Introduction to Anaerobes

ANAEROBES AS PATHOGENS

Anaerobic bacteria differ in their pathogenicity. Not all of them are believed to be clinically significant, while others are known to be highly pathogenic. Table 1 lists the major anaerobes that are most frequently encountered clinically. The taxonomy of anaerobic bacteria has changed in recent years because of their improved characterization using genetic studies (1). The ability to differentiate between similar strains enables better characterization of type of infection and predicted antimicrobial susceptibility. The species of anaerobes most frequently isolated from clinical infections are in decreasing frequency: the clinically important anaerobes are of gram-negative rods (Bacteroides, Prevotella, Porphyromonas, Fusobacterium, Bilophila and Sutterella), gram-positive cocci (primarily Peptostreptococcus), gram-positive spore-forming (Clostridium) and non-spore-forming bacilli (Actinomyces, Propionibacterium, Eubacterium, Lactobacillus, and Bifidobacterium), and gram-negative cocci (mainly Veillonella) (2). About 95% of the anaerobes isolated from clinical infections are members of these genera. The remaining isolates belong to species not yet described, but these usually can be assigned to the appropriate genus on the basis of morphologic characteristics and fermentation products. The frequency of recovery of the different anaerobic strains differs in various infectious sites. The 12 years experience in recovering anaerobic bacteria from adults and children at two medical centers is presented in Table 2 (3). The main isolates were anaerobic gram-negative bacilli (Bacteroids, Prevotella, and Porphyromonas; 43% of anaerobic isolates), anaerobic gram-positive cocci (26%), Clostridium spp. (7%), and Fusobacterium spp. (5%). This chapter discusses the main anaerobic species and their role in infectious processes.

CLASSIFICATION OF ANAEROBES

Anaerobes do not multiply in oxygen but have different susceptibility to oxygen. Most normal flora anaerobes are extremely oxygen sensitive, while those that cause infections are more aero-tolerant. The aero-tolerance of several anaerobes is through the production of superoxide dismutase, they produce on exposure to oxygen. The negative oxidation-reduction potential (Eh) of the environment is a critical factor in the survival of anaerobic bacteria.

Anaerobes do not grow on solid media in room air (10% CO2, 18% 02); facultative anaerobes grow both in the presence and absence of air, and microaerophilic bacteria grow poorly or not at all aerobically but grow better under 10% CO2 or anaerobically. Anaerobes are divided into "strict anaerobes" that are unable to grow in the presence of more than 0.5% 02 or "moderate anaerobes" that are capable of growing at between 2% and 8% 02.

GRAM-POSITIVE SPORE-FORMING BACILLI

Anaerobic spore-forming bacilli belong to the genus Clostridium. Morphologically, the clostridia are highly pleomorphic, ranging from short, thick bacilli to long filamentous forms, and are either ramrod straight or slightly curved. The clostridia found most frequently in clinical infections are Clostridium perfringens (Fig. 1), Clostridium septicum, Clostridium butyricum, Clostridium sordellii, Clostridium ramosum, and Clostridium innocuum.

C. perfringens is an inhabitant of soil and of intestinal contents of humans and animals and is the most frequently encountered histotoxic clostridial species (4). This microorganism, which

TABLE 1 Anaerobic Bacteria Most Frequently Encountered in Clinical Specimens Organism

Infectious site

Gram-positive cocci Peptostreptococcus spp. Microaerophilic streptococci3 Gram-positive bacilli Non-spore-forming Actinomyces spp.

Propionibacterium acnes Bifidobacterium spp. Spore-forming Clostridium spp. C. perfringens C. septicum C. sordellii C. difficile C. botulinum C. tetani Gram-negative bacilli Bacteroides fragilis group (B. fragilis,

B. thetaiotamicron) Pigmental Prevotella and Porphyromonas spp. Prevotella oralis Prevotella B. oris-buccae P. bivia, P. disiens Fusobacterium spp. F. nucleatum F. necrophorum

Respiratory tract, intra-abdominal and subcutaneous infections Sinusitis, brain abscesses

Intracranial abscesses, chronic mastoiditis, aspiration pneumonia, head and neck infections Shunt infections (cardiac, intracranial) Chronic otitis media, cervical lymphadenitis

Wounds and abscesses, sepsis Sepsis

Necrotizing infections Diarrheal disease, colitis Botulism Tetanus

Intra-abdominal and female genital tract infections, sepsis, neonatal infection Orofacial infections, aspiration pneumonia, periodontitis Orofacial infections

Orofacial infections, intra-abdominal infections Female genital tract infections

Orofacial and respiratory tract infections, brain abscesses, bacteremia Aspiration pneumonia, bacteremia

1 Not obligate anaerobes.

elaborates a number of necrotizing extracellular toxins, is easily isolated and identified in the clinical laboratory. C. perfringens seldom produces spores in vivo. It can be characterized in direct smears of a purulent exudate by the presence of stout gram-variable rods of varying length, frequently surrounded by a capsule. C. perfringens can cause a devastating illness with high mortality. Clostridial bacteremia is associated with extensive tissue necrosis, hemolytic anemia, and renal failure. The incidence of clostridial endometritis, a common event following septic abortions, has decreased as medically supervised abortions have increased (2).

  1. perfringens accounted for 48% of all clostridial isolates in our hospitals (Table 2) and was primarily isolated from wounds (26% of C. perfringens) isolates, blood (16%), abdomen (14%), and obstetrical and gynecological infections (13%).
  2. septicum, long known as an animal pathogen, has been found in humans within the last decade, often associated with malignancy. The intestinal tract is thought to be the source of the organism, and most of the isolates are recovered from the blood.
  3. sordellii causes life threatening infections after trauma, childbirth, gynecological procedures, medically induced abortions, surgery and injection of elicit drugs. It can cause rapid progressive tissue necrosis, shock, multiorgan failure and death in about 3/4 of patients (4a).

Although Clostridium botulinum usually is associated with food poisoning, wound infections caused by this organism are being recognized with increasing frequency. Proteolytic strains of types A and B have been reported from wound infections. Disease caused by C. botulinum usually is an intoxication produced by ingestion of contaminated food (uncooked meat, poorly processed fish, improperly canned vegetables), containing a highly potent neurotoxin. Such food may not necessarily seem spoiled, nor may gas production be evident. The polypeptide neurotoxin is relatively heat labile, and food containing this toxin may be rendered innocuous by exposure to 100°C for 10 minutes.

TABLE 2 Percentage of Recovery of Anaerobes in Each Infection Site at Walter Reed Army Medical and Naval Medical Centers 1973-1985

Total Number

Total number of anaerobic

Specimen number of anaerobic Isolates/ Bacteroides Fusobac- Clostridium Lactobacillus Eubacterium Propionibac- Bilidobac- Actinomyces Veillonella Peptostrepto-

source specimens Isolates specimen spp. /erfi/mspp. spp. spp. spp. /em/mspp. /em/mspp. spp. spp. coccj/sspp.

Abdomen

359

550

1.53

299 (55)a

43 (8)

71 (13)

4(1)

31 (6)

23 (4)

8(1)

71 (13)

Abscess

820

1416

1.73

725 (51)

97 (7)

71 (5)

7 (0.5)

44 (3)

54 (4)

5 (0.5)

2 (0.2)

28 (2)

383 (27)

Bile

66

75

1.14

29 (39)

1 (1)

27 (36)

9(12)

9(12)

Bites

9

12

1.33

5(42)

1 (8)

2(17)

4 (33)

Blood

587

634

1.08

222 (35)

24 (4)

70 (11)

1 (0.2)

13(2)

229 (36)

1 (0.2)

7(1)

67 (11)

Bone

37

69

1.86

24 (35)

4(6)

2(3)

1 (1)

9(13)

2(3)

27 (39)

Central nervous

220

225

1.02

16(7)

2(1)

4(2)

163 (72)

1 (0.5)

39 (17)

system

Chest

191

283

1.48

101 (37)

31 (11)

18(6)

1 (0.4)

9(3)

51 (18)

4(1)

9(3)

59 (21)

Cysts

206

348

1.69

153 (44)

5(1)

6(2)

4(1)

6(2)

24 (7)

1 (0.3)

10(3)

139 (40)

Ear

25

47

1.88

12 (26)

1 (2)

1 (2)

7(15)

1 (2)

25 (53)

Eye

55

66

1.20

8(12)

3(5)

11 (17)

36 (55)

4(6)

4(6)

Genitourinary

30

52

1.73

29 (56)

2(4)

1 (2)

1 (2)

3(6)

3(6)

2(4)

11 (21)

Grafts

13

15

1.15

4(27)

1 (7)

5 (33)

5 (33)

Joints

63

69

1.10

9(13)

8(12)

39 (57)

13 (19)

Lymph glands

70

76

1.09

11 (15)

3(4)

1 (1)

48 (63)

1 (1)

2(3)

10 (13)

Obstetric/

871

1328

1.52

654 (49)

42 (3)

50 (4)

15(1)

28 (2)

28 (2)

12(1)

1 (1)

28 (2)

470 (35)

gynecologic

Sinuses

102

159

1.56

53 (33)

11 (!)

2(1)

2(1)

36 (23)

1 (1)

7(4)

47 (30)

Tumors

61

79

1.30

33 (42)

1 (1)

1 (1)

1 (1)

22 (28)

1 (1)

1 (1)

19 (24)

Wounds

622

987

1.59

425 (43)

20 (2)

124 (13)

6(1)

18(2)

66 (7)

1 (0.1)

14(1)

313 (31)

Miscellaneous

51

67

1.31

23 (34)

3(4)

3(4)

2(3)

20 (30)

1 (1)

2(3)

13 (19)

Total

4458

6557

1.47

2835 (43)

294 (5)

471 (7)

40 (1)

158 (2)

874 (13)

27 (0.4)

5(0.1)

125 (2)

1728 (26)

a In parentheses: percentage of all anaerobic bacteria isolated from source indicated. Source: From Ref. 3.

FIGURE 1 Gram stain of Clostridium per-fringens.

  1. botulinum is usually associated with food poisoning (2); botulism is an intoxication caused by ingestion of contaminated food containing its highly potent neurotoxin. However, wound infections caused by proteolytic strains of types A and B has been reported with increasing frequency and can also produce botulism.
  2. botulinum has also been associated with newborns presenting with hypotonia, respiratory arrest, areflexia, ptosis, and poorly responding pupils. Botulism in infants is caused by toxin from the germination of ingested spores and C. botulinum in the bowel lumen. C. butyricum can also be recovered from infection of the abdomen, abscesses, bile, wounds, and blood.

Clostridium difficile has been incriminated as the causative agent of antibiotic-associated and spontaneous diarrhea and colitis (5). A formerly infrequently isolated strain of C. difficile known as BI/NAP1 has recently been implicated in geographically diverse outbreaks of C. difficile-associated disease which have severe clinical presentations and poor outcomes (5).

Clostridium tetani is rarely isolated from human feces. Infections caused by this bacillus are a result of contamination of wounds with soil containing C. tetani spores. The spores will germinate in devitalized tissue and produce the neurotoxin that is responsible for the clinical findings of tetanus. C. tetani has been recovered from patients presenting with otogenous tetanus (6).

Clostridia can be isolated from various infectious sites. These organisms are especially prevalent in abscesses (mostly abdominal, rectal area, and oropharyngeal), and peritonitis (1). The distribution of clostridia in these infections is explained by their prevalence in the normal gastrointestinal and cervical flora from where they may originate (7).

Clostridia strains (C. perfringens, C. butyricum, and C. difficile) have been recovered from blood and peritoneal cultures of necrotizing enterocolitis and from infants with sudden death syndrome (8-10). Strains of Clostridium were recovered from children with bacteremia of gastrointestinal origin (11) and with sickle cell disease (12). Clostridial strains have been recovered from specimens obtained from patients with acute (13) and chronic (14) otitis media, chronic sinusitis and mastoiditis (15,16), peritonsillar abscesses (17), peritonitis (18,19), liver and spleen abscesses (20), abdominal abscesses (21), and neonatal conjunctivitis (22,23).

GRAM-POSITIVE NON-SPORE-FORMING BACILLI

Anaerobic, gram-positive, non-spore-forming rods comprise part of the microflora of the gingival crevices, the gastrointestinal tract, the vagina, and the skin. Since many of them appear to be morphologically similar, they have been difficult to separate by the usual bacteriologic tests. Several distinct genera are recognized: Actinomyces, Arachnid, Bifidobacterium, Eubacterium, Lactobacillus, and Propionibacterium.

The Actinomyces, Arachnia, and Bifidobacterium of the family Actinomycetaceae are grampositive, pleomorphic, anaerobic to microaerophilic bacilli. Species of the genus Bifidobacterium are part of the commensal flora of the mouth gastrointestinal tract and female genital tract and constitute a high proportion of the normal intestinal flora in humans, especially in breast-fed infants (24). Although some infections caused by these organisms have been reported (25-28), little is known about their pathogenic potential.

Eubacterium spp. are part of the flora of the mouth and the bowel. They have been recognized as pathogens in chronic periodontal disease (29) and in infections associated with intra-uterine devices (30), and have been isolated from patients with bacteraemia associated with malignancy (31) and from female genital tract infection (32). Lactobacillus spp. are ubiquitous inhabitants of the human oral cavity, the vagina, and the gastrointestinal tract (33). They have been implicated in various serious deep-seated infections, amnionitis (33) and bacteraemia (34). Eubacterium, Lactobacillus, and Bifidobacterium spp. have been isolated in pure culture in only a few instances and are usually isolated in mixed culture from clinical specimens (1). The infections where they have been found most often are chronic otitis media and sinusitis, aspiration pneumonia, and intra-abdominal, obstetric and gynecological and skin, and soft-tissue infections (1,35,36).

Actinomyces israelii and Actinomyces naeslundii are normal inhabitants of the human mouth and throat (particularly gingival crypts, dental calculus, and tonsillar crypts) and are the most frequently isolated pathogenic actinomycetes. These organisms have been recovered from intracranial abscesses (37), chronic mastoiditis (16), aspiration pneumonia (38), and peritonitis (18). Although actinomycetes often are present in mixed culture, they are clearly pathogenic in their own right and may produce widespread devastating disease anywhere in the body (39). The lesions of actinomycosis occur most commonly in the tissues of the face and neck, lungs, pleura, and ileocecal regions. Bone, pericardial, and anorectal lesions are less common, but virtually any tissue may be invaded; a disseminated, bacteremic form has been described.

Propionibacterium spp. are part of the normal bacterial flora that colonize the skin (40), conjunctiva (41), oropharynx, and gastrointestinal tract (42). These non-spore-forming, anaerobic, gram-positive bacilli are frequent contaminants of specimens of blood and other sterile body fluids and have been generally considered to play little or no pathogenic role in humans.

Propionibacterium acnes and other Propionibacterium spp. have, however, been recovered with or without other aerobic or anaerobic organisms as etiologic agents of multiple infection sites (43-54). These include conjunctivitis (43), intracranial abscesses (44,45), peritonitis (46), and dental, parotid (47,48), pulmonary (47,48), and other serious infections (49). They have often been recovered as a sole isolate in specimens obtained from patients with infections associated with a foreign body (such as an artificial valve), endocarditis (50,51), and central nervous system shunt infections (50,52). The possible role of P. acnes in the pathogenesis of acne vulgaris was suggested. The data that support this are based on the recovery of this organism in large numbers from sebaceous follicles, especially in patients with acne, on its ability to elaborate enzymes such as lipase, protease, and hyaluronidase, and on its ability to activate the complement system and enhance chemotactic activity of neutrophils (53).

GRAM-NEGATIVE BACILLI

The anaerobic gram-negative rods are differentiated into genera on the basis of the fermentation acids they produce. The family Bacteroidaceae contains several genera of medical importance: Bacteroides fragilis group, Prevotella, Porphyromonas, Bacteroides, and Fusobacterium.

Acne Myths Uncovered

Acne Myths Uncovered

What is acne? Certainly, most of us know what it is, simply because we have had to experience it at one time or another in our lives. But, in case a definition is needed, here is a short one.

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