Streptococcus bovis Contributes to the Development of Colorectal Cancer via Recruiting CD11b⁺TLR-4⁺ Cells.

BACKGROUND An increasing number of studies have demonstrated that Streptococcus bovis and its concomitant inflammatory factors concentrate in the intestine in colorectal cancer (CRC). However, the molecular mechanism of S. bovis on colorectal tumorigenesis remains unclear. This study aimed to explore the role of S. bovis in carcinogenesis and its potential mechanism in CRC of mice orally pretreated with S. bovis. MATERIAL AND METHODS The colons of experimental mice were collected and evaluated for the extent of neoplasm. In addition, comparative feces DNA sequencing was adopted to verify the abundance change of S. bovis during the progression of CRC in patients. RESULTS The results of this study found that S. bovis is more likely to be present at higher levels in patients with progressive colorectal carcinoma compared to those adenoma patients and healthy volunteers (P<0.05). Pretreatment with S. bovis aggravated tumor formation in mice, resulting in more substantial and a higher number of tumor nodes (P<0.05). A cytokine expression pattern with increased levels of IL-6, Scyb1, Ptgs2, IL-1ß, TNF, and Ccl2 was detected in S. bovis pretreated CRC mice (all P<0.05). Furthermore, S. bovis recruited myeloid cells, especially CD11b⁺TLR-4⁺ cells, which could promote pro-tumor immunity in the tumor microenvironment (P<0.05). CONCLUSIONS Collectively, our study indicates that S. bovis may induce a suppressive immunity that is conducive to CRC by recruiting tumor-infiltrating CD11b⁺TLR-4⁺ cells. In conclusion, S. bovis contributes to colorectal tumorigenesis via recruiting CD11b⁺TLR-4⁺ cells.

[Presence of Streptococcus bovis in urine samples from patients experiencing symptoms of urinary tract]. / Presencia de Streptococcus bovis en muestras de orina de sujetos con síntomas de infección del aparato urinario.

Given the relevance of proper clinical validation of Streptococcus bovis, we here consider revising its presence in urine samples in order to determine its relative frequency and the pattern of antibiotic susceptibility. The susceptibility to antibiotics of 91 isolates of S. bovis from urine samples was retrospectively reviewed over a period of 4 years (2012-2015). The mean age of patients was 55 years, 81% of whom were women and 37.4% were hospitalized patients suffering from urological diseases (61%). Susceptibility to penicillin, vancomycin and teicoplanin was 97.8%. Due to the fact that S. bovis can be infrequent in urine isolates and given its presence in patients suffering from urological diseases, further pathogenic studies, showing the true ability of this group of bacteria to produce disease, are required.

[Streptococcus gallolyticus (ex S. bovis) bacteremia and its relationship with colonic or hepatobiliary disease and endocarditis]. / Bacteriemias por Streptococcus gallolyticus (ex S. bovis) y su relación con patología colónica o hepatobiliar y endocarditis.

BACKGROUND: Bacteremia due to Streptococcus bovis (now S. gallolyticus) has been traditionally associated to colon or hepatobiliar disease and endocarditis but there is no information on this matter in Chile. AIMS: To describe clinical features of adult patients suffering bacteremia by S. bovis/S. gallolyticus, identify the source of the bacteremia and the frequency of endocarditis. METHODS: Retrospective-descriptive study using laboratory records. RESULTS: Between January 2003 and August 2014, 23 S. bovis/S. gallolyticus bacteremic events were identified among 22 patients. Mean age was 72.7 years (range 46-96). Co-morbidities were frequent (9.1 to 47.6%). The primary source of bacteremia was intestinal in 52.2%; hepatobiliar in 17.4% and in 34.8% it was not elucidated. Six patients had infective endocarditis (26.1%) and one patient had espondylodiscitis (4.3%). S. bovis represented 39.1% of isolates (all until 2008), S. gallolyticus subsp pasteurianus 39.1% and, S. gallolyticus subsp infantarius and S. gallolyticus subsp gallolyticus 8.7% each one, respectively. Association studies between the bacteremic source or endocarditis with specific S. gallolyticus subspecies were limited by the small number of isolates. Seven patients (30.4%) underwent surgical interventions. In-hospital mortality reached 21.7% (n=5). CONCLUSIONS: Although infrequent, bacteremic events by S. gallolyticus/S. bovis have increased in-hospital mortality, require surgical intervention and affect older patients with co-morbidities. Near two-thirds suffer from colonic or hepatobiliary disease that act as the primary source of bacteremia. In addition, near one fourth is affected by infective endocarditis. Detection of S. gallolyticus/S. bovis in blood cultures prompts a thorough clinical evaluation in order to clarify the source of the bloodstream infection and the presence of complications.

[Diseases associated with bloodstream infections caused by the new species included in the old Streptococcus bovis group]. / Cuadros clínicos asociados a bacteriemia causada por las nuevas especies incluidas en el antiguo grupo Streptococcus bovis.

OBJECTIVE: We sought to identify possible diseases associated with bloodstream infections caused by new species of S. bovis group isolated in blood cultures and by studying patient records METHODS: Forty-four consecutive blood culture isolates initially designated S. bovis were further characterised using phenotypic methods Patient records were examined. RESULTS: We identified 15 Streptococcus gallolyticus subsp. gallolyticus, 24 Streptococcus gallolyticus subsp. pasteurianus, and 5 Streptococcus infantarius isolates in 44 BSI episodes. CONCLUSIONS: The association between S. bovis bacteraemia and endocarditis and/or colon carcinoma is highly dependent on the causative species. Streptococcus gallolyticus subsp. gallolyticus is a surrogate for endocarditis and/or bowel disease, whereas Streptococcus gallolyticus subsp. pasteurianus is a surrogate for hepato-biliary disease.

Clinical Importance of Streptococcus gallolyticus infection among colorectal cancer patients: systematic review and meta-analysis.

BACKGROUND: Streptococcus bovis has long been associated with colorectal cancer (CRC). However, not all genospecies are as closely related to CRC. With this systematic review, we aim to increase the awareness of the association between S. bovis biotype I (Streptococcus gallolyticus) and CRC and urge for uniform molecular microbiological classification. METHODS: In January 2011, the PubMed database was searched for all studies that investigated the association between S. bovis, infective endocarditis (IE), and CRC. A total of 191 studies were screened for eligibility and yielded 52 case reports and 31 case series, of which 11 were used for meta-analysis on the association between S. bovis biotype, IE, and adenomas/carcinomas (CRC). RESULTS: Among the S. bovis-infected patients who underwent colonic evaluation, the median percentage of patients who had concomitant adenomas/carcinomas was 60% (interquartile range, 22%), which largely exceeds the disease rate reported in the general asymptomatic population. Meta-analysis showed that patients with S. bovis biotype I infection had a strongly increased risk of having CRC (pooled odds ratio [OR], 7.26; 95% confidence interval [CI], 3.94-13.36) and IE (pooled OR, 16.61; 95% CI, 8.85-31.16), compared with S. bovis biotype II-infected patients. Notably, CRC occurred more often among patients with S. bovis IE than among patients with S. bovis infection at other sites (pooled OR, 3.72; 95% CI, 2.03-6.81). CONCLUSIONS: Our meta-analysis clearly indicates that S. bovis should no longer be regarded as a single species in clinical practice, because S. gallolyticus (S. bovis biotype I) infection, in particular, has an unambiguous association with CRC.

Identification and pathogenicity analysis of Streptococcus equinus FMD1, a beta-hemolytic strain isolated from forest musk deer lung.

Streptococcus spp. cause a wide range of diseases in animals and humans. A Streptococcus strain (FMD1) was isolated from forest musk deer lung. To identify the bacterium at the species level and investigate its pathogenicity, whole genome sequencing and experimental infections of mice were performed. The genome had 97.63% average nucleotide identity with the S. equinus strain. Through virulence gene analysis, a beta-hemolysin/cytolysin genome island was found in the FMD1 genome, which contained 12 beta-hemolysin/cytolysin-related genes. Hemolytic reaction and histopathological analysis established the strain's pathogenicity in mice. This is the first report of a beta-hemolytic S. equinus strain in forest musk deer identified based on phenotypic and genotypic analyzes; this strategy could be useful for analyzing pathogens affecting rare animals.

Surface-exposed histone-like protein a modulates adherence of Streptococcus gallolyticus to colon adenocarcinoma cells.

Streptococcus gallolyticus (formerly known as Streptococcus bovis biotype I) is a low-grade opportunistic pathogen which is considered to be associated with colon cancer. It is thought that colon polyps or tumors are the main portal of entry for this bacterium and that heparan sulfate proteoglycans (HSPGs) at the colon tumor cell surface are involved in bacterial adherence during the first stages of infection. In this study, we have shown that the histone-like protein A (HlpA) of S. gallolyticus is a genuine anchorless bacterial surface protein that binds to lipoteichoic acid (LTA) of the gram-positive cell wall in a growth phase-dependent manner. In addition, HlpA was shown to be one of the major heparin-binding proteins of S. gallolyticus able to bind to the HSPG-expressing colon tumor cell lines HCT116 and HT-29. Strikingly, although wild-type levels of HlpA appeared to contribute to adherence, coating of additional HlpA at the bacterial surface resulted in reduced binding to colon tumor cells. This may be explained by the fact that heparan sulfate and LTA compete for the same binding site in HlpA. Altogether, this study implies that HlpA serves as a fine-tuning factor for bacterial adherence.

Ecology and pathogenicity of gastrointestinal Streptococcus bovis.

Streptococcus bovis is an indigenous resident in the gastrointestinal tracts of both humans and animals. S. bovis is one of the major causes of bacterial endocarditis and has been implicated in the incidence of human colon cancer, possibly due to chronic inflammatory response at the site of intestinal colonization. Certain feeding regimens in ruminants can lead to overgrowth of S. bovis in the rumen, resulting in the over-production of lactate and capsular polysaccharide causing acute ruminal acidosis and bloat, respectively. There are multiple strategies in controlling acute lactic acidosis and bloat. The incidence of the two diseases may be controlled by strict dietary management. Gradual introduction of grain-based diets and the feeding of coarsely chopped roughage decrease the incidence of the two disease entities. Ionophores, which have been used to enhance feed conversion and growth rate in cattle, have been shown to inhibit the growth of lactic acid bacteria in the rumen. Other methods of controlling lactic acid bacteria in the ruminal environment (dietary supplementation of long-chain fatty acids, induction of passive and active immune responses to the bacteria, and the use of lytic bacteriophages) have also been investigated. It is anticipated that through continued in-depth ecological analysis of S. bovis the characteristics responsible for human and animal pathogenesis would be sufficiently identified to a point where more effective control strategies for the control of this bacteria can be developed.

Infectious agents and colorectal cancer: a review of Helicobacter pylori, Streptococcus bovis, JC virus, and human papillomavirus.

Based on the high volume of bacteria and viruses that the intestine is exposed to and the importance of infectious agents in some gastrointestinal and anogenital cancers, it is not surprising the many studies have evaluated the association between colorectal cancer and infectious agents. This review highlights investigations of four agents in relation to colorectal cancer. Helicobacter pylori, Streptococcus bovis, JC virus, and human papillomavirus have all been evaluated as possible etiologic agents for colorectal cancer. For each of these agents, a review of possible mechanisms for carcinogenesis and epidemiologic evidence is discussed, and future directions for research are proposed.

Adherence characteristics of endocarditis-derived Streptococcus gallolyticus ssp. gallolyticus (Streptococcus bovis biotype I) isolates to host extracellular matrix proteins.

Members of the Streptococcus bovis group are frequent colonizers of the intestinal tract, which can also cause endocarditis. However, their ability to adhere to and colonize host tissues and the factors associated with pathogenicity are largely unknown. Here, we assessed 17 endocarditis-derived human isolates [identified here as 15 Streptococcus gallolyticus ssp. gallolyticus (S. bovis biotype I), one S. gallolyticus ssp. pasteurianus (biotype II/2) and one Streptococcus infantarius ssp. coli (biotype II/1)] for their in vitro adherence to components of the extracellular matrix (ECM). Adherence to collagen type I was found to be the most common phenotype exhibited by 76% of isolates, followed by collagen type IV (53%), fibrinogen (47%), collagen type V (35%) and fibronectin (35%). Pulsed-field gel electrophoresis analyses showed that >50% of endocarditis-derived S. gallolyticus ssp. gallolyticus isolates are genetically diverse, although two clusters of two and four isolates were observed. The diversity of strains and differences observed in adherence characteristics to distinct host ECM proteins suggest that isolates of S. gallolyticus ssp. gallolyticus produce different surface components, similar to other gram-positive pathogens, to colonize the host and cause infection.

Characteristics and regional variations of group D streptococcal endocarditis in France.

The proportion of infective endocarditis (IE) caused by group D streptococci (GDS; formerly Streptococcus bovis) increased markedly in France, to account for 25% of all cases of IE by 1999. In an attempt to explain this phenomenon, a comparative analysis of GDS and oral streptococci (OS) causing IE was performed. This study was based on data collected from a large cross-sectional population-based survey that was conducted in 1999. In total, 559 cases of definite IE were recorded, of which 142 involved GDS and 79 involved OS. Patients with GDS IE were older (62.7 vs. 56.6 years, p 0.01) and had a history of valve disease less frequently than did patients with OS IE (33.8% vs. 67.1%, p <0.0001). At-risk procedures for IE were performed less frequently in patients with GDS than in patients with OS (14.8% vs. 24.1%, p 0.08), but co-morbidities were more frequent in the GDS group (59.9% vs. 32.9%, p 0.0001). Diabetes, colon diseases and cirrhosis were also more frequent in the GDS group (p 0.006, p <0.0001 and p 0.08, respectively). Rural residents accounted for 31.0% of the GDS group, but for only 15.2% of the OS group (p 0.001). Likewise, the proportion of GDS IE was higher in regions with mixed (urban and rural) populations (Franche-Comté 81.8%, Marne 68.7%, Lorraine 70.3% and Rhône-Alpes 65.3%) than in exclusively urban regions (Paris and Ile de France 58.0%). Further investigations are required to elucidate the link in France between the incidence of GDS IE, rural residency and nutritional factors.

Cloning and characterization of the glutamate dehydrogenase gene in Streptococcus bovis.

Streptococcus bovis, an etiologic agent of rumen acidosis in cattle, is a rumen bacterium that can grow in a chemically defined medium containing ammonia as a sole source of nitrogen. To understand its ability to assimilate inorganic ammonia, we focused on the function of glutamate dehydrogenase. In order to identify the gene encoding this enzyme, we first amplified an internal region of the gene by using degenerate primers corresponding to hexameric family I and NAD(P)+ binding motifs. Subsequently, inverse PCR was used to identify the whole gene, comprising an open reading frame of 1350 bp that encodes 449 amino acid residues that appear to have the substrate binding site of glutamate dehydrogenase observed in other organisms. Upon introduction of a recombinant plasmid harboring the gene into an Escherichia coli glutamate auxotroph lacking glutamate dehydrogenase and glutamate synthase, the transformants gained the ability to grow on minimal medium without glutamate supplementation. When cell extracts of the transformant were resolved by blue native polyacrylamide gel electrophoresis followed by activity staining, a single protein band appeared that corresponded to the size of S. bovis glutamate dehydrogenase. Based on these results, we concluded that the gene obtained encodes glutamate dehydrogenase in S. bovis.

A prospective controlled study of the association of Streptococcus bovis with colorectal carcinoma.

AIM: To investigate the ability of Streptococcus bovis to colonise colorectal cancers. PATIENTS: 19 patients with colorectal cancer and 23 controls without malignancy. SETTING: University teaching hospital. METHODS: Prospective study comparing unselected patients with known colorectal cancer with age and sex matched controls. Carcinoma tissue from patients with colorectal cancer and normal colonic mucosa, stool, and blood from both patients and control subjects were cultured. RESULTS: In contrast to published data, the faecal carriage rate was similar in cancer (11%) and control groups (13%). CONCLUSIONS: Faecal colonisation by Str bovis in colorectal cancer patients is lower than previously reported and does not differ significantly from controls.

Streptococcus bovis infections in pigeons: virulence of different serotypes.

In a first experiment, the relative virulence for pigeons of 5 strains of S. bovis was assessed by experimental inoculations. Two S. bovis serotype 1 strains, one serotype 2 strain and two serotype 3 strains were examined. One of the serotype 1 strains and the serotype 2 strain were isolated from pigeons that died from septicaemia. The other strains were isolated from cloaca samples of healthy pigeons. For each strain, 10-20 pigeons were intravenously inoculated with 1 x 10(9) CFU. Morbidity after infection with the serotype 1 and 2 strains varied between 75% and 90%. Disease signs included inability to fly, lameness, emaciation, production of slimy, green droppings, polyuria and sudden death. In groups of pigeons inoculated with the serotype 3 strains, morbidity was 0% and 6%, respectively. Results demonstrate that serotype 3 strains are less virulent for pigeons than serotype 1 and 2 strains. In a second experiment, bacteriological and histological examinations were performed on organs of pigeons serially killed between 1 and 10 days after experimental inoculation with an S. bovis serotype 3 strain of low virulence. Results were compared with results of studies carried out with a highly virulent serotype 1 strain. Notwithstanding bacterial spread and replication in various organs of inoculated pigeons, clinical disease was not observed and histological lesions were scarce and of limited extent.

Extracellular proteins and virulence in Streptococcus bovis isolates from pigeons.

The association between virulence and the occurrence of the extracellular proteins A, T1, T2 and T3 in the culture supernatant of pigeon Streptococcus bovis strains, was examined in experimental infection studies. Fourteen groups of 10-17 pigeons were inoculated intravenously with 1 x 10(9) CFU of S. bovis strains that belonged to the phenotypes A + T1, A - T1, A + T2, A - T2, A + T3 and A - T3, respectively. The overall postinoculation morbidity in the phenotype groups was 85%, 87%, 70%, 5%, 100% and 37%, respectively. These results indicate that strains producing A or T1 are of high virulence, those producing T3 only are of moderate virulence and those producing T2 are of low virulence. Virulence of S. bovis for pigeons was more clearly correlated with supernatant-phenotype than with serotype.

Immunity in pigeons against homologous and heterologous serotypes of Streptococcus bovis after infection.

Groups of 20, 20 and 10 pigeons were intravenously inoculated with 1 x 10(9) CFU of a Streptococcus bovis serotype 1, 2 and 3 strain, respectively. Groups which received the highly virulent serotypes 1 or 2 strains were treated with antibiotics starting from 2 days post inoculation in order to prevent the development of clinical signs. Pigeons inoculated with the low virulence S. bovis serotype 3 strain were left untreated. Another group of 10 non-inoculated pigeons was used as challenge control. Four weeks later, pigeons were challenged intravenously with S. bovis serotype 1. Morbidity after challenge infection was 19%, 36% and 70% in groups previously inoculated with serotypes 1, 2 and 3, respectively, whereas it was 100% in the control group. Results demonstrate that pigeons developed significant protective immunity to S. bovis serotype 1 septicaemia following infection with S. bovis serotype 1 and 2 strains but not after infection with the serotype 3 strain. Protection did not correlate with the presence of antibodies as detected in ELISA.

Secreted antigens as virulence associated markers in Streptococcus bovis strains from pigeons.

SDS-PAGE and Western blot analysis were performed on the culture supernatant of 7 pigeon S. bovis reference strains belonging to the serotypes 1, 2, 3 and 5. The culture supernatant of highly virulent serotype 1, 2 and 5 strains contained four antigens that were absent in low virulent serotype 3 strains, notably a 185 kDa minor protein band (A) and a triplet (T1) of 70 kDa. The less virulent serotype 3 strains on the other hand contained protein triplets, that had a molecular mass of either 68 kDa (T2) or 74 kDa (T3). The prevalence of A, T1, T2 and T3 was examined in 68 S. bovis strains isolated from healthy pigeons and in 68 S. bovis strains isolated from pigeons that died of S. bovis septicaemia. Six supernatant phenotypes were identified: A-T1 (32 strains), A- T2 (10 strains), A- T3 (7 strains), A+ T1 (84 strains), A+ T2 (1 strain) and A+ T3 (2 strains). Ninety-four percent of the strains lacking the A and T1 proteins were isolated from healthy pigeons, and only 6% were isolated from septicaemia. Strains expressing A and/or T1, however, were isolated form septicaemia in 57% of the cases. These observations may indicate that the A and/or T1 proteins are associated with virulence.

Intracellular survival and multiplication of virulent and less virulent strains of Streptococcus bovis in pigeon macrophages.

The intracellular fate of pigeon S. bovis strains ingested by macrophages was studied in vivo and in vitro. During in vivo experiments, histological and electron microscopical examinations demonstrated numerous cocci, which appeared to be actively multiplying, within splenic macrophages of pigeons experimentally inoculated with a highly virulent S. bovis serotype 1 strain. In pigeons inoculated with a low virulence serotype 3 strain, intracellular cocci were only occasionally observed. For in vitro experiments, pigeon peritoneal macrophages were inoculated with a S. bovis serotype 1 or serotype 3 strain and incubated. Following an initial decrease, an increase in the number of intracellular bacteria was observed in tests performed with the S. bovis serotype 1 strain, demonstrating intracellular multiplication. Macrophages in these experiments had all died after 7 h of incubation, possibly indicating that the intracellular replication of S. bovis resulted in the release of substances toxic for macrophages. In experiments performed with the S. bovis serotype 3 strain, the number of intracellular bacteria continuously decreased, reflecting killing of organisms. Significant changes in the number of adhering macrophages in S. bovis serotype 3 inoculated cultures were not observed. These results indicate S. bovis in pigeons is a facultative intracellular bacterium and intracellular multiplication may be involved in virulence.

In vitro evidence for a bacterial pathogenesis of equine laminitis.

Utilizing an in vitro laminitis explant model, we have investigated how bacterial broth cultures and purified bacterial proteases activate matrix metalloproteinases (MMPs) and alter structural integrity of cultured equine lamellar hoof explants. Four Gram-positive Streptococcus spp. and three Gram-negative bacteria all induced a dose-dependent activation of MMP-2 and MMP-9 and caused lamellar explants to separate. MMP activation was deemed to have occurred if a specific MMP inhibitor, batimastat, blocked MMP activity and prevented lamellar separation. Thermolysin and streptococcal pyrogenic exotoxin B (SpeB) both separated explants dose-dependently but only thermolysin was inhibitable by batimastat or induced MMP activation equivalent to that seen with bacterial broths. Additionally, thermolysin and broth MMP activation appeared to be cell dependent as MMP activation did not occur in isolation. These results suggest the rapid increase in streptococcal species in the caecum and colon observed in parallel with carbohydrate induced equine laminitis may directly cause laminitis via production of exotoxin(s) capable of activating resident MMPs within the lamellar structure. Once activated, these MMPs can degrade key components of the basement membrane (BM) hemidesmosome complex, ultimately separating the BM from the epidermal basal cells resulting in the characteristic laminitis histopathology of hoof lamellae. While many different causative agents have been evaluated in the past, the results of this study provide a unifying aetiological mechanism for the development of carbohydrate induced equine laminitis.

Identification of virulence associated markers in the cell wall of pigeon Streptococcus gallolyticus strains.

The cell wall protein profiles of 56 isolates of Streptococcus gallolyticus of differing virulence for pigeons were compared by SDS-PAGE. Additionally, Western blot analysis was performed on the cell wall proteins of 14 strains using sera of pigeons, experimentally infected with A(+)T1 or A(-)T2 strains of S. gallolyticus. The profile of silver stained gels exhibited a complex array of 20-50 bands ranging from less than 6.5-210kDa. A band with molecular mass of 114kDa was only observed in isolates that belonged to the highly virulent A(+)T1, A(+)T2, A(+)T3 and A(-)T1 culture supernatant groups. A band with a slightly higher molecular mass (115kDa) as well as a 207kDa band were only detected in isolates that belonged to the moderately A(-)T3 or low A(-)T2 virulent culture supernatant groups. The 114 and 115kDa band were recognised by all homologous and heterologous pigeon sera used whereas the 207kDa band was only recognised by sera of pigeons infected with a A(-)T2 strain. These findings may indicate that the 114, 115 and 207kDa bands are useful as additional virulence associated markers for pigeon S. gallolyticus strains.