1983). The growth of P. acnes seems to

be promoted by the utilization of molecular oxygen at low concentration, but at higher concentration, the bacterium induces production of toxic forms of oxygen at concentrations exceeding the capabilities of the protective mechanism by the cells (Gibbon et al., 1994). It appears that P. acnes carries out an aerobic respiration through

formation of succinate by transmitting electrons from cytochromes b and c and vitamin

K2 to fumarate when oxygen serves as a final electron acceptor (Gibbon et al. 1994). It seems that oxygen could activate the reversed randomised pathway, which causes the change of organic acid formation and oxidation of propionate. These pathways are shown in Figure 2.6 (Ye et al., 1999).

2.2.7 Incubation temperature of P. acnes

The optimum growth temperature for P. acnes is 33 °C at pH 5.6 in anaerobic condition

using continuous culture technique (Gribbon et al., 1993). However, the optimum

growth of the bacterium can be also obtained from 35-37 °C (Schlecht et al., 1997).

2.2.8 Media for cultivation the P. acnes

Under anaerobic conditions, P. acnes can grow well on media containing glucose,

glycerol, ribose, fructose, mannose and N-acetylglucosamine. Different solid media for the cultivation of anaerobes were compared in the study of Heginbothem et al. (1990).

In this study, P. acnes had detectable (moderate to good) growth after 48 h on BHI agar

(Brain Heart Infusion agar), FA agar (Fastidious Anaerobe) Agar and GAA (Gibco Anaerobe Agar). In addition, BHI broth is the commonly used liquid medium for growing P. acnes culture (Miskin et al., 1997; Chomnawang et al., 2005).