Anaerobic Infections Part 1

General Considerations

Anaerobic bacteria and oxygen tolerance

Obligate anaerobes require reduced oxygen tension (< 10%) for growth; they do not survive as surface growths on solid medium in ambient air (i.e., 20% oxygen). In contrast, facultative bacteria can grow in air with or without 10% carbon dioxide, whereas microaerophilic or capnophilic bacteria grow poorly in ambient air, but they grow better in air with reduced oxygen tension (i.e., 10% oxygen and 10% carbon dioxide). Obligate anaerobes vary greatly in their sensitivity to oxygen. Extremely oxygen-sensitive anaerobes, such as spirochetes and some Clostridium species, cannot tolerate even 0.5% oxygen. As a general rule, clinical isolates commonly recovered from anaerobic infections are relatively aerotolerant, able to survive in 2% to 8% oxygen.

Obligate anaerobes in the normal flora

Quantitatively, obligate anaerobes are the predominant normal flora on mucocutaneous surfaces—especially the oral cavity, skin, and gastrointestinal and genital tracts—where they outnumber facultative bacteria by a factor of 10 to 103 [see Table 1]. The major genera of obligate anaerobes and their distribution in the normal flora vary according to body site [see Table 2]. These indigenous bacteria have unique ecologic niches at different body sites. For example, in the oral cavity, Actinomyces viscosus (along with the aerobes Streptococcus sanguis, S. mutans, and S. mitis) preferentially colonizes the tooth surface; in contrast, Veil-lonella parvula and S. salivarius have a predilection for the tongue and buccal mucosa.1 Bacteroides vulgatus, B. thetaiotaomicron, B. fragilis, and B. distasonis are primarily indigenous in the colon2; Prevotella bivia and P. disiens are primarily resident in the female genital tract.3 In addition, this ecosystem is readily influenced by a variety of physiologic and other host factors, such as age, pregnancy, menses, diet, underlying disease, hospitalization, and antimicrobial therapy [see Table 3].

Pathogenesis of anaerobic infections

Host Conditions

Anaerobic infections characteristically are polymicrobial (mixed) and include both anaerobic and facultative organisms. The organisms tend to be acquired endogenously. The particular mix of pathogens reflects the combined influence of the complex commensal flora at a specific body site and the unique microbio-ta of the underlying conditions. Because these organisms are generally of low pathogenicity, anaerobic or mixed infections generally develop as a consequence of either structural alterations in the normal mucosal barrier or tissue ischemia with lowered oxidation-reduction potential. Knowledge of the anatomic location of the primary source of infection and the underlying condition of the host, therefore, is essential in predicting the probable organisms causing anaerobic and mixed infections associated with the indigenous microflora.

Table 1 The Predominant Normal Flora at Various Body Sites

	Anaerobes		Aerobes
Site	Concentration	Predominant Genera	Concentration	Predominant Genera
Skin	104-105/cm2	Propionibacterium	102-103/cm2	Staphylococcus Micrococcus Diphtheroids
Oropharynx	106-10U/ml	Peptostreptococcus Veillonella Actinomyces Prevotella Porphyromonas Fusobacterium	104-106/ml	Streptococcus
Stomach and upper small bowel Lower small bowel Colon	101-104/ml 104-107/ml 1011-1012/ml	Peptostreptococcus Veillonella Bacteroides Bifidobacterium Bacteroides Bifidobacterium Eubacterium Clostridium Peptostreptococcus Veillonella	10l-104/ml 104-107/ml 108-109/ml	Streptococcus Lactobacillus Coliforms Escherichia Enterococcus Lactobacillus
Female genitalia (vagina and endocervix)	108-10l°/g	Peptostreptococcus Lactobacillus Bacteroides Prevotella	107-109/g	Lactobacillus Streptococcus Staphylococcus

Table 2 Classification of Anaerobic Bacteria and Their Distribution in Normal Flora and in Infection

Anaerobic Bacteria	Genera	Normal Flora				Predominant Species from Anaerobic Infections
		Skin	Oropharynx	Intestine	Genitalia
Sporulating bacilli	Clostridium	0	±	2	±	C. perfringens, C. difficile, C. ramosum, C. septicum, C. novyi
Nonsporulating bacilli Gram negative	Bacteroides Prevotella Porphyromonas Fusobacterium	000 0	2 2 2 2	2 1 1 ±	1 1 1 1	B. fragilis group (B. fragilis, B. vulgatus, B. thetaiotamicron, B. distasonis) P. melaninogenica, P. intermedia, P. bivia, P. disiens P. asaccharolytica, P. gingivalis F. nucleatum, F. necrophorum, F. varium, F. mortiferum
Gram positive	Actinomyces Bifidobacterium Eubacterium Lactobacillus Propionibacterium and Arachnia	0 0 ± 0 2	1 ± 1 ± ±	± 2 2 1 ±	± ± ± 2 1	A. israelii, A. naeslundii, A. viscosus B. eriksonii, B. breve E. lentum, E. limosum L. acidophilus, L. casei, L. plantarum P. acnes, P. granulosum, A. propionica
Cocci Gram negative Gram positive	Veillonella Peptostreptococcus	0 1	2 2	1 2	1 1	V. parvula P. asaccharolyticus, P. prevotii, P. magnus, P. variabilis, P. anaerobius, P. micros, P. productus
Spirochetes	Treponema	0	1	±	±	T. vincentii, T. denticola

0, absent or rare; ±, irregularly present; 1, usually present; 2, predominant

Microbial Synergy

Two thirds of anaerobic infections involve both anaerobes and facultative bacteria. The infectivity of obligate anaerobes in these instances is often facilitated by the coexistence of facultative organisms. Such examples of microbial synergy are particularly well demonstrated in periodontal infection and in various animal models of intra-abdominal and subcutaneous ab-scesses.4,5 For example, microbial synergy is common between Bacteroides species and aerobic bacteria or anaerobic cocci and between most Peptostreptococcus species and Pseudomonas aeruginosa or Staphylococcus aureus. Anaerobes may require symbiotic facultative bacteria for providing necessary growth factors, lowering the oxidation-reduction potential of the environment, or impairing local host defenses. Conversely, the presence of obligate anaerobes may benefit coexisting facultative bacteria by growth enhancement,6 protection from phagocytosis (e.g., succinic acid production by Bacteroides species),7 or protection from p-lactam antibiotics (e.g., p-lactamase production).8 Infective synergy between anaerobes and facultative bacteria is best demonstrated within tissues in which bacterial clearance is normally slow (e.g., subcutaneous abscesses or fibrin clot in intraperitoneal infection) or is hampered by underlying disease. An understanding of the dynamic interactions between different components of a complex flora in mixed infections has important therapeutic implications. Microorganisms in mixed infections may respond to antimicrobial agents differently than do those in monomicrobial infections, and it may not be necessary to eradicate every bacterial species in mixed infection to achieve a cure.

Virulence Factors

A number of microbial virulence factors are considered important in the pathogenesis of anaerobic infections [see Table 4].

Extracellular or membrane-bound enzymes Obligate anaerobes possess a number of extracellular or membrane-bound enzymes that promote tissue destruction. These include lipas-es, proteases, nucleases, and heparinases.9 Membrane-bound enzymes, such as superoxide dismutase10 and p-lactamases,8 may be important for protecting virulent organisms from the toxic effects of oxygen and p-lactam antibiotics, respectively. Catalase may serve a function similar to that of superoxide dis-mutase. Organisms lacking these enzymes are susceptible to killing by toxic oxygen radicals and common antibiotics in the environment.

Table 3 Effect of Host Conditions on the Indigenous Microflora at Various Body Sites

Site	Host Condition	Change in Normal Flora
Gingiva	Dental caries and periodontal disease	Increased motile anaerobic gram-negative bacilli and spirochetes
Oropharynx	Hospitalization, antibiotics, or serious illness	Increased facultative gram-negative bacilli
Upper small bowel	Achlorhydria, vagotomy, pyloroplasty	Increased E. coli, B. fragilis, and Bifidobacterium
Small bowel	Regional enteritis, decreased motility, or stasis secondary to blind loop, obstruction, diverticuli, irradiation, etc. Disrupted anatomic continuity after bowel resection or bypass surgery	Colonic flora
Large bowel	Colonic resection with ileostomy	Decreased anaerobes and some facultative bacteria
Vagina	Parturition, hysterectomy, or irradiation	Increased E. coli and B. fragilis

Table 4 Microbial Virulence Factors Important in Mixed Anaerobic Infections

Microbial Factor	Pathogenic Effect
Histolytic enzymes (e.g., collagenase, fibrinolysin, hemolysins, hyaluronidase, protease, lipase, ribonuclease, deoxyribonuclease; Bacteroides fragilis, Porphyromonas gingi-valis, Prevotella melaninogenica)	Tissue destruction
Hemolysins (Clostridium perfringens)	Hemolysis
Enterotoxins (C. perfringens, C. difficile)	Alteration of cell function with diarrhea or cell death
Neurotoxins (C. tetani, C. botulinum)	Blockade of neuromuscular junctions with either spasticity (tetanus) or paralysis (botulism)
Endotoxin (gram-negative anaerobes; B. fragilis, Fusobacterium nuclea-tum, P. intermedia, Veillonella parvula)	Direct toxicity; Hageman factor, complement activation
Heparinase (P. gingivalis, Bac-teroides species, Fusobacterium species)	Promotion of coagulation and tissue ischemia
Capsular polysaccharide (B. frag- ilis, P. melaninogenica, Pep-tostreptococcus species)	Inhibition of phagocytosis; abscess formation
IgA protease (P. melaninogenica, P. gingivalis, P. intermedia)	Impairment of secretory and muco-sal immunity
Succinic acid (B. fragilis)	Inhibition of phagocytosis
p-Lactamase (B. fragilis, P. melaninogenica, P. gingivalis, F. nucleatum)	Antibiotic resistance
Oxygen-scavenging enzymes (e.g., superoxide dismutase, catalase, peroxidase; P. gingi-valis, B. fragilis, C. perfringens)	Oxygen tolerance
Surface ligands and charge (P. gingivalis, P. intermedia, F. nucleatum, Actinomyces naeslundii)	Adherence and bacterial interaggregation
Bacteriocin and metabolites (e.g., fatty acids, H2S, NH3; Propionibacterium acnes, B. frag-ilis, F. necrophorum)	Inhibition of other normal flora

Capsular polysaccharide B. fragilis, Prevotella melaninogeni-ca, and a number of Peptostreptococcus species are encapsulated and exhibit increased virulence in abscess formation and systemic invasion.11 Interestingly, organisms that are normally nonencapsulated and unable to induce abscesses by themselves during experimental infection may become heavily encapsulated after participating in a mixed infection with other aerobic and anaerobic bacteria.5 These heavily encapsulated strains are able to induce abscesses thereafter when inoculated alone. This phenomenon may help explain how nonpathogen-ic organisms that are part of the normal host flora can become pathogens. The capsular materials of B. fragilis and P. melanino-genica have been extracted and purified. These large-molecular-weight polysaccharides have been demonstrated to inhibit phagocytosis in vitro and to promote abscess formation in several animal models.12 In addition, several oral anaerobes (e.g., P. melaninogenica, P. gingivalis, and P. intermedia) can secrete IgA proteases that may impair secretory and local mucos-al immunity.

Lipopolysaccharide Like their aerobic counterparts, anaerobic gram-negative bacteria possess lipopolysaccharides (LPS) in their outer cell membrane. However, the structure and biologic activity of LPS from several anaerobic bacteria are distinctly different from those of the classic LPS of Enterobacteriaceae. For example, LPS of B. fragilis and P. intermedia lack 2-keto-3-deoxyoc-tanoic acid and L-glycero-D-mannoheptose, and they have little endotoxic potency.14 The LPS of F. nucleatum and V. parvula, on the other hand, have biochemical and biologic properties similar to those of classic endotoxin.

Fatty acids and other metabolites B. fragilis and other anaerobes produce various short-chain fatty acids that are deleterious to mammalian cell function. Infections with B. fragilis are associated with the production of high concentrations of succinic acid, which impairs the generation of the respiratory burst and profoundly reduces phagocytic killing and chemotactic responses of neutrophils.15 This effect is most evident at the low pH and the low Eh (redox potential) conditions present in abscesses and mixed infections. Succinic acid production may represent an important virulence mechanism by Bacteroides species in the patho-genesis of synergistic mixed infections.

Toxins Several Clostridium species produce potent exotoxins. The most important of these is C. perfringens a-toxin, which is a lecithinase that exhibits hemolytic, necrotizing, and lethal properties.16 a-Toxin disrupts membranes containing phospho-lipid-lecithin complexes, including human cell and mitochond-rial membranes, and has direct myocardial depressant properties. A second clostridial exotoxin, p-toxin, is a potent cytotoxin that has cytolytic activity, particularly against endothelial cells. Toxin production at a site of injury allows rapid invasion and destruction of healthy tissues. The paucity of leukocytes in the exudate of clostridial myonecrosis may reflect the presence of these cytotoxins.

C. difficile, the major cause of antibiotic-associated diarrhea and enterocolitis, produces two high-molecular-weight entero-toxins, toxin A (enterotoxin) and toxin B (cytotoxin).17 Both toxins inactivate Rho proteins, a family of small guanosine triphosphate-binding proteins that regulate actin cytoskeleton and various signal transduction processes. Toxin A causes intestinal fluid secretion, mucosal injury, and inflammation. Toxin B has no demonstrable effect on cell permeability and fluid secretion, but like toxin A, it disrupts tight junctions in human epithelial cell monolayers. Toxin B is 10 times more potent on a molar basis than toxin A in mediating damage to human colonic mucosa.

Table 5 Anaerobic Bacteria Associated with Specific Infections^57-86

Site	Infections Likely to Involve Anaerobes	Anaerobes Recovered (Percentage of Infections)	Predominant Isolates
Head and neck	Periapical abscess	90	Anaerobes: Peptostreptococcus species, Prevotella melanogenica, P. intermedia, Porphyromonas gingivalis, P. asaccharolyticus, Actinomyces species, Fusobacterium nucleatum, F. necrophorum
	Periodontal infection	100
	Fascial space infections	100
	Peritonsillar abscess	84	Aerobes: Streptococcus species
	Chronic sinusitis	53
	Chronic otitis media	56
Intracranial	Brain abscess (nontraumatic)	89	Anaerobes: Peptostreptococcus species, Bacteroides fragilis, F. nucleatum, A. israelii
	Subdural empyema	~50
			Aerobes: Streptococcus milleri, other Streptococcus species
Pleuropulmonary	Aspiration pneumonia	87	Anaerobes: Peptostreptococcus species, B. fragilis group, F. nucleatum, pig-mented and nonpigmented Prevotella species, Clostridium species, Eu-bacterium species, Actinomyces species, Lactobacillus species, Veillonella parvula
	Lung abscess	93
	Empyema	76
	Necrotizing pneumonia	94
	Bronchiectasis	27	Aerobes: S. pneumoniae, Haemophilus influenzae, Staphylococcus aureus,
	Hospital-acquired pneumonia	35	Pseudomonas aeruginosa, Enterobacteriaceae species
Intra-abdominal	Intra-abdominal sepsis	83	Anaerobes: B. fragilis, other members of B. fragilis group, Peptostreptococ-cus species, Clostridium species, Fusobacterium species, Biliophila wadsworthia
	Hepatic abscess	25
	Appendiceal abscess	92
			Aerobes: Escherichia coli, P. aeruginosa, Klebsiella species, Enterococcus species, S. aureus
Female genital tract	Vulvovaginal abscess	75	Anaerobes: Prevotella bivia, P. disiens, Bacteroides species, Peptostreptococ-cus asaccharolyticus, P. anaerobius, Actinomyces species
	Salpingitis and pelvic peritonitis	25
	Tubo-ovarian abscess	92
	Posthysterectomy wound infections	67	Aerobes: Gardnerella vaginalis, E. coli, group B streptococci, coagulase-negative staphylococci, Neisseria gonorrhaeae, Chlamydia trachomatis
	Septic abortion and postpartum en-dometritis	73
Skin, soft tissue, and bone Blood	Crepitant cellulitis	75	Anaerobes: Peptostreptococcus species, Bacteroides fragilis, P. melaninogeni- ca, F. nucleatum, C. perfringens Aerobes: Proteus mirabilis, E. coli, Enterococcus species, S. aureus, P. aeruginosa, S. anginosus, S. aureus, Eikenella corrodens B. fragilis group, F. necrophorum, Peptostreptococcus species, C. perfringens, C. septicum
	Synergistic necrotizing cellulitis	89
	Necrotizing fasciitis	47
	Myonecrosis	100
	Infected pressure ulcer	63
	Pilonidal abscess	73
	Perirectal abscess	77
	Diabetic foot ulcer	63
	Breast abscess	79
	Bite wound infections Primary bacteremia	53 4

Tetanus toxin (tetanospasmin) is produced in a tetanus-infected wound and is transported intra-axonally along motor nerves to the spinal cord. Here, the toxin alters normal control of the reflex arc by suppressing the inhibitory neurotransmit-ter y-aminobutyric acid (GABA), producing severe muscle spasms. Like tetanospasmin, botulinum toxin also binds irreversibly to presynaptic nerve endings of cranial and peripheral nerves. Once bound, botulinum toxin prevents the release of the neurotransmitter acetylcholine, producing flaccid paralyses.

Specific Anaerobic Infections

Anaerobic bacteria can cause infections throughout the body. These infections can be conveniently divided into three categories on the basis of unique clinical, microbiologic, and epi-demiologic features. These include infections caused by (1) Bac-teroides and other mixed anaerobes, (2) Actinomyces species, and (3) Clostridium species.