Demonstration of biofilm in infectious crystalline keratopathy using ruthenium red and electron microscopy

Fulcher, Tim; Dart, John; McLaughlin-Borlace, Louise; Howes, R. C.; Matheson, Melville; Cree, Ian A.

doi:10.1016/s0161-6420(01)00561-9

Cited by 67 publications

(52 citation statements)

References 25 publications

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“…Ruthenium redtreated tissue, counterstained with uranyl acetate and lead citrate, revealed dark fiber-like structures between P. aeruginosa cells in TEM (Fig. 3), which are consistent with previously demonstrated biofilms (10,11). These fibrous structures were not visualized in areas devoid of P. aeruginosa.…”

Section: Resultssupporting

confidence: 89%

“…For visualization by TEM, tissue sections were treated with ruthenium red, a polyanionic stain that stabilizes the structural integrity of the polysaccharide-rich BFM, which can be lost during the dehydration process (10,11). Ruthenium redtreated tissue, counterstained with uranyl acetate and lead citrate, revealed dark fiber-like structures between P. aeruginosa cells in TEM (Fig.…”

Section: Resultsmentioning

confidence: 99%

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Pseudomonas aeruginosa Forms Biofilms in Acute Infection Independent of Cell-to-Cell Signaling

et al. 2007

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Biofilms are bacterial communities residing within a polysaccharide matrix that are associated with persistence and antibiotic resistance in chronic infections. We show that the opportunistic pathogen Pseudomonas aeruginosa forms biofilms within 8 h of infection in thermally injured mice, demonstrating that biofilms contribute to bacterial colonization in acute infections as well. Using light, electron, and confocal scanning laser microscopy, P. aeruginosa biofilms were visualized within burned tissue surrounding blood vessels and adipose cells. Although quorum sensing (QS), a bacterial signaling mechanism, coordinates differentiation of biofilms in vitro, wild-type and QS-deficient P. aeruginosa strains formed similar biofilms in vivo. Our findings demonstrate that P. aeruginosa forms biofilms on specific host tissues independently of QS.Bacterial biofilms are communities of microorganisms residing within a polysaccharide matrix that have been imaged in dental plaques, medical prostheses, and contact lenses (7,31,33). It is well accepted that biofilms play important roles in bacterial persistence and antibiotic resistance in chronic infections, such as cystic fibrosis and otitis media (3,8,9,30). However, the existence and/or roles of biofilms in acute infections, which are defined by short time courses and high severity, have not been examined. The opportunistic gram-negative pathogen Pseudomonas aeruginosa causes both chronic and acute infections and is one of the leading causes of morbidity and mortality in thermally injured patients (27,37). In this study we examined the production of P. aeruginosa biofilms in the thermally injured mouse model of acute infections.The differentiation or maturation of P. aeruginosa biofilms in vitro depends on intercellular signaling systems or quorum sensing (QS) (5,22). QS systems in many gram-negative bacteria rely on acylated homoserine lactones (AHLs), which are produced at high levels when cell density is high and act as ligands for transcriptional regulators. The P. aeruginosa synthases LasI and RhlI synthesize two AHLs, N-3-oxododecanoyl homoserine lactone (3OC 12 -HSL) and N-butyryl-homoserine lactone (C 4 -HSL), which bind and modulate the activity of the transcriptional regulators LasR and RhlR, respectively (28). These transcriptional regulators then regulate the transcription of many genes whose products, including proteases, elastases, toxins, and hemolysins, are thought to be crucial for virulence (28). P. aeruginosa strains lacking functional QS systems are less virulent than wild-type strains (29) and form flat, undifferentiated biofilms on glass surfaces (5). These undifferentiated biofilms are less stable than the differentiated biofilms formed by wild-type P. aeruginosa as they can be easily disrupted by the detergent sodium dodecyl sulfate (5). However, the role of QS in biofilm formation has not previously been examined in vivo. Therefore, in this study we have also examined the role of QS in P. aeruginosa biofilm formation in the acute infection model. ...

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Section: Resultssupporting

confidence: 89%

Section: Resultsmentioning

confidence: 99%

Pseudomonas aeruginosa Forms Biofilms in Acute Infection Independent of Cell-to-Cell Signaling

et al. 2007

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“…The definition given by Donlan and Costerton (16) states that a biofilm is "a microbially derived sessile community characterized by cells that are irreversibly attached to a substratum or interface or to each other, are embedded in a matrix of extracellular polymeric substances that they have produced, and exhibit an altered phenotype with respect to growth rate and gene transcription." For our mycelial mat-like structure, the criteria of attachment is certainly satisfied; furthermore, a thin layer of ruthenium red-stainable substance (possibly exopolysaccharide) (24) coats the filaments (data not shown). The filamentous cells have a growth profile (Fig.…”

Section: Vol 72 2006mentioning

confidence: 86%

Temperature-Regulated Formation of Mycelial Mat-Like Biofilms by Legionella pneumophila

Piao

Sze

Barysheva

et al. 2006

Appl Environ Microbiol

101

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Fifty strains representing 38 species of the genus Legionella were examined for biofilm formation on glass, polystyrene, and polypropylene surfaces in static cultures at 25°C, 37°C, and 42°C. Strains of Legionella pneumophila, the most common causative agent of Legionnaires' disease, were found to have the highest ability to form biofilms among the test strains. The quantity, rate of formation, and adherence stability of L. pneumophila biofilms showed considerable dependence on both temperature and surface material. Glass and polystyrene surfaces gave between two-to sevenfold-higher yields of biofilms at 37°C or 42°C than at 25°C; conversely, polypropylene surface had between 2 to 16 times higher yields at 25°C than at 37°C or 42°C. On glass surfaces, the biofilms were formed faster but attached less stably at 37°C or 42°C than at 25°C. Both scanning electron microscopy and confocal laser scanning microscopy revealed that biofilms formed at 37°C or 42°C were mycelial mat like and were composed of filamentous cells, while at 25°C, cells were rod shaped. Planktonic cells outside of biofilms or in shaken liquid cultures were rod shaped. Notably, the filamentous cells were found to be multinucleate and lacking septa, but a recA null mutant of L. pneumophila was unaffected in its temperature-regulated filamentation within biofilms. Our data also showed that filamentous cells were able to rapidly give rise to a large number of short rods in a fresh liquid culture at 37°C. The possibility of this biofilm to represent a novel strategy by L. pneumophila to compete for proliferation among the environmental microbiota is discussed.

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“…Biofilms adherent to epithelial or artificial surfaces represent the most common form, but intracellular (3) and stromal (8,12) localizations have also been reported. Biofilms attached to ductal walls have been demonstrated in patients with prostatitis (26), but to the best of our knowledge, a stromal biofilm has never been demonstrated in prostate tissue.…”

Section: Discussionmentioning

confidence: 99%

Direct Visualization ofPropionibacterium acnesin Prostate Tissue by Multicolor Fluorescent In Situ Hybridization Assay

Alexeyev¹,

Marklund²,

Shannon³

et al. 2007

J Clin Microbiol

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Prostate tissues from patients with prostate cancer and benign prostatic hyperplasia (BPH) frequently contain histological inflammation, and a proportion of these patients show evidence of Propionibacterium acnes infection in the prostate gland. We developed a multicolor fluorescent in situ hybridization (FISH) assay targeting P. acnes 23S rRNA along with a 14-kb region of the P. acnes genome. This assay was used to analyze prostate tissues from patients with prostate cancer and BPH. P. acnes infection of the prostate gland was demonstrated in prostatic tissue in 5 of 10 randomly selected prostate cancer patients. FISH analysis and confocal laser microscopy imaging revealed intracellular localization and stromal biofilmlike aggregates as common forms of P. acnes infection in prostate tissues from both prostate cancer and BPH patients. A sequential analysis of prostate tissue from individual patients suggested that P. acnes can persist for up to 6 years in the prostate gland. These results indicate that P. acnes can establish a persistent infection in the prostate gland. Further study is needed to clarify the link between this bacterium and prostatic inflammation which may contribute to the development of BPH and prostate cancer.Clinical benign prostatic hyperplasia (BPH) occurs in half of all men by the age of 80 years (20), while prostate cancer is a leading cause of morbidity and death among men in the United States and Western Europe (13). Chronic or recurrent inflammation has been implicated in the development and progression of both BPH (20) and prostate cancer (9,24,25,30), and various degrees of histological inflammation are seen in the majority of prostate tissue specimens examined. The possibility of an infectious etiology for this prostatic inflammation has been raised in the context of both BPH (20) and prostate cancer, although the accumulation of a larger amount of evidence supports this hypothesis for the latter. For example, epidemiological studies have reported an increased risk of prostate cancer in patients with a history of sexually transmitted infections (29,31,32). The genetic signatures of bacteria were found in prostate tissues from 21 of 107 patients with prostate cancer (21). Moreover, a positive correlation between prostatic inflammation and the detection of bacterial 16S rRNA gene sequences in tissue specimens obtained by radical prostatectomy has been reported (16).Recently, a single bacterial species, Propionibacterium acnes, has been shown to infect a considerable proportion (35%) of prostate glands removed at radical prostatectomy (6). Moreover, the presence of P. acnes was strongly correlated with histological inflammation, suggesting that this bacterium might be linked to cancer development. The possible involvement of P. acnes in prostate pathology was further highlighted by the detection of P. acnes 16S rRNA gene sequences in BPH tissues, with a positive association between the detection of P. acnes DNA and subsequent prostate cancer development reported (1). We have developed ...

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Demonstration of biofilm in infectious crystalline keratopathy using ruthenium red and electron microscopy

Cited by 67 publications

References 25 publications

Pseudomonas aeruginosa Forms Biofilms in Acute Infection Independent of Cell-to-Cell Signaling

Pseudomonas aeruginosa Forms Biofilms in Acute Infection Independent of Cell-to-Cell Signaling

Temperature-Regulated Formation of Mycelial Mat-Like Biofilms by Legionella pneumophila

Direct Visualization ofPropionibacterium acnesin Prostate Tissue by Multicolor Fluorescent In Situ Hybridization Assay

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