Anaerobes as Part of the Human Indigenous Microbial Flora

The human mucous and epithelial surfaces are colonized with aerobic and anaerobic microorganisms (1). These surfaces are the skin, conjunctiva, mouth, nose, throat, lower intestinal tract, vagina, and the urethra. The trachea, bronchi, esophagus, stomach, and upper urinary tract are not normally colonized by indigenous flora. However, a limited number of transient organisms may by present at these locations. Differences in the environment, such as oxygen tension and pH and variations in bacterial adherence, account for the changing patterns of bacterial colonization. The microflora also varies within the different body sites; in the oral cavity, for example, the organisms in the buccal folds vary in their concentration and types from those from the tongue or gingival sulci. However, the bacteria that prevail in a system generally belong to certain major bacterial species. The relative and total bacterial counts can be influenced by various factors, such as age, diet, anatomic variations, illness, hospitalization, and antimicrobial therapy.

Anaerobes outnumber aerobes in all mucous surfaces, and certain types predominate in the different sites (Tables 1 and 2). Their recovery is inversely related to the oxygen tension. Their predominance in the skin, mouth, nose, and throat which are exposed to oxygen is explained by the anaerobic microenvironment generated by the facultative bacteria that consume oxygen.

Recognizing the unique composition of the flora at certain sites is useful for predicting which organisms may be involved in an adjacent infection and can assist in the selection of empiric antimicrobial therapy. It can also be useful in determining the source and significance of microorganisms recovered from body sites. For example, bacterial endocarditis caused by Enterococcus faecalis is more often associated with urinary tract infection, while alpha hemolytic streptococcal endocarditis is more often observed in patients with poor dental hygiene and tooth extraction.

Knowledge of the indigenous microflora is helpful in determining the consequence of overgrowth of one microorganism by another. Antimicrobials that suppress the intestinal anaerobes may select for the over growth of Clostridium difficile which can result in the production of a potent enterotoxin inducing colitis. Recognition of the normal flora can also help the microbiology laboratory to select proper selective culture media inhibiting certain organisms regarded as contaminants. Furthermore, proper media can enhance the growth of expected pathogens. The recovery of certain organisms from the blood can suggest a possible port of entry (i.e., Clostridium and Bacteroides fragilis usually originate from the gastrointestinal tract) (2).

The normal flora is not exclusively a potential hazard for the host. It can also serve as a beneficial partner. An example of such a benefit is the development of vitamin K deficiency following antimicrobial therapy, which suppresses the gut flora that produces this vitamin.

The normal flora also serves as protector from colonization and subsequent invasion by potential pathogens. Bacterial interference (BI) may play a major role in the maintenance of the normal flora of skin and mucous membranes, by preventing colonization and subsequent invasion by exogenous bacteria (Fig. 1). BI is expressed through several mechanisms. These includes the production of antagonistic substances, changes in the microenvironment and reduction of needed nutritional substances (3). The mediators of BI include the production of

TABLE1

Normal Aerobic and Anaerobic Flora

Aerobes

Anaerobes

Predominant anaerobic organisms

Skin

109-ii

Propionbacterium acnes, Peptostreptococcus spp.

Oral cavity

108-9

Pigmented Prevotella, and Porphyromonas,

Fusobacterium spp.

Upper GIa

102"5

103"7

Bacteroides fragilis group

Lower GIb

105"9

1010"12

Clostridium spp.

Vagina

108

109

Prevotella bivia, Prevotella disiens

Number of organisms per 1 g secretion or contents. a The small intestine and accending colon. b The transverse, descending colon, and rectum.

bacteriocins, bacteriophages, or bacteriolytic enzymes, and molecules such as hydrogen peroxide, lactic or fatty acids and ammonia (3).

THE SKIN

The commonest members of the cutaneous microflora are Staphylococcus, Micrococcus, Coryne-bacterium, Propionibacterium, Brevibacterium, and Acinetobacter and the yeast Pityrosporum (Table 2). The skin flora varies depending on the skin site and its characteristics.

Several potential pathogens are only transient residents around orifices. The oral region or sites that can be in contact with the oropharyngeal flora (i.e., nipples, fingers, genitalia) can become colonized with oral flora organisms (4). These include Haemophilus, Peptostreptococcus, Fusobacterium, and pigmented Prevotella and Porphyromonas spp. Similarly the rectal, vulvovaginal areas, and lower extremities may become colonized with colonic and vaginal organisms. These include B.fragilis group, Clostridium spp., (of rectal origin), or Neisseria gonorrheae, group B Streptococci, and Prevotella (of vaginal origin). These can cause local (i.e., wounds, abrasions, infected burns, decubitus ulcers) or serious infections including bacteremia (2).

The anaerobic microflora of the skin generally is made up of the genus Propionibacterium (5). Propionibacterium acnes predominates, while Propionibacterium granulosum and Propionibacterium

TABLE 2 Predominant Human Microbial Flora at Body Sites

Oral Lower gastro- Genitourinary Type of bacteria Skin Conjunctiva Nasopharynx cavity intestinal tract tract

Aerobic and facultative

Staphylococcus spp. C C +

Streptococcus spp.

C

C

C

C

+a

C

Haempohilus spp.

C

Moraxella catarrhalis

C

Enterobacteriaceae

C

C

Anaerobic

Veillonella sp.

C

C

C

Peptostreptococcus spp.

C

C

C

C

C

Actinomyces spp.

C

C

Bifidobacterium spp.

C

C

C

Eubacterium spp.

C

C

C

Lactobacillus spp.

C

C

Propionibacterium spp.

C

C

C

C

C

Clostridium spp.

C

Fusobacterium spp.

C

Bacteriodes spp.

C

C

Prevotella spp.

+ b

+1

Porphyromonas spp

+ b

b Pigmented species.

° Prevotella bivia and Prevotella disiens.

a Enterococcus spp.

b Pigmented species.

° Prevotella bivia and Prevotella disiens.

Human Microbial Normal Flora

Potential pathogens

Normal flora

Potential pathogens

Normal flora

FIGURE 1 Normal flora organisms prevent colonization by potential pathogens by physically competing with them on colonization sites and essential nutrients, and by production of bacteriocins.

avidum are rare. P. acnes and P. granulosum are found on skin with a high sebum content; P. acnes is found in all postpubertal individuals; whereas P. granulosum is found in 10% and 20% of individuals in numbers about 100 to 1000 fold fewer than P. acnes. Eubacterium and Peptos-treptococcus may also be encountered.

These microorganisms grow within the sebaceous glands openings and consequently their distribution is proportional to the number of glands, the amount of sebum, and the composition of skin surface lipids (6).

Propionibacteria produce free fatty acids from triglycerides by generating lipase (7). These acids are antibacterial and antifungal and interfere with the growth of nonidigenous microorganism such as Staphylococcus spp., Streptococcus pyogenes, and aerobic gram negative bacilli. These fatty acids may, however, play a deleterious role in the development of acne by causing inflammation (8). The numbers of P. acnes are higher in adults than in young children. Because of their prevalence in the skin and the ear canal, they can contaminate blood cultures and aspirates of cerebrospinal fluid, abscesses, and middle ear fluid.

THE ORAL CAVITY

The establishment of the normal oral flora is initiated at birth. Lactobacilli and Peptostrepto-cocci, reach high numbers within a few days. Actinomyces, Fusobacterium, and Nocardia are acquired by six months. Following that time, Prevotella, Porphyromonas, Leptotrichia, Propionibacterium, and Candida also are established (9). Fusobacterium populations attain high numbers after dentition.

The predominant facultative organisms are the alpha-hemolytic streptococci (the species mitis, milleri, sanguis, intermedins, and salivarius) (10). Other organisms are Moraxella catarrhalis and Haemophillus influenzae, which may cause otitis, sinusitis, or bronchitis. Encapsulated H. influenzae can cause meningitis and bacteremia. The oropharynx also contains Staphylococcus aureus and Staphylococcus epidermidis that can cause chronic infections.

The oropharynx is seldom colonized by Enterobacteriaceae. In contrast, hospitalized patients are often colonized with these organisms. This may be due to selection following the administration of antimicrobials (11) and can contribute to the development of anaerobic gram negative bacilli (AGNB) pneumonia.

Oropharyngeal selective decontamination using topical polymyxin B, neomycin, and vancomycin is effective in reducing colonization and pneumonia with S. aureus and AGNB, without suppression of anaerobes organisms (12).

Anaerobes are present in large numbers in the mouth and the oropharynx, particularly in patients with poor dental hygiene, caries, or periodontal disease (Fig. 2). They outnumber the aerobes 10:1 to 100:1. The predominant anaerobes are Peptostreptococcus, Veillonella, Bacteroides, pigmented Prevotella and Porphyromonas, and Fusobacterium spp., Porphyromonas gingivalis, Bacteroides ureolyticus. Actinomyces spp., treponemas, Leptotrichia buccalis, Bifidobacterium, Eubacterium, and Propionibacterium spp., (1). Some of these organisms are a potential source of chronic infections such as otitis, sinusitis, aspiration pneumonia and lung abscesses, and oropharyngeal and dental abscesses.

Anaerobes can adhere to dental surfaces and contribute through the elaboration of metabolic products to the production of both caries and periodontal disease ranging from gingivitis to periodontitis (10).

Oropharyngeal Flora

Aerobic/Anaerobic

Nasal washings: 10-104/102-105/mL

Tooth surfaces: 106/109-1010/mL

Saliva:

107-108/108-109/mL

Gingival scrapings: 107/1011-1012/mL

Cricothyroid membrane

FIGURE 2 The microbiology of the oral flora.

The oral cavity is an open ecosystem, with a dynamic balance between the entrance of organisms, colonization, and the host defenses directed at their removal. To avoid elimination, bacteria adhere to either hard dental surfaces or epithelial surfaces and form a biofilm. Biofilm is defined as a community of bacteria intimately associated with each other and included within an exopolymer matrix: this biological unit exhibits its own properties. The oral biofilm formation and development have been correlated with all common oral and otolaryngological pathologies, such as dental caries, periodontal disease peri-implantitis otitis, sinusilitis and tonsillitis (13) (Fig. 3) (9).

The recovery rate of aerobic (H. influenzae, M. catarrhalis, and S. aureus) and anaerobic (Prevotella, Porphyromonas, and Fusobacterium) beta-lactamase producing bacteria (BLPB) in the oropharynx has increased in recent years, and these organisms were isolated in over half of the patients with head and neck infections (14). BLPB can protect not only themselves from the activity of penicillin but also penicillin-susceptible organisms as the enzyme is released into the infected tissue or abscess fluid (15). The high incidence of isolation of BLPB may be due to their selection following penicillin therapy (16).

Images Anaerobic BacteriaIsolation Anaerobic Bacterai

THE GASTROINTESTINAL TRACT

Gastrointestinal tract colonization is initiated during delivery as the newborn aspirates cervical canal material (17). The development of the flora is a gradual, and is determined by factors such as composition of the maternal gut micro flora, environmental, and genetic aspects. Variables such as, dietary constituents, gestational age, degree of hygiene, mode of delivery, use of antibiotics or other medication and a need for nursing in incubators, can all effect the microbial colonization (18).

Streptococci, enterococci, and staphylococci usually are present in the first days of life. At the end of one week the fecal flora is predominately anaerobic and contains Bifidobacterium, Bacteroides, and Clostridium spp. The commonest facultative fecal flora is Escherichia coli and E. faecalis (19). Both prematurity and breast feeding were less frequently associated with colonization by anaerobes, B.fragilis was less likely to be recovered in breast-fed infants than in their formula-fed counterparts, and Bifidobacterium predominates in breast-fed infants (20). After weaning the numbers of Bifidobacterium decrease, while Bacteroides increases.

The gut flora plays an essential role in the development of the gut immunity. Intestinal micro-organisms can down-regulate an allergic inflammation by counterbalancing type 2 T-helper cell responses and by enhancing antigen exclusion through an immunoglobulin (Ig)A response (21).

The gastrointestinal flora is dynamic and varies at different locations and levels. These changes depend on factors such as anatomical changes, diet, state of health, and ingestion of medication that alter the stomach acidity, secretory Igs, intestinal motility, and BI (22). Factors that interfere with colonization are active peristalsis, gastric acidity, and high oxidation-reduction potential.

The esophagus, stomach, duodenum, jejunum, and proximal ileum normally contain relatively few bacteria. However, the flora becomes more complex and the number of different bacterial species increases in the distal portions.

Even though the stomach is constantly seeded with oropharyngeal organisms (22), the gastric acidity decreases their number. Those who receive acid reducing medications, or suffer from gastric bleeding have a higher pH, and subsequently more surviving bacteria (23). The bacterial counts in the small intestine are relatively low, with total counts of 102 to 105 aerobic and anaerobic organisms per milliliter. The predominate organisms up to the ileocecal valve are gram-positive facultatives, while Bacteroides (mostly B. fragilis group), Bifidobacterium, Lactobacillus, and coliform predominate below that structure (24). The colon is colonized by the largest numbers of microorganisms of any inhabited region of the human body; 300 to 400 different species and 1012 bacteria per gram fecal material. Approximately 99.9% of these bacteria are anaerobic (ratio aerobes to anaerobes; 1 to 1000 or 10,000) (Fig. 4).

Bacteroides is the predominant bacterial genus in the intestine, present at approximately 1011 organisms per gram dry weight (24). The most frequently isolated are Bacteroides vulgatus, B. thetaiotaomicron, B. distasonis, B. fragilis, and B. ovatus. Among the gram-positive rods, Bifidobacterium adolescents, Eubacterium aerofaciens, Eubacterium lentum, and C. ramosum predominate (24).

B. fragilis group and other AGNB undergoes morphological changes as it transforms itself to become a pathogen (25). About 80% of AGNB recovered from blood and abscesses were encapsulated, while only 10% of stool or pharynx isolates were encapsulated (p < 0.001). Pili were observed in 6% of blood, 75% of abscesses, and 69% of normal flora isolates (p < 0.001).

AGNB expresses different morphological features at different sites as some structures are advantageous or detrimental (25). Pili enables mucosal adherence to those who colonize. Because they are not exposed to macrophages, capsules do not provide them with any advantage. In abscesses, capsules provide protection from macrophages, and pili enable attachment. In contrast, the presence of pili may interfere with systemic spread, since piliated organisms may be more easily phagocytosed (26). The gut harbors numerous AGNB but only those that can adapt to the changing environment can cause illness.

Anatomic and physiologic derangement in the gut can lead to bacterial overgrowth in the upper small bowel (22). This was demonstrated in patients with hypochlorydia, atropic

Oropharynx 1010-1012

Stomach and jejunum 0-105

Stomach and jejunum 0-105

Indigenous Microbial Flora

gastritis, intake of antacids or cimetidine, ineffective peristalsis, multiple diverticula, cirrhosis, chronic malnutrition, excessive small bowel resection, and abdominal irradiation (22). Proliferation of a colonic-type flora in the small intestine can cause a variety of metabolic disturbances, including steatorrhea, vitamin B12 deficiencies, and carbohydrate malabsorption.

Acute diarrhea produces profound alterations in the gut flora. Under certain conditions the resident microflora is eclipsed by a pathogen (27). The rapid transit of diarrheal stool results in a marked reduction in the large bowel anaerobic population. Resolution of diarrhea is accompanied by rapid restitution of the normal flora.

The normal colonic flora is relatively constant and constitutes a defense mechanism against infections by pathogens. Suppression of the anaerobic flora by antimicrobials effective against most anaerobic bacteria except C. difficile, can cause pseudomembranous colitis (28). The ability of colonic flora to interfere with the establishment of pathogens is termed "colonization resistance" (29). Antibiotics effective against anaerobes increase the gut population and subsequently the potentials for translocation of Enterobacteriaceae (30).

Numerous studies utilized selective gut decontamination in an attempt to eradicate only the Enterobacteriaceae and preserve the anaerobes by using antimicrobials that are only effective against Enterobacteriaceae (31). The subjects of these studies were generally immunosupressed individuals and those prone to infections. The antimicrobials were either nonabsorbable (i.e., polymyxin, neomycin, bacitracin) or absorbable (i.e., trimethoprim/ sulfamethoxazole, quinolones) (31). However, there is no consensus yet regarding the practical implications of using selective decontamination.

Oropharynx 1010-1012

VAGINAL AND CERVICAL FLORA

The vagina contains a complex microbial flora (32). Lactobacilli colonize the vagina shortly after birth, because of the mother's hormonal stimulation. As this effect wanes, lactobacilli are replaced with aerobic gram-positive cocci. At puberty the cyclic hormonal stimulation ensues, the squamous epithelium glycogen content increases and lactobacilli returns. Lactobacilli metabolize glycogen, producing lactic acid, which contributes to a low vaginal pH (4.5-5.5) in adults. The low pH select for certain microorganisms, such as Candida and anaerobes, but inhibits the growth of fastidious bacteria including Enterobacteriaceae.

The mean bacterial counts in the vagina and cervix are approximately 10s organisms/ml. About 50% of these are anaerobic (32). The cervical canal contains mixed aerobic and anaerobic flora. The aerobic components consist of lactobacilli, group B streptococci, Enterococcus spp., S. epidermidis, S. aureus, and Enterobacteriaceae.

The anaerobic component consists predominately of lactobacillus and peptostreptococci. Clostridium spp. include bifermentans, perfringens, ramosum, and difficile. The predominant gram negative bacilli are P. disiens, P. bivia, pigmented Prevotella and Porphyromonas, B. fragilis, and Prevotella oralis. Veillonella, bifidobacteria, and eubacteria are also present.

Variations in cervical-vaginal flora are related to the effects of age, pregnancy, and menstrual cycle. Estrogen can increase the bacterial population of the female genital tract, while progesterone decreases it (33). The flora before puberty, during childbearing years, pregnancy, and after menopause is not uniform. Colonization with lactobacilli is low in children and in postclimactic years, and is high in pregnancy and the reproductive years.

The influence of pregnancy on the vaginal flora is important because the newborn is exposed to it during delivery or through exposure to infected amniotic fluid (17). The major change during pregnancy is an increase in the colonization by lactobacilli (17,32). This increase in the number of non-virulent lactobacilli at the expense of the more virulent microorganisms may serve to protect the fetus from exposure to pathogens.

COLONIZATION OF GASTROINTESTINAL TRACT IN THE NORMAL INFANT

The developing fetus is protected from the bacterial flora of the maternal genital tract. Initial colonization of the newborn and of the placenta usually occurs after rupture of the maternal membranes. During a vaginal delivery the neonate is exposed to the cervical birth canal flora, which includes many aerobic and anaerobic bacteria (34,35).

The predominant aerobic bacteria present in the cervical flora are staphylococci, diphtheroids, alpha-hemolytic streptococci, Gardnerella vaginalis, lactobacilli, and E. coli. The most common anaerobic organisms are Prevotella bivia, Prevotella disiens, B. fragilis group, P. acnes, Peptostreptococci, pigmented prevotella and porphyromonas, clostridia, and lactobacilli (36).

The newborn is colonized initially on the skin and mucosa of the nasopharynx, oropharynx, conjunctivae, umbilical cord, and the external genitalia. In most infants, the organisms colonize these sites without causing any inflammatory changes.

The colonization of the gastrointestinal tract by bacteria begins immediately after delivery. Conjunctival and gastric contents of vaginally delivered infants contain many aerobic and anaerobic bacteria that are identical to the maternal genital flora (17,37). As the newborn infant's birth weight and duration of pregnancy increased, more potentially pathogenic aerobic (such as E. coli and S. aureus) and anaerobic bacteria (such as the B. fragilis group) were found in gastric contents; also prolongation of labor brought about increased numbers of anaerobes. These organisms represent a transient load of bacteria acquired during delivery (38). The only organism whose recovery from gastric aspirates has clinical importance is Group B streptococci (39). This has particular importance in newborns with signs of infection.

The bacterial flora is usually heterogeneous during the first few days of life, independently of feeding habits. After the first week of life, a stable bacterial flora is usually established (40). In full-term infants a diet of breast milk induces the development of a flora rich in Bifidobacterium spp. Other obligate anaerobes, such as Clostridium spp. and Bacteroides spp., are rarely isolated and also Enterobacteriaceae and enterococci are relatively few. During the corresponding period, formula-fed babies are often colonized by other anaerobes in addition to bifidobacteria and by facultatively anaerobic bacteria.

The initially sterile meconium becomes colonized in most instances within 24 hours with aerobic and anaerobic bacteria, predominantly micrococci, E. coli, Clostridium spp., and streptococci (41). The presence of various types of clostridia can be demonstrated at that age (18,42,43). Facultatively anaerobic bacteria colonize from the first days of life followed closely by bifidobacteria. The number of facultative bacteria fall by the third day, and the suppression is attributed to the establishment of an acetate and acetic acid buffer of low pH in the intestinal lumen (44). Bifidobacteria reach high levels to become the predominant organisms, although other anaerobes such as Bacteroides spp., clostridia, and anaerobic streptococci are also present.

Several factors influence the composition of the fecal bacterial flora. These include the type of feeding (breast or formula), the route of delivery, gestational age term and exposure to antimicrobials. Anaerobes other than bifidobacteria tend not to persist in breast-fed infants during the period of exclusive breast feeding (45). Formula-fed neonates harbor higher number of facultative anaerobes, and colonization by bifidobacteria generally is slower compared to breast-fed infants (46). Anaerobic bacteria other than bifidobacteria are also found in the feces of formula-fed infants during the first week of life, and these persist beyond the neonatal period. The isolation rates of B. fragilis and other anaerobic bacteria in term babies approach that of adults within a week. The percentage of stools containing anaerobic bacteria increased with age and by four or six days of age 96% of infants were colonized with anaerobic bacteria, and 61% were colonized with B. fragilis. E. coli, Klebsiella spp., Enterobacter spp., and Proteus spp. were the most frequently colonizing aerobic gram-negative bacilli.

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Responses

  • SINIKKA ROVANPER
    Why do obligate anaerobes need to grow without oxygen?
    7 years ago
  • stefan
    Is morexella catarrhalis aerobic or facultative anaerobic?
    6 years ago
  • Oskar Johnston
    Is Prevotella bivia anerobic or aerobic bacteria?
    6 years ago
  • dirk
    Is prevotella bivia part of your normal vaginal flora?
    6 years ago

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