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Streptococcus bovis: Answers and questions
Clinical Microbiology Newsletter Vol. 29, No. 7
www.cmnewsletter.com
April 1, 2007
Streptococcus bovis: Answers and Questions Laura J. Tafe, M.D. and Kathryn L. Ruoff, Ph.D., Department of Pathology, Dartmouth Hitchcock Medical Center, Lebanon, New Hampshire
Abstract Bacteremia caused by Streptococcus bovis can indicate serious conditions, like endocarditis and colonic cancer. S. bovis has also been documented as an agent of central nervous system and abdominal infections and osteomyelitis. The link between S. bovis and colonic cancer is still not well understood, and observations on this relationship may have been clouded in the past by the inability to accurately identify S. bovis and related organisms. This article explores the clinical significance of bacteria identified as S. bovis, their link with gastrointestinal cancer, and the evolution of the taxonomy and classification of these organisms. Better methods of identification of S. bovis organisms may aid in unraveling the true clinical role of these bacteria.
Introduction Streptococcus bovis was first described as a species isolated from the milk and dung of cows. From these humble beginnings, organisms resembling the original S. bovis have been divided into an array of new taxonomic groups isolated from animals and humans. Sherman, in his seminal 1937 review of the streptococci, observed that S. bovis, first described in 1919 by Orla-Jensen, bore a strong resemblance to an organism known as the “Bargen streptococcus.” Bargen had hypothesized that this streptococcus, isolated from human stool cultures, was the causative agent of ulcerative colitis (1). Thus, the groundwork was laid for the association of S. bovis with colonic cancer. S. bovis has also been documented as an agent of endocarditis and other infections. This article will trace the changes in classification of S. bovis and review the significance of these bacteria
Mailing address: Kathryn L. Ruoff, Department of Pathology, Dartmouth Hitchcock Medical Center, One Medical Center Dr., Lebanon, NH 03756-0001. Tel.: 603650-8728. Fax: 603-650-4845. E-mail: Kathryn.L.Ruoff@ Hitchcock.org Clinical Microbiology Newsletter 29:7,2007
in human infection. The reader should note that while the name “S. bovis” is used throughout much of this article, the new species and subspecies included in the current classification scheme for theses organisms are described in the sections on microbiology and taxonomy and in Table 1. S. bovis endocarditis and the link with colonic cancer Infective endocarditis is a serious cardiac disease that is associated with a number of causative agents, including S. bovis. Several review articles have dealt with comparisons of the clinical, echocardiographic, surgical rate, mortality rate, and associated pathologic findings in patients with S. bovis endocarditis versus those with other causative microorganisms. All patients reviewed met the Duke criteria for the clinical diagnosis of infective endocarditis as classified by: (i) two major criteria, (ii) one major and three minor criteria, or (iii) five minor criteria (2). In 1998, Kupferwasser et al. reviewed 177 consecutive patients with infective endocarditis (22 with S. bovis infection) during a 13-year period in Mainz, Germany. Statistically significant (P < 0.05) clinical findings included S. bovis asso© 2007 Elsevier
ciated with older patients (67 years of age; range, 49 to 76), the overall need for surgical treatment/intervention (73%), delay of therapeutic intervention (14 days; standard deviation [SD], ±6 days), and a higher mortality rate (45% versus 25%; P, 0.04). Echocardiography (transthoracic and/or transesophageal) suggested that the simultaneous involvement of two or three valves was a common finding in those with S. bovis endocarditis. The mean vegetations per valve were 1.8 (SD, 0.7) versus 1.3 (SD, 0.5) in those with non-S. bovis endocarditis (P, 0.05). The valvar damage in S. bovis endocarditis led to a higher percentage of affected patients presenting with valvar regurgitation (86% versus 76%; P, 0.05), which also led to a higher number of surgical
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Table 1. Phenotypic characteristics and taxonomic and nomenclature changes in organisms identified as S. bovisa S. Bovis Biotype
Former nomenclature
Currently proposed nomenclature
Mannitol fermentation
Starch hydrolysis
Esculin hydrolysis
Betaglucuronidase
I
S. gallolyticus
S. gallolyticus subsp. gallolyticus
+
+
+
–
II/2
S. gallolyticus, S. pasteurianus
S. gallolyticus subsp. pasteurianus –
–
+
+
II/1
S. infantarius
S. infantarius subsp. infantarius
–
+
V
–
II/1
S. infantarius, S. infantarius subsp. coli
S. lutetiensis
–
V
+
–
a
+, positive; -, negative; V, variable. Information in this table is based on publications by Facklam (32) and Schlegel et al. (29) and other literature cited in the text.
interventions. Interestingly, there were significantly fewer cerebral or peripheral embolic events in S. bovis endocarditis patients, which the authors suggested appeared to be associated with the morphology of the valvar vegetations: typically smaller, fixed, and without oscillating parts, which are features of stability making them less likely to embolize. The increased mortality in patients with S. bovis could have been due to the greater age of these patients and the increased likelihood that they had underlying extracardiac comorbidities. Therapeutic delays, the increased rates of congestive heart failure due to regurgitation, and the need for surgical interventions were also likely to contribute (3). A number of studies have shown a correlation between S. bovis and colonic carcinoma. In Kupferwasser’s review, 21 of the 22 patients underwent gastroscopy and colonoscopy, and in 10 patients (45%), 16 lesions were identified (two gastric ulcers, one gastric cancer, two duodenal ulcers, two colon cancers, four colon adenomas, one inflammatory bowel disease, two colonic diverticula, one angiodysplasia, and one liver cirrhosis). A variety of gastrointestinal-systemassociated lesions were identified in
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these patients rather than a single characteristic lesion. Some authors believe that S. bovis endocarditis patients should be followed up for 2 years after the infective endocarditis episode for subsequent development of gastrointestinal malignancies (3). In 2003, Gonzalez-Juanatey et al. (4) reviewed 20 cases of S. bovis endocarditis over a 13-year period in a welldefined area of southern Europe with some similar conclusions. The patients were older (63.1 versus 58.6 years; P, 0.13), and 95% were men (P, 0.08), but these findings were not significant (P < 0.05). All patients had an abnormal transthoracic echocardiogram, with 19 of 20 patients showing aortic valve involvement (P, 0.01) and 11/20 (55%) with both aortic and mitral valve involvement. All but one patient had developed valve regurgitation (65.5% with grade III or IV regurgitation; P, 0.02), and 40% experienced heart failure. Non-significant findings included the number of embolic events, the proportion of patients requiring valve replacement, mortality rates during the active phase of the disease, and the causes of death. The overall mortality was 40%, which is similar to that reported by Kupferwasser and coworkers. Perhaps the differences in
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surgical interventions mentioned in both review articles had to do with differences in treatment philosophies of the two different countries in which the data were acquired. In the review of Gonzalez-Juanatey et al., 13 of 20 patients underwent a colonoscopy during hospitalization and 8 of the 13 (61.5%) patients had colonic neoplasms: seven adenomatous polyps and one adenocarcinoma. One patient had diverticular disease. Upon followup, one patient with an adenomatous polyp developed adenocarcinoma 6 years after the acute endocarditis event (4). A low incidence of embolic events has not been observed in all studies. In a prospective study of 199 patients with infective endocarditis, Tripodi and colleagues (5), found 34 embolic events, with 22 of them occurring among the 30 patients who had S. bovis endocarditis. The events involved the brain, spleen, kidney, and lung and were significantly more frequent than in nonS. bovis endocarditis patients (73.3% versus 40.2%; P, 0.002). In 2002, investigators from the Mayo Clinic published an evaluation of S. bovis strains that were collected from 1975 to 1985. Fourteen patients with infective
Clinical Microbiology Newsletter 29:7,2007
endocarditis were included in their study and 10 of these underwent colonic evaluation. Two had colon cancer and 6 had other gastrointestinal pathologies (2 with ulcerative colitis, one with sigmoid colon villous adenoma, one with cecal polyp, one with gastric adenomatous polyps, and one with sigmoid diverticulosis (6). Treatment for S. bovis endocarditis consists primarily of single-agent therapy with penicillin G for 4 weeks. Alternatively, penicillin G plus gentamicin for 2 weeks can be used. Ceftriaxone for 4 weeks is recommended for patients with non-immediate penicillin allergies. Vancomycin for 4 weeks may be used in patients with severe or immediate β-lactam allergy (2). As has been previously mentioned, there appears to be a correlation between S. bovis and colon cancer. S. bovis is occasionally present in the human colonic flora, and it has been reported that fecal carriage of the bacteria is increased in colon cancer patients. Various studies have shown that 25 to 80% of patients with S. bovis bacteremia also had colorectal adenomatous polyps or carcinoma (7). Beebe and Koneman reviewed 23 studies and case reports of patients with S. bovis bacteremia and/or endocarditis. In total, there were 202 patients, 98 (48.5%) of whom exhibited a gastrointestinal disorder; 40 (19.8%) had carcinoma, 34 (16.8%) had adenomatous polyps, and 24 (12%) had other disorders (not specified) (8). Interestingly, there have been case studies reporting S. bovis bacteremia in patients with pancreatic cancer, endometrial cancer, metastatic melanoma, esophageal and gastric carcinoma, and lymphomas and other malignancies (8,9). Further studies are warranted to determine the nature of this relationship. Carcinogenesis of colon cancer The relationship between S. bovis and colonic carcinoma has been established; however, the mechanism of carcinogenesis is not known. Bacteria have been linked to cancer via two possible mechanisms: induction of chronic inflammation and the production of carcinogenic metabolites. Most colorectal carcinomas occur sporadically (with the exception of well-defined familial syndromes) with a well-described set of genetic mutations that ultimately lead to cancer. Clinical Microbiology Newsletter 29:7,2007
Two pathogenetically distinct models have been described. The first is the adenoma-carcinoma sequence proposed by Fearon and Vogelstein, which is thought to explain about 80 to 90% of sporadic colorectal carcinomas. This model is called the APC/β-catenin pathway and involves a stepwise accumulation of mutations in oncogenes, tumor suppressor genes, and overexpression of cyclooxygenase — 2 (COX-2). These changes progress through morphologically identifiable stages from epithelial proliferation to progressively enlarging and more dysplastic adenomas, and ultimately to invasive carcinomas. COX-2, via prostaglandins, promotes cellular proliferation and angiogenesis and inhibits apoptosis, thus acting as a promoter in the cancer pathway. The second model is the microsatellite instability pathway and is involved in 10 to 15% of sporadic carcinomas. In this pathway, there is also accumulation of mutations, but involving different genes, and there does not seem to be a morphologic correlation as there is in the adenoma-carcinoma pathway. (10). In the chronic-inflammation mechanism of carcinogenesis, bacteria present in the gastrointestinal tract induce an inflammatory response that may ultimately contribute to the development of carcinoma. A well-discussed example of this theory is the association of Helicobacter pylori infection with gastric carcinoma. H. pylori infection is associated with a twofold increased risk of gastric cancer. However, infection and subsequent inflammation seem most likely to be promoters in the multistep development of carcinoma (from chronic gastritis to atrophy, intestinal metaplasia, dysplasia, and, ultimately, cancer) rather than the causative agents (11,12). Bacteria in general are thought to contribute to carcinogenesis by formation of potentially toxic by-products of carbohydrate and bile acid metabolism, as well as hydrolysis of other mutagenic precursors (10,11). The gut is colonized by many species of bacteria, and it is nearly impossible to narrow carcinogenesis down to one organism, but it is possible that a specific bacterium may induce a favorable environment for mutagens to inflict their damage (11). Ellmerich and coworkers designed a rat model to determine the effects of S. bovis and its bacterial cell wall antigens © 2007 Elsevier
on early azoxymethane (AMO) induced preneoplastic changes in the intestinal tract (13). They found that, compared to controls, the mucosa of rats receiving S. bovis and S. bovis wall-extracted antigen (WEA) gavages had four- and three-times higher expression of interleukin-8 (IL-8), respectively, as well as increased polyamine content and proliferating cell nuclear antigen (PCNA) expression. IL-8 is an important cytokine in the inflammatory response to bacteria. It is a potent neutrophil and T-cell chemoattractant and may lead to free radical and proteolytic enzyme release from activated neutrophils, which can damage the mucosa (13). In H. pylori infection, gastric levels of IL-8 strongly correlate with the severity of gastritis. In a study by Ellmerich et al., there was a 1.8-fold increase in the number of aberrant colonic crypts (P < 0.01) in rats exposed to S. bovis and S. bovis WEA, as well as presence of colonic adenomas in 50% of the S. bovis WEA group. They also found that healthy rats treated with S. bovis did not develop hyperplastic colonic crypts, which suggests that a preneoplastic lesion may be necessary for the bacteria to have pathological activity in the colonic mucosa. This suggests that S. bovis may be just one of the environmental factors that lead to cancer (14). This group of workers continued their studies of the pro-inflammatory activity of S. bovis WEA. From a WEA purified fraction, they were able to identify 12 different proteins that were able to trigger release of IL-8 and prostaglandin E2 (PGE2) (which correlated with in vitro over-expression of COX-2) from cells of the human colonic epithelial cell line Caco-2. They also showed that these proteins were able to stimulate the phosphorylation of three classes of mitogen-activated protein kinases (MAPKs). When an inhibitor of the MAPKs was added, there was a marked inhibition of IL-8 and PGE2 release by the Caco-2 cells, illustrating that phosphorylation of the MAPKs is necessary for IL-8 and PGE2 release. In in vivo studies in AMO- pre-treated rats, rat IL-8 (CINC/GRO) increased by 12 and 44% in rats administered WEA and the fractionated proteins, respectively, in comparison with AMO controls. Also, the amount of PGE2 was increased by 51% in the rats treated with the frac0196-4399/00 (see frontmatter)
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tionated proteins. Therefore, the results seen in vitro with the Caco-2 cells were reproducible in the in vivo rat model. While one polyp was identified in the WEA-treated rats, five polyps were detected in the fractionated protein treated rats (3/6 rats) and none in the AOM controls (7). Taken together, these studies suggest that S. bovis plays a role in inducing chronic colonic inflammation and probably promotes progression of pre-neoplastic mucosal lesions (at least in the rat model). The release of IL-8, a potent chemoattractant, results in the activation of inflammatory cells, which release more inflammatory mediators and can affect the surrounding colonic mucosa. The release of PGE2 is correlated with the overexpression of COX-2, which is seen in about 85% of colon cancers, and, through its association with enhanced angiogenesis and inhibition of apoptosis, is favorable to the development of carcinoma (7). Liver disease associations Tripodi et al. (5) observed chronic liver disease in 17 (60.7%) of 28 patients with S. bovis endocarditis. The liver disease was associated with viral infection in 14 patients (12 with hepatitus C virus [HCV] and 2 with HBV infection) and alcohol abuse in 2 patients and was cryptogenic in 1 patient. The patterns of liver injury included liver cirrhosis (11 patients), diffuse fibrosis (3 patients), and chronic hepatitis (3 patients). Chronic liver disease among these patients with S. bovis endocarditis was significantly higher than among those with endocarditis of other etiologies (60% versus 15.3%; P < 0.001). Gonzlez-Quintela et al. (15) observed that in 20 cases of S. bovis bacteremia (10 with endocarditis), there were 11 (55%) patients with chronic liver disease, nine of whom had cirrhosis (7 alcohol associated, 3 alcohol and HCV associated, and one HCV associated). It is worth considering that chronic liver disease may predispose to bacteremia due to impaired bacterial clearance from the portal blood (5). S. bovis association with other infections Central nervous system infections (meningitis, abscess, and subdural empyema) have also been reported as being rarely caused by S. bovis. Several 52
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case reports document this finding, and in almost all, the patient had an underlying disease or condition (i.e., gastroenteritis, endocarditis, colonic lesions, polymyalgia rheumatica, or HIV infection). In the five cases for which S. bovis biotyping results are available, all typed as S. bovis biotype II in contrast to the majority of endocarditis isolates, which are biotype I (see Microbiology of S. bovis below) (16-18). Vilaichone et al. (19) documented spontaneous bacterial peritonitis with S. bovis in 14 patients with cirrhosis and 1 with acute alcoholic hepatitis. Clinical characteristics included infection of the elderly with equal male and female frequency. Most patients presented with fever and abdominal pain and distention. The causes of the cirrhosis included cryptogenic, alcoholic, and viral factors. In 10 of the 15 (67%) cases, the isolates were identified as S. bovis biotype II (typing was not available for the five additional cases). S. bovis is also associated with osteomyelitis. In Tripodi et al.’s report of 30 patients with S. bovis endocarditis, a higher incidence of diskitis (23.3% versus 0.6%; P < 0.001) in S. bovis patients versus those with endocarditis from another cause was observed. For the seven patients with diskitis, this was the initial clinical manifestation of S. bovis disease. The etiology of the patients’ diskitis was not clear; it may have been an embolic phenomenon (five had experienced embolic events before presentation) or a result of seeding during bacteremia (5). Microbiology of S. bovis Organisms known as S. bovis are bile esculin-positive, salt-intolerant streptococci with Lancefield’s group D antigen. Thus, they represent the “nonenterococcal” group D streptococci. S. bovis organisms are closely related to the viridans streptococci and are VPpositive and arginine-hydrolysis negative, like members of the S. mutans and S. salivarius viridans species groups. S. bovis strains can be differentiated from S. mutans on the basis of sorbitol fermentation (S. mutans is positive; S. bovis is negative) and from S. salivarius by the ability of S. bovis to ferment starch, its inability to produce urease, and other characteristics (20). Early characterization of human S. bovis isolates suggested that they could © 2007 Elsevier
be subdivided into biotypes on the basis of mannitol fermentation and starch hydrolysis. Typical S. bovis strains (also referred to as biotype I) were mannitol fermentation and starch hydrolysis positive, and S. bovis variant, or biotype II, strains were mannitol fermentation negative and usually starch hydrolysis negative. S. bovis biotype I, but not biotype II, strains produce extracellular polysaccharides when grown on sucrosecontaining media. Additional details of the phenotypic traits of these organisms can be found in the ensuing discussion and in Table 1. Like most streptococci, S. bovis isolates display susceptibility to beta-lactam antibiotics (20). In a recently published prospective 16-year study, Corredoira and colleagues (21) noted the lack of resistance to penicillin, cefotaxime, and vancomycin among 64 S. bovis human isolates but found that 60% of the strains were resistant to erythromycin and trimethoprim-sulfamethoxazole and 50% were resistant to clindamycin. S. bovis taxonomy In 1984, Farrow and coworkers (22) reported on DNA-DNA hybridization studies performed on a large number of strains of S. bovis and the similar species S. equinus, isolated mostly from animals. They were able to divide these organisms into a number of subgroups and concluded that S. bovis and S. equinus were closely related, representing a single species. Knight and Shlaes (23), also employing DNA-DNA reassociation techniques, observed that human S. bovis isolates were not related to the S. bovis/S. equinus group described by Farrow and colleagues. They noted that typical mannitol-fermenting, starchhydrolyzing human strains identified as S. bovis biotype I were related to mannitol and starch hydrolysis-negative S. bovis variant, or biotype, II strains. Another paper on S. bovis DNA-DNA reassociation studies by Coykendall and Gustafson (24) appeared in the same issue of the same journal as Knight and Shlaes’ publication. Coykendall and Gustafson found, like Knight and Shlaes, that typical S. bovis biotype I strains showed homology with some, but not all, of the S. bovis biotype II isolates. Coykendall and Gustafson also examined their strains with the Rapid Strep system (DMS Laboratories, Flemington, NJ), the forerunner of the API 20 Strep Clinical Microbiology Newsletter 29:7,2007
system (bioM´erieux, Hazelwood, MO). With this phenotypic method, the biotype II strains could be subdivided into biotypes II/1 and II/2. Biotype II/2 was identical to biotype I in the hybridization studies, in spite of its phenotypic differences with biotype I in the mannitol fermentation and starch hydrolysis tests (24). A new species resembling S. bovis was proposed in 1995 by Osawa et al. (25), who named the taxon S. gallolyticus because the strains studied were able to decarboxylate gallic acid. This species was proposed on the basis of phenotypic and DNA-DNA reassociation studies. Osawa et al.’s strains were isolated from both animals and humans, and it was suggested on the basis of whole-cell protein analysis studies by Devriese and colleagues (26) that human isolates identified as S. bovis biotypes I and II/2 were members of the new S. gallolyticus species. Conflicting results were published by Clarridge and coworkers (27), who employed 16S rRNA gene sequencing methods. They concluded that human biotype II/2 strains formed a separate species that was different from S. gallolyticus. Poyart and colleagues (28), using superoxide dismutase (sodA) gene sequencing data, proposed that S. bovis biotype II/2 be classified in a separate species, which they named S. pasteurianus. Additional 16S rRNA gene sequencing studies published by Herrero and colleagues (6) suggested that none of the human S. bovis isolates they examined clustered with the S. gallolyticus sequence. The majority of their strains were phenotypically S. bovis I, and their 16S rRNA gene sequences were identical to or differed in one base pair from what was then designated as the type strain of S. bovis (ATCC 43143). Herrero and coworkers also reported that one of their strains was identified as Streptococcus macedonicus, an organism isolated from dairy sources. Soon after the appearance of these two 16S rRNA gene sequencing studies, Schlegel and colleagues (29), employing DNA hybridization methods, advocated the subdivision of S. gallolyticus into three subspecies, S. gallolyticus subsp. gallolyticus (S. bovis biotype I), S. gallolyticus subsp. pasteurianus (S. bovis biotype II/2, formerly called S. Clinical Microbiology Newsletter 29:7,2007
pasteurianus), and S. gallolyticus subsp. macedonicus (isolated from dairy products). One of Clarridge and coworkers’ biotype II/2 strains was included in the study of Schlegel and coworkers and was classified as S. gallolyticus subsp. pasteurianus. The S. bovis type strain (ATCC 43143) examined in Herrero and coworkers’ study was reclassified as S. gallolyticus subsp. gallolyticus when examined in Schlegel’s study. Thus, the new subspecies of S. gallolyticus could accommodate two of the three S. bovis biotypes. The third S. bovis biotype, II/1, contains strains that are beta-glucuronidase negative, in contrast to the biotype II/2 isolates. In 1997, Bouvet and colleagues (30) examined biotype II/1 strains that were unable to hydrolyze esculin and proposed the species name Streptococcus infantarius for this group of organisms. Further investigations by Schlegel and coworkers (31) suggested that S. infantarius should be divided into two subspecies, which would accommodate esculin hydrolysis-variable biotype II/1 strains. S. infantarius subsp. infantarius strains were esculin hydrolysis positive, and S. infantarius subsp. coli strains were variable for esculin hydrolysis. Most recently, Poyart and coworkers (28) proposed that S. infantarius subsp. coli be reclassified as the new species Streptococcus lutetiensis. Most of the taxonomic changes discussed above were chronicled by Facklam in his 2002 review of the evolution of streptococcal classification (32) and are summarized in Table 1. The table contains currently accepted nomenclature (List of Prokaryotic Names with Standing in Nomenclature [http://www.bacterio.cict.fr]), although for ease of comprehension the epithets S. bovis I, II/2, and II/1 will continue to be used in the present discussion.
Implications of Taxonomy for Clinical Microbiologists Early studies that used phenotypic identification methods suggested that the majority of S. bovis human isolates were mannitol-fermenting and starchhydrolyzing biotype I strains that should, in the current classification, be called S. gallolyticus subsp. gallolyticus. Biotype I isolates seemed more likely to be clinically important; they were isolated more often and tended to be more strongly © 2007 Elsevier
associated with endocarditis and gastrointestinal malignancies than biotype II strains (33). Other more recent surveys of have supported the predominance and clinical significance of biotype I isolates (5,6,21). However, Clarridge and coworkers (27) found that biotype II/2 strains were the most common isolates in the collection of S. bovis strains they examined. They noted that their isolates originated from a population composed primarily of adult males, which may have affected the distribution of biotypes. Clinically significant isolates of S. bovis organisms are encountered on a regular basis in the clinical microbiology laboratory, and microbiologists need to be prepared to identify these organisms. If only a few phenotypic tests like reaction on bile esculin medium, presence of Lancefield group D antigen, and growth in 6.5% salt broth, are used, mistakes in identification are likely to occur (34). Commercially available identification systems that rely on a larger number of phenotypic tests are usually fairly accurate at identifying S. bovis isolates. The API 20 Strep system is useful for providing biotypes and has been employed in some of the taxonomic studies of human S. bovis strains. However, no method is perfect, and isolates that are atypical or produce ambiguous identifications should always be expected. All three biotypes of S. bovis (I, II/2 and II/1) have been recovered as significant isolates from cultures of blood and other specimens. Should clinical microbiologists strive to accurately identify these organisms as S. gallolyticus subsp. gallolyticus, S. gallolyticus subsp. pasteurianus, S. infantarius subsp. infantarius, or S. lutetiensis? In a perfect world, the answer would be “yes.” In a perfect world we could do unlimited phenotypic tests and also determine the organism’s 16S rRNA gene sequence. Accurate identification of S. bovis organisms would contribute to a base of knowledge about their incidence, clinical significance, and pathogenic potentials. Unfortunately, for most of us who live in the real world, this would involve an unwarranted expenditure of resources, not to mention all of the explaining we would have to do once the “taxonomically correct” laboratory report was sent out. Current 0196-4399/00 (see frontmatter)
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nomenclature of S. bovis organisms will probably be meaningless to the majority of clinicians. We can realistically provide fairly accurate species and biotype identification with relatively simple methods (e.g., the API 20 Strep system or other phenotypic methods). Biotyping does provide some clinically useful information, even though these designations have no standing in current taxonomy. Table 1 suggests that the biotype designation of an S. bovis isolate can usually predict its current taxonomic standing with fair accuracy. Retaining the S. bovis nomenclature of the previous millennium will not distinguish us as forwardthinking, cutting-edge scientific types, but it might be practical for the present. Eventually the new nomenclature and taxonomy will become better known and accepted by both microbiologists and clinicians. While many studies and observations have been undertaken to explain the clinical significance and taxonomic relationships of S. bovis organisms, much work remains to be done. The intriguing association of S. bovis (and other bacteria) with cancers and what to name the S. bovis isolates we recover in the clinical microbiology laboratory will no doubt occupy many scientists in the future. References 1. Sherman, J.M. 1937. The streptococci. Bacteriol. Rev. 1:3-97. 2. Fauci, A.S. et al. (ed.). 1998. Infective endocarditis, p. 785-791. In Harrison’s principles of internal medicine, 14th ed. McGraw-Hill, New York. 3. Kupferwasser, H.D. et al. 1998. Clinical and morphologic characteristics in Streptococcus bovis endocarditis: a comparison with other causative microorganisms in 177 cases. Heart 80:276-280. 4. Gonzalez-Juanatey, C. et al. 2003. Infective endocarditis due to Streptococcus bovis in a series of nonaddict patients: clinical and morphological characteristics of 20 cases and review of the literature. Can. J. Cardiol. 19:1139-1145. 5. Tripodi, M.F. et al. 2004. Streptococcus bovis endocarditis and its association with chronic liver disease: an underestimated risk factor. Clin. Infect. Dis. 38:1394-1400. 6. Herrero, I.A. et al. 2002. Reevaluation
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of Streptococcus bovis endocarditis cases from 1975 to 1985 by 16S ribosomal DNA sequence analysis. J. Clin. Microbiol. 40:3848-3850. 7. Biarc, J. et al. 2004. Carcinogenic properties of proteins with pro-inflammatory activity from Streptococcus infantarius (formerly S. bovis). Carcinogenesis 25:1477-1484. 8. Beebe, J.L. and E.W. Koneman. 1995. Recovery of uncommon bacteria from blood: association with neoplastic disease. Clin. Microbiol. Rev. 8:347-348. 9. Gold, J.S., S. Bayar, and R.R. Salem. 2004. Association of Streptococcus bovis bacteremia with colonic neoplasia and extracolonic malignancy. Arch. Surg. 139:760-765. 10. Kumar, V., A.K. Abbas, and N. Fausto (ed.). 2005. The gastrointestinal tract, p. 862-865. In Robbins and Cotran pathologic basis of disease, 7th ed. Elsevier, Philadelphia, PA. 11. Parsonett, J. 1995. Bacterial infection as a cause of cancer. Environ. Health Perspect. 103(Suppl. 8):263-268. 12. Leung, W.K. 2006. Helicobacter pylori and gastric neoplasia. Contrib. Microbiol. 13:66-80. 13. Ellmerich, S. et al. 1999. Production of cytokines by monocytes, epithelial and endothelial cells activated by Streptococcus bovis. Cytokine 12:26-31. 14. Ellmerich, S. et al. 2000. Promotion of intestinal carcinogenesis by Streptococcus bovis. Carcinogenesis 21:753-756. 15. Gonzlez-Quintela, A. et al. 2001. Prevalence of liver disease in patients with Streptococcus bovis bacteraemia. J. Infect. 42:116-119. 16. Cohen, L.F. et al. 1997. Streptococcus bovis infection of the central nervous system: report of two cases and review. Clin. Infect. Dis. 25:819-823. 17. Grant, R.J., T.R. Whitehead, and J.E. Orr. 2000. Streptococcus bovis meningitis in an infant. J. Clin. Microbiol. 38:462-463. 18. Vilarrasa, N. et al. 2002. Streptococcus bovis meningitis in a healthy adult patient. Scand. J. Infect. Dis. 34:61-62. 19. Vilaichone, R.K., V. Mahachai, and P. Kullavanijaya. 2002. Spontaneous bacterial peritonitis caused by Streptococcus bovis: case series and review of the literature. Am. J. Gastroenterol. 97:1476-1479. 20. Ruoff, K.L., R.A. Whiley, and D. Beighton. 2003. Streptococcus, p. 405421. In P.R. Murray et al. (ed.), Manual of clinical microbiology, 8th ed. ASM Press, Washington, DC.
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21. Corredoira, J.C. et al. 2005. Clinical characteristics and significance of Streptococcus salivarius bacteremia and Streptococcus bovis bacteremia: a prospective 16-year study. Eur. J. Clin. Microbiol. Infect. Dis. 24:250-255. 22. Farrow, J.A.E. et al. 1984. Taxonomic studies on streptococci of serological groups C, G, and L, and possibly related taxa. Syst. Appl. Microbiol. 5:467-482. 23. Knight, R.G. and D.M. Shlaes. 1985. Physiological characteristics and deoxyribonucleic acid relatedness of human isolates of Streptococcus bovis and Streptococcus bovis (var.). Int. J. Syst. Bacteriol. 35:357-361. 24. Coykendall, A.L. and K.B. Gustafson. 1985. Deoxyribonucleic acid hybridizations among strains of Streptococcus salivarius and Streptococcus bovis. Int. J. Syst. Bacteriol. 35:274-280. 25. Osawa, R., T. Fujisawa, and L.I. Sly. 1995. Streptococcus gallolyticus sp. nov.: gallate degrading organisms formerly assigned to Streptococcus bovis. Syst. Appl. Microbiol. 18:74-78. 26. Devriese, L.A. et al. 1998. Differentiation between Streptococcus gallolyticus strains of human clinical and veterinary origins and Streptococcus bovis strains from the intestinal tracts of ruminants. J. Clin. Microbiol. 36:3520-3523. 27. Clarridge, J.E. et al. 2001. 16S ribosomal DNA sequence analysis distinguishes biotypes of Streptococcus bovis: Streptococcus bovis biotype II/2 is a separate genospecies and the predominant clinical isolate in adult males. J. Clin. Microbiol. 39:1549-1552. 28. Poyart, C., G. Quesne, and P. Trieu-Cuot. 2002. Taxonomic dissection of the Streptococcus bovis group by analysis of manganese-dependent superoxide dismutase gene (sodA) sequences: reclassification of “Streptococcus infantarius subsp. coli” as Streptococcus lutetiensis sp. nov. and of Streptococcus bovis biotype II.2 as Streptococcus pasteurianus sp. nov. Int. J. Syst. Evol. Microbiol. 52:12471255. 29. Schlegel, L. et al. 2003. Reappraisal of the taxonomy of the Streptococcus bovis/Streptococcus equinus complex and related species: description of Streptococcus gallolyticus subsp. gallolyticus subsp. nov., S. gallolyticus subsp. macedonicus subsp. nov. and S. gallolyticus subsp. pasteurianus subsp. nov. Int. J. Syst. Evol. Microbiol. 53:631-645. 30. Bouvet, A. et al. 1997. Streptococcus infantarius sp. nov. related to S. bovis
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and S. equinus. Adv. Exp. Med. Biol. 418:393-395. 31. Schlegel, L. et al. 2000. Streptococcus infantarius subsp. infantarius subsp. nov. and Streptococcus infantarius subsp. coli subsp. nov., isolated from humans and food. Int. J. Syst. Evol.
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Microbiol. 50:1425-1434. 32. Facklam, R. 2002. What happened to the streptococci: overview of taxonomic and nomenclature changes. Clin. Microbiol. Rev. 15:613-630. 33. Ruoff, K.L. et al. 1989. Bacteremia with Streptococcus bovis and Streptococcus
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salivarius: clinical correlates of more accurate identification of isolates. J. Clin. Microbiol. 27:305-308. 34. Ruoff, K.L. et al. 1984. Identification of Streptococcus bovis and Streptococcus salivarius in clinical laboratories. J. Clin. Microbiol. 20:223-226.
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April 1, 2007
Streptococcus bovis: Answers and Questions Laura J. Tafe, M.D. and Kathryn L. Ruoff, Ph.D., Department of Pathology, Dartmouth Hitchcock Medical Center, Lebanon, New Hampshire
Abstract Bacteremia caused by Streptococcus bovis can indicate serious conditions, like endocarditis and colonic cancer. S. bovis has also been documented as an agent of central nervous system and abdominal infections and osteomyelitis. The link between S. bovis and colonic cancer is still not well understood, and observations on this relationship may have been clouded in the past by the inability to accurately identify S. bovis and related organisms. This article explores the clinical significance of bacteria identified as S. bovis, their link with gastrointestinal cancer, and the evolution of the taxonomy and classification of these organisms. Better methods of identification of S. bovis organisms may aid in unraveling the true clinical role of these bacteria.
Introduction Streptococcus bovis was first described as a species isolated from the milk and dung of cows. From these humble beginnings, organisms resembling the original S. bovis have been divided into an array of new taxonomic groups isolated from animals and humans. Sherman, in his seminal 1937 review of the streptococci, observed that S. bovis, first described in 1919 by Orla-Jensen, bore a strong resemblance to an organism known as the “Bargen streptococcus.” Bargen had hypothesized that this streptococcus, isolated from human stool cultures, was the causative agent of ulcerative colitis (1). Thus, the groundwork was laid for the association of S. bovis with colonic cancer. S. bovis has also been documented as an agent of endocarditis and other infections. This article will trace the changes in classification of S. bovis and review the significance of these bacteria
Mailing address: Kathryn L. Ruoff, Department of Pathology, Dartmouth Hitchcock Medical Center, One Medical Center Dr., Lebanon, NH 03756-0001. Tel.: 603650-8728. Fax: 603-650-4845. E-mail: Kathryn.L.Ruoff@ Hitchcock.org Clinical Microbiology Newsletter 29:7,2007
in human infection. The reader should note that while the name “S. bovis” is used throughout much of this article, the new species and subspecies included in the current classification scheme for theses organisms are described in the sections on microbiology and taxonomy and in Table 1. S. bovis endocarditis and the link with colonic cancer Infective endocarditis is a serious cardiac disease that is associated with a number of causative agents, including S. bovis. Several review articles have dealt with comparisons of the clinical, echocardiographic, surgical rate, mortality rate, and associated pathologic findings in patients with S. bovis endocarditis versus those with other causative microorganisms. All patients reviewed met the Duke criteria for the clinical diagnosis of infective endocarditis as classified by: (i) two major criteria, (ii) one major and three minor criteria, or (iii) five minor criteria (2). In 1998, Kupferwasser et al. reviewed 177 consecutive patients with infective endocarditis (22 with S. bovis infection) during a 13-year period in Mainz, Germany. Statistically significant (P < 0.05) clinical findings included S. bovis asso© 2007 Elsevier
ciated with older patients (67 years of age; range, 49 to 76), the overall need for surgical treatment/intervention (73%), delay of therapeutic intervention (14 days; standard deviation [SD], ±6 days), and a higher mortality rate (45% versus 25%; P, 0.04). Echocardiography (transthoracic and/or transesophageal) suggested that the simultaneous involvement of two or three valves was a common finding in those with S. bovis endocarditis. The mean vegetations per valve were 1.8 (SD, 0.7) versus 1.3 (SD, 0.5) in those with non-S. bovis endocarditis (P, 0.05). The valvar damage in S. bovis endocarditis led to a higher percentage of affected patients presenting with valvar regurgitation (86% versus 76%; P, 0.05), which also led to a higher number of surgical
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Table 1. Phenotypic characteristics and taxonomic and nomenclature changes in organisms identified as S. bovisa S. Bovis Biotype
Former nomenclature
Currently proposed nomenclature
Mannitol fermentation
Starch hydrolysis
Esculin hydrolysis
Betaglucuronidase
I
S. gallolyticus
S. gallolyticus subsp. gallolyticus
+
+
+
–
II/2
S. gallolyticus, S. pasteurianus
S. gallolyticus subsp. pasteurianus –
–
+
+
II/1
S. infantarius
S. infantarius subsp. infantarius
–
+
V
–
II/1
S. infantarius, S. infantarius subsp. coli
S. lutetiensis
–
V
+
–
a
+, positive; -, negative; V, variable. Information in this table is based on publications by Facklam (32) and Schlegel et al. (29) and other literature cited in the text.
interventions. Interestingly, there were significantly fewer cerebral or peripheral embolic events in S. bovis endocarditis patients, which the authors suggested appeared to be associated with the morphology of the valvar vegetations: typically smaller, fixed, and without oscillating parts, which are features of stability making them less likely to embolize. The increased mortality in patients with S. bovis could have been due to the greater age of these patients and the increased likelihood that they had underlying extracardiac comorbidities. Therapeutic delays, the increased rates of congestive heart failure due to regurgitation, and the need for surgical interventions were also likely to contribute (3). A number of studies have shown a correlation between S. bovis and colonic carcinoma. In Kupferwasser’s review, 21 of the 22 patients underwent gastroscopy and colonoscopy, and in 10 patients (45%), 16 lesions were identified (two gastric ulcers, one gastric cancer, two duodenal ulcers, two colon cancers, four colon adenomas, one inflammatory bowel disease, two colonic diverticula, one angiodysplasia, and one liver cirrhosis). A variety of gastrointestinal-systemassociated lesions were identified in
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these patients rather than a single characteristic lesion. Some authors believe that S. bovis endocarditis patients should be followed up for 2 years after the infective endocarditis episode for subsequent development of gastrointestinal malignancies (3). In 2003, Gonzalez-Juanatey et al. (4) reviewed 20 cases of S. bovis endocarditis over a 13-year period in a welldefined area of southern Europe with some similar conclusions. The patients were older (63.1 versus 58.6 years; P, 0.13), and 95% were men (P, 0.08), but these findings were not significant (P < 0.05). All patients had an abnormal transthoracic echocardiogram, with 19 of 20 patients showing aortic valve involvement (P, 0.01) and 11/20 (55%) with both aortic and mitral valve involvement. All but one patient had developed valve regurgitation (65.5% with grade III or IV regurgitation; P, 0.02), and 40% experienced heart failure. Non-significant findings included the number of embolic events, the proportion of patients requiring valve replacement, mortality rates during the active phase of the disease, and the causes of death. The overall mortality was 40%, which is similar to that reported by Kupferwasser and coworkers. Perhaps the differences in
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surgical interventions mentioned in both review articles had to do with differences in treatment philosophies of the two different countries in which the data were acquired. In the review of Gonzalez-Juanatey et al., 13 of 20 patients underwent a colonoscopy during hospitalization and 8 of the 13 (61.5%) patients had colonic neoplasms: seven adenomatous polyps and one adenocarcinoma. One patient had diverticular disease. Upon followup, one patient with an adenomatous polyp developed adenocarcinoma 6 years after the acute endocarditis event (4). A low incidence of embolic events has not been observed in all studies. In a prospective study of 199 patients with infective endocarditis, Tripodi and colleagues (5), found 34 embolic events, with 22 of them occurring among the 30 patients who had S. bovis endocarditis. The events involved the brain, spleen, kidney, and lung and were significantly more frequent than in nonS. bovis endocarditis patients (73.3% versus 40.2%; P, 0.002). In 2002, investigators from the Mayo Clinic published an evaluation of S. bovis strains that were collected from 1975 to 1985. Fourteen patients with infective
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endocarditis were included in their study and 10 of these underwent colonic evaluation. Two had colon cancer and 6 had other gastrointestinal pathologies (2 with ulcerative colitis, one with sigmoid colon villous adenoma, one with cecal polyp, one with gastric adenomatous polyps, and one with sigmoid diverticulosis (6). Treatment for S. bovis endocarditis consists primarily of single-agent therapy with penicillin G for 4 weeks. Alternatively, penicillin G plus gentamicin for 2 weeks can be used. Ceftriaxone for 4 weeks is recommended for patients with non-immediate penicillin allergies. Vancomycin for 4 weeks may be used in patients with severe or immediate β-lactam allergy (2). As has been previously mentioned, there appears to be a correlation between S. bovis and colon cancer. S. bovis is occasionally present in the human colonic flora, and it has been reported that fecal carriage of the bacteria is increased in colon cancer patients. Various studies have shown that 25 to 80% of patients with S. bovis bacteremia also had colorectal adenomatous polyps or carcinoma (7). Beebe and Koneman reviewed 23 studies and case reports of patients with S. bovis bacteremia and/or endocarditis. In total, there were 202 patients, 98 (48.5%) of whom exhibited a gastrointestinal disorder; 40 (19.8%) had carcinoma, 34 (16.8%) had adenomatous polyps, and 24 (12%) had other disorders (not specified) (8). Interestingly, there have been case studies reporting S. bovis bacteremia in patients with pancreatic cancer, endometrial cancer, metastatic melanoma, esophageal and gastric carcinoma, and lymphomas and other malignancies (8,9). Further studies are warranted to determine the nature of this relationship. Carcinogenesis of colon cancer The relationship between S. bovis and colonic carcinoma has been established; however, the mechanism of carcinogenesis is not known. Bacteria have been linked to cancer via two possible mechanisms: induction of chronic inflammation and the production of carcinogenic metabolites. Most colorectal carcinomas occur sporadically (with the exception of well-defined familial syndromes) with a well-described set of genetic mutations that ultimately lead to cancer. Clinical Microbiology Newsletter 29:7,2007
Two pathogenetically distinct models have been described. The first is the adenoma-carcinoma sequence proposed by Fearon and Vogelstein, which is thought to explain about 80 to 90% of sporadic colorectal carcinomas. This model is called the APC/β-catenin pathway and involves a stepwise accumulation of mutations in oncogenes, tumor suppressor genes, and overexpression of cyclooxygenase — 2 (COX-2). These changes progress through morphologically identifiable stages from epithelial proliferation to progressively enlarging and more dysplastic adenomas, and ultimately to invasive carcinomas. COX-2, via prostaglandins, promotes cellular proliferation and angiogenesis and inhibits apoptosis, thus acting as a promoter in the cancer pathway. The second model is the microsatellite instability pathway and is involved in 10 to 15% of sporadic carcinomas. In this pathway, there is also accumulation of mutations, but involving different genes, and there does not seem to be a morphologic correlation as there is in the adenoma-carcinoma pathway. (10). In the chronic-inflammation mechanism of carcinogenesis, bacteria present in the gastrointestinal tract induce an inflammatory response that may ultimately contribute to the development of carcinoma. A well-discussed example of this theory is the association of Helicobacter pylori infection with gastric carcinoma. H. pylori infection is associated with a twofold increased risk of gastric cancer. However, infection and subsequent inflammation seem most likely to be promoters in the multistep development of carcinoma (from chronic gastritis to atrophy, intestinal metaplasia, dysplasia, and, ultimately, cancer) rather than the causative agents (11,12). Bacteria in general are thought to contribute to carcinogenesis by formation of potentially toxic by-products of carbohydrate and bile acid metabolism, as well as hydrolysis of other mutagenic precursors (10,11). The gut is colonized by many species of bacteria, and it is nearly impossible to narrow carcinogenesis down to one organism, but it is possible that a specific bacterium may induce a favorable environment for mutagens to inflict their damage (11). Ellmerich and coworkers designed a rat model to determine the effects of S. bovis and its bacterial cell wall antigens © 2007 Elsevier
on early azoxymethane (AMO) induced preneoplastic changes in the intestinal tract (13). They found that, compared to controls, the mucosa of rats receiving S. bovis and S. bovis wall-extracted antigen (WEA) gavages had four- and three-times higher expression of interleukin-8 (IL-8), respectively, as well as increased polyamine content and proliferating cell nuclear antigen (PCNA) expression. IL-8 is an important cytokine in the inflammatory response to bacteria. It is a potent neutrophil and T-cell chemoattractant and may lead to free radical and proteolytic enzyme release from activated neutrophils, which can damage the mucosa (13). In H. pylori infection, gastric levels of IL-8 strongly correlate with the severity of gastritis. In a study by Ellmerich et al., there was a 1.8-fold increase in the number of aberrant colonic crypts (P < 0.01) in rats exposed to S. bovis and S. bovis WEA, as well as presence of colonic adenomas in 50% of the S. bovis WEA group. They also found that healthy rats treated with S. bovis did not develop hyperplastic colonic crypts, which suggests that a preneoplastic lesion may be necessary for the bacteria to have pathological activity in the colonic mucosa. This suggests that S. bovis may be just one of the environmental factors that lead to cancer (14). This group of workers continued their studies of the pro-inflammatory activity of S. bovis WEA. From a WEA purified fraction, they were able to identify 12 different proteins that were able to trigger release of IL-8 and prostaglandin E2 (PGE2) (which correlated with in vitro over-expression of COX-2) from cells of the human colonic epithelial cell line Caco-2. They also showed that these proteins were able to stimulate the phosphorylation of three classes of mitogen-activated protein kinases (MAPKs). When an inhibitor of the MAPKs was added, there was a marked inhibition of IL-8 and PGE2 release by the Caco-2 cells, illustrating that phosphorylation of the MAPKs is necessary for IL-8 and PGE2 release. In in vivo studies in AMO- pre-treated rats, rat IL-8 (CINC/GRO) increased by 12 and 44% in rats administered WEA and the fractionated proteins, respectively, in comparison with AMO controls. Also, the amount of PGE2 was increased by 51% in the rats treated with the frac0196-4399/00 (see frontmatter)
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tionated proteins. Therefore, the results seen in vitro with the Caco-2 cells were reproducible in the in vivo rat model. While one polyp was identified in the WEA-treated rats, five polyps were detected in the fractionated protein treated rats (3/6 rats) and none in the AOM controls (7). Taken together, these studies suggest that S. bovis plays a role in inducing chronic colonic inflammation and probably promotes progression of pre-neoplastic mucosal lesions (at least in the rat model). The release of IL-8, a potent chemoattractant, results in the activation of inflammatory cells, which release more inflammatory mediators and can affect the surrounding colonic mucosa. The release of PGE2 is correlated with the overexpression of COX-2, which is seen in about 85% of colon cancers, and, through its association with enhanced angiogenesis and inhibition of apoptosis, is favorable to the development of carcinoma (7). Liver disease associations Tripodi et al. (5) observed chronic liver disease in 17 (60.7%) of 28 patients with S. bovis endocarditis. The liver disease was associated with viral infection in 14 patients (12 with hepatitus C virus [HCV] and 2 with HBV infection) and alcohol abuse in 2 patients and was cryptogenic in 1 patient. The patterns of liver injury included liver cirrhosis (11 patients), diffuse fibrosis (3 patients), and chronic hepatitis (3 patients). Chronic liver disease among these patients with S. bovis endocarditis was significantly higher than among those with endocarditis of other etiologies (60% versus 15.3%; P < 0.001). Gonzlez-Quintela et al. (15) observed that in 20 cases of S. bovis bacteremia (10 with endocarditis), there were 11 (55%) patients with chronic liver disease, nine of whom had cirrhosis (7 alcohol associated, 3 alcohol and HCV associated, and one HCV associated). It is worth considering that chronic liver disease may predispose to bacteremia due to impaired bacterial clearance from the portal blood (5). S. bovis association with other infections Central nervous system infections (meningitis, abscess, and subdural empyema) have also been reported as being rarely caused by S. bovis. Several 52
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case reports document this finding, and in almost all, the patient had an underlying disease or condition (i.e., gastroenteritis, endocarditis, colonic lesions, polymyalgia rheumatica, or HIV infection). In the five cases for which S. bovis biotyping results are available, all typed as S. bovis biotype II in contrast to the majority of endocarditis isolates, which are biotype I (see Microbiology of S. bovis below) (16-18). Vilaichone et al. (19) documented spontaneous bacterial peritonitis with S. bovis in 14 patients with cirrhosis and 1 with acute alcoholic hepatitis. Clinical characteristics included infection of the elderly with equal male and female frequency. Most patients presented with fever and abdominal pain and distention. The causes of the cirrhosis included cryptogenic, alcoholic, and viral factors. In 10 of the 15 (67%) cases, the isolates were identified as S. bovis biotype II (typing was not available for the five additional cases). S. bovis is also associated with osteomyelitis. In Tripodi et al.’s report of 30 patients with S. bovis endocarditis, a higher incidence of diskitis (23.3% versus 0.6%; P < 0.001) in S. bovis patients versus those with endocarditis from another cause was observed. For the seven patients with diskitis, this was the initial clinical manifestation of S. bovis disease. The etiology of the patients’ diskitis was not clear; it may have been an embolic phenomenon (five had experienced embolic events before presentation) or a result of seeding during bacteremia (5). Microbiology of S. bovis Organisms known as S. bovis are bile esculin-positive, salt-intolerant streptococci with Lancefield’s group D antigen. Thus, they represent the “nonenterococcal” group D streptococci. S. bovis organisms are closely related to the viridans streptococci and are VPpositive and arginine-hydrolysis negative, like members of the S. mutans and S. salivarius viridans species groups. S. bovis strains can be differentiated from S. mutans on the basis of sorbitol fermentation (S. mutans is positive; S. bovis is negative) and from S. salivarius by the ability of S. bovis to ferment starch, its inability to produce urease, and other characteristics (20). Early characterization of human S. bovis isolates suggested that they could © 2007 Elsevier
be subdivided into biotypes on the basis of mannitol fermentation and starch hydrolysis. Typical S. bovis strains (also referred to as biotype I) were mannitol fermentation and starch hydrolysis positive, and S. bovis variant, or biotype II, strains were mannitol fermentation negative and usually starch hydrolysis negative. S. bovis biotype I, but not biotype II, strains produce extracellular polysaccharides when grown on sucrosecontaining media. Additional details of the phenotypic traits of these organisms can be found in the ensuing discussion and in Table 1. Like most streptococci, S. bovis isolates display susceptibility to beta-lactam antibiotics (20). In a recently published prospective 16-year study, Corredoira and colleagues (21) noted the lack of resistance to penicillin, cefotaxime, and vancomycin among 64 S. bovis human isolates but found that 60% of the strains were resistant to erythromycin and trimethoprim-sulfamethoxazole and 50% were resistant to clindamycin. S. bovis taxonomy In 1984, Farrow and coworkers (22) reported on DNA-DNA hybridization studies performed on a large number of strains of S. bovis and the similar species S. equinus, isolated mostly from animals. They were able to divide these organisms into a number of subgroups and concluded that S. bovis and S. equinus were closely related, representing a single species. Knight and Shlaes (23), also employing DNA-DNA reassociation techniques, observed that human S. bovis isolates were not related to the S. bovis/S. equinus group described by Farrow and colleagues. They noted that typical mannitol-fermenting, starchhydrolyzing human strains identified as S. bovis biotype I were related to mannitol and starch hydrolysis-negative S. bovis variant, or biotype, II strains. Another paper on S. bovis DNA-DNA reassociation studies by Coykendall and Gustafson (24) appeared in the same issue of the same journal as Knight and Shlaes’ publication. Coykendall and Gustafson found, like Knight and Shlaes, that typical S. bovis biotype I strains showed homology with some, but not all, of the S. bovis biotype II isolates. Coykendall and Gustafson also examined their strains with the Rapid Strep system (DMS Laboratories, Flemington, NJ), the forerunner of the API 20 Strep Clinical Microbiology Newsletter 29:7,2007
system (bioM´erieux, Hazelwood, MO). With this phenotypic method, the biotype II strains could be subdivided into biotypes II/1 and II/2. Biotype II/2 was identical to biotype I in the hybridization studies, in spite of its phenotypic differences with biotype I in the mannitol fermentation and starch hydrolysis tests (24). A new species resembling S. bovis was proposed in 1995 by Osawa et al. (25), who named the taxon S. gallolyticus because the strains studied were able to decarboxylate gallic acid. This species was proposed on the basis of phenotypic and DNA-DNA reassociation studies. Osawa et al.’s strains were isolated from both animals and humans, and it was suggested on the basis of whole-cell protein analysis studies by Devriese and colleagues (26) that human isolates identified as S. bovis biotypes I and II/2 were members of the new S. gallolyticus species. Conflicting results were published by Clarridge and coworkers (27), who employed 16S rRNA gene sequencing methods. They concluded that human biotype II/2 strains formed a separate species that was different from S. gallolyticus. Poyart and colleagues (28), using superoxide dismutase (sodA) gene sequencing data, proposed that S. bovis biotype II/2 be classified in a separate species, which they named S. pasteurianus. Additional 16S rRNA gene sequencing studies published by Herrero and colleagues (6) suggested that none of the human S. bovis isolates they examined clustered with the S. gallolyticus sequence. The majority of their strains were phenotypically S. bovis I, and their 16S rRNA gene sequences were identical to or differed in one base pair from what was then designated as the type strain of S. bovis (ATCC 43143). Herrero and coworkers also reported that one of their strains was identified as Streptococcus macedonicus, an organism isolated from dairy sources. Soon after the appearance of these two 16S rRNA gene sequencing studies, Schlegel and colleagues (29), employing DNA hybridization methods, advocated the subdivision of S. gallolyticus into three subspecies, S. gallolyticus subsp. gallolyticus (S. bovis biotype I), S. gallolyticus subsp. pasteurianus (S. bovis biotype II/2, formerly called S. Clinical Microbiology Newsletter 29:7,2007
pasteurianus), and S. gallolyticus subsp. macedonicus (isolated from dairy products). One of Clarridge and coworkers’ biotype II/2 strains was included in the study of Schlegel and coworkers and was classified as S. gallolyticus subsp. pasteurianus. The S. bovis type strain (ATCC 43143) examined in Herrero and coworkers’ study was reclassified as S. gallolyticus subsp. gallolyticus when examined in Schlegel’s study. Thus, the new subspecies of S. gallolyticus could accommodate two of the three S. bovis biotypes. The third S. bovis biotype, II/1, contains strains that are beta-glucuronidase negative, in contrast to the biotype II/2 isolates. In 1997, Bouvet and colleagues (30) examined biotype II/1 strains that were unable to hydrolyze esculin and proposed the species name Streptococcus infantarius for this group of organisms. Further investigations by Schlegel and coworkers (31) suggested that S. infantarius should be divided into two subspecies, which would accommodate esculin hydrolysis-variable biotype II/1 strains. S. infantarius subsp. infantarius strains were esculin hydrolysis positive, and S. infantarius subsp. coli strains were variable for esculin hydrolysis. Most recently, Poyart and coworkers (28) proposed that S. infantarius subsp. coli be reclassified as the new species Streptococcus lutetiensis. Most of the taxonomic changes discussed above were chronicled by Facklam in his 2002 review of the evolution of streptococcal classification (32) and are summarized in Table 1. The table contains currently accepted nomenclature (List of Prokaryotic Names with Standing in Nomenclature [http://www.bacterio.cict.fr]), although for ease of comprehension the epithets S. bovis I, II/2, and II/1 will continue to be used in the present discussion.
Implications of Taxonomy for Clinical Microbiologists Early studies that used phenotypic identification methods suggested that the majority of S. bovis human isolates were mannitol-fermenting and starchhydrolyzing biotype I strains that should, in the current classification, be called S. gallolyticus subsp. gallolyticus. Biotype I isolates seemed more likely to be clinically important; they were isolated more often and tended to be more strongly © 2007 Elsevier
associated with endocarditis and gastrointestinal malignancies than biotype II strains (33). Other more recent surveys of have supported the predominance and clinical significance of biotype I isolates (5,6,21). However, Clarridge and coworkers (27) found that biotype II/2 strains were the most common isolates in the collection of S. bovis strains they examined. They noted that their isolates originated from a population composed primarily of adult males, which may have affected the distribution of biotypes. Clinically significant isolates of S. bovis organisms are encountered on a regular basis in the clinical microbiology laboratory, and microbiologists need to be prepared to identify these organisms. If only a few phenotypic tests like reaction on bile esculin medium, presence of Lancefield group D antigen, and growth in 6.5% salt broth, are used, mistakes in identification are likely to occur (34). Commercially available identification systems that rely on a larger number of phenotypic tests are usually fairly accurate at identifying S. bovis isolates. The API 20 Strep system is useful for providing biotypes and has been employed in some of the taxonomic studies of human S. bovis strains. However, no method is perfect, and isolates that are atypical or produce ambiguous identifications should always be expected. All three biotypes of S. bovis (I, II/2 and II/1) have been recovered as significant isolates from cultures of blood and other specimens. Should clinical microbiologists strive to accurately identify these organisms as S. gallolyticus subsp. gallolyticus, S. gallolyticus subsp. pasteurianus, S. infantarius subsp. infantarius, or S. lutetiensis? In a perfect world, the answer would be “yes.” In a perfect world we could do unlimited phenotypic tests and also determine the organism’s 16S rRNA gene sequence. Accurate identification of S. bovis organisms would contribute to a base of knowledge about their incidence, clinical significance, and pathogenic potentials. Unfortunately, for most of us who live in the real world, this would involve an unwarranted expenditure of resources, not to mention all of the explaining we would have to do once the “taxonomically correct” laboratory report was sent out. Current 0196-4399/00 (see frontmatter)
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nomenclature of S. bovis organisms will probably be meaningless to the majority of clinicians. We can realistically provide fairly accurate species and biotype identification with relatively simple methods (e.g., the API 20 Strep system or other phenotypic methods). Biotyping does provide some clinically useful information, even though these designations have no standing in current taxonomy. Table 1 suggests that the biotype designation of an S. bovis isolate can usually predict its current taxonomic standing with fair accuracy. Retaining the S. bovis nomenclature of the previous millennium will not distinguish us as forwardthinking, cutting-edge scientific types, but it might be practical for the present. Eventually the new nomenclature and taxonomy will become better known and accepted by both microbiologists and clinicians. While many studies and observations have been undertaken to explain the clinical significance and taxonomic relationships of S. bovis organisms, much work remains to be done. The intriguing association of S. bovis (and other bacteria) with cancers and what to name the S. bovis isolates we recover in the clinical microbiology laboratory will no doubt occupy many scientists in the future. References 1. Sherman, J.M. 1937. The streptococci. Bacteriol. Rev. 1:3-97. 2. Fauci, A.S. et al. (ed.). 1998. Infective endocarditis, p. 785-791. In Harrison’s principles of internal medicine, 14th ed. McGraw-Hill, New York. 3. Kupferwasser, H.D. et al. 1998. Clinical and morphologic characteristics in Streptococcus bovis endocarditis: a comparison with other causative microorganisms in 177 cases. Heart 80:276-280. 4. Gonzalez-Juanatey, C. et al. 2003. Infective endocarditis due to Streptococcus bovis in a series of nonaddict patients: clinical and morphological characteristics of 20 cases and review of the literature. Can. J. Cardiol. 19:1139-1145. 5. Tripodi, M.F. et al. 2004. Streptococcus bovis endocarditis and its association with chronic liver disease: an underestimated risk factor. Clin. Infect. Dis. 38:1394-1400. 6. Herrero, I.A. et al. 2002. Reevaluation
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of Streptococcus bovis endocarditis cases from 1975 to 1985 by 16S ribosomal DNA sequence analysis. J. Clin. Microbiol. 40:3848-3850. 7. Biarc, J. et al. 2004. Carcinogenic properties of proteins with pro-inflammatory activity from Streptococcus infantarius (formerly S. bovis). Carcinogenesis 25:1477-1484. 8. Beebe, J.L. and E.W. Koneman. 1995. Recovery of uncommon bacteria from blood: association with neoplastic disease. Clin. Microbiol. Rev. 8:347-348. 9. Gold, J.S., S. Bayar, and R.R. Salem. 2004. Association of Streptococcus bovis bacteremia with colonic neoplasia and extracolonic malignancy. Arch. Surg. 139:760-765. 10. Kumar, V., A.K. Abbas, and N. Fausto (ed.). 2005. The gastrointestinal tract, p. 862-865. In Robbins and Cotran pathologic basis of disease, 7th ed. Elsevier, Philadelphia, PA. 11. Parsonett, J. 1995. Bacterial infection as a cause of cancer. Environ. Health Perspect. 103(Suppl. 8):263-268. 12. Leung, W.K. 2006. Helicobacter pylori and gastric neoplasia. Contrib. Microbiol. 13:66-80. 13. Ellmerich, S. et al. 1999. Production of cytokines by monocytes, epithelial and endothelial cells activated by Streptococcus bovis. Cytokine 12:26-31. 14. Ellmerich, S. et al. 2000. Promotion of intestinal carcinogenesis by Streptococcus bovis. Carcinogenesis 21:753-756. 15. Gonzlez-Quintela, A. et al. 2001. Prevalence of liver disease in patients with Streptococcus bovis bacteraemia. J. Infect. 42:116-119. 16. Cohen, L.F. et al. 1997. Streptococcus bovis infection of the central nervous system: report of two cases and review. Clin. Infect. Dis. 25:819-823. 17. Grant, R.J., T.R. Whitehead, and J.E. Orr. 2000. Streptococcus bovis meningitis in an infant. J. Clin. Microbiol. 38:462-463. 18. Vilarrasa, N. et al. 2002. Streptococcus bovis meningitis in a healthy adult patient. Scand. J. Infect. Dis. 34:61-62. 19. Vilaichone, R.K., V. Mahachai, and P. Kullavanijaya. 2002. Spontaneous bacterial peritonitis caused by Streptococcus bovis: case series and review of the literature. Am. J. Gastroenterol. 97:1476-1479. 20. Ruoff, K.L., R.A. Whiley, and D. Beighton. 2003. Streptococcus, p. 405421. In P.R. Murray et al. (ed.), Manual of clinical microbiology, 8th ed. ASM Press, Washington, DC.
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21. Corredoira, J.C. et al. 2005. Clinical characteristics and significance of Streptococcus salivarius bacteremia and Streptococcus bovis bacteremia: a prospective 16-year study. Eur. J. Clin. Microbiol. Infect. Dis. 24:250-255. 22. Farrow, J.A.E. et al. 1984. Taxonomic studies on streptococci of serological groups C, G, and L, and possibly related taxa. Syst. Appl. Microbiol. 5:467-482. 23. Knight, R.G. and D.M. Shlaes. 1985. Physiological characteristics and deoxyribonucleic acid relatedness of human isolates of Streptococcus bovis and Streptococcus bovis (var.). Int. J. Syst. Bacteriol. 35:357-361. 24. Coykendall, A.L. and K.B. Gustafson. 1985. Deoxyribonucleic acid hybridizations among strains of Streptococcus salivarius and Streptococcus bovis. Int. J. Syst. Bacteriol. 35:274-280. 25. Osawa, R., T. Fujisawa, and L.I. Sly. 1995. Streptococcus gallolyticus sp. nov.: gallate degrading organisms formerly assigned to Streptococcus bovis. Syst. Appl. Microbiol. 18:74-78. 26. Devriese, L.A. et al. 1998. Differentiation between Streptococcus gallolyticus strains of human clinical and veterinary origins and Streptococcus bovis strains from the intestinal tracts of ruminants. J. Clin. Microbiol. 36:3520-3523. 27. Clarridge, J.E. et al. 2001. 16S ribosomal DNA sequence analysis distinguishes biotypes of Streptococcus bovis: Streptococcus bovis biotype II/2 is a separate genospecies and the predominant clinical isolate in adult males. J. Clin. Microbiol. 39:1549-1552. 28. Poyart, C., G. Quesne, and P. Trieu-Cuot. 2002. Taxonomic dissection of the Streptococcus bovis group by analysis of manganese-dependent superoxide dismutase gene (sodA) sequences: reclassification of “Streptococcus infantarius subsp. coli” as Streptococcus lutetiensis sp. nov. and of Streptococcus bovis biotype II.2 as Streptococcus pasteurianus sp. nov. Int. J. Syst. Evol. Microbiol. 52:12471255. 29. Schlegel, L. et al. 2003. Reappraisal of the taxonomy of the Streptococcus bovis/Streptococcus equinus complex and related species: description of Streptococcus gallolyticus subsp. gallolyticus subsp. nov., S. gallolyticus subsp. macedonicus subsp. nov. and S. gallolyticus subsp. pasteurianus subsp. nov. Int. J. Syst. Evol. Microbiol. 53:631-645. 30. Bouvet, A. et al. 1997. Streptococcus infantarius sp. nov. related to S. bovis
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and S. equinus. Adv. Exp. Med. Biol. 418:393-395. 31. Schlegel, L. et al. 2000. Streptococcus infantarius subsp. infantarius subsp. nov. and Streptococcus infantarius subsp. coli subsp. nov., isolated from humans and food. Int. J. Syst. Evol.
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Microbiol. 50:1425-1434. 32. Facklam, R. 2002. What happened to the streptococci: overview of taxonomic and nomenclature changes. Clin. Microbiol. Rev. 15:613-630. 33. Ruoff, K.L. et al. 1989. Bacteremia with Streptococcus bovis and Streptococcus
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salivarius: clinical correlates of more accurate identification of isolates. J. Clin. Microbiol. 27:305-308. 34. Ruoff, K.L. et al. 1984. Identification of Streptococcus bovis and Streptococcus salivarius in clinical laboratories. J. Clin. Microbiol. 20:223-226.
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