Culture of Small Colony Variant of &lt;em&gt;Pseudomonas aeruginosa&lt;/em&gt; and Quantitation of its Alginate

Ahmar, Roy Al; Kirby, Brandon D.; Yu, Hongwei D.

doi:10.3791/60466

Cited by 5 publications

(7 citation statements)

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“…This provides an additional explanation for survivor P. putida mucoidy since alginate lyases are responsible for the degradation of alginate 49 . Utilizing an established carbazole assay as well as alginate antibody dot blots 50 – 52 , we sought to determine if the increased transcription of genes associated with mucoid conversion observed from survivor P. putida resulted in increased production of alginate when compared to parent P. putida . Comparing supernatants, we observed that survivor P. putida indeed produced more alginate when compared to parent P. putida (Fig.…”

Section: Resultsmentioning

confidence: 99%

Functional genomics study of Pseudomonas putida to determine traits associated with avoidance of a myxobacterial predator

Akbar

Stevens

2021

Sci Rep

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Predation contributes to the structure and diversity of microbial communities. Predatory myxobacteria are ubiquitous to a variety of microbial habitats and capably consume a broad diversity of microbial prey. Predator–prey experiments utilizing myxobacteria have provided details into predatory mechanisms and features that facilitate consumption of prey. However, prey resistance to myxobacterial predation remains underexplored, and prey resistances have been observed exclusively from predator–prey experiments that included the model myxobacterium Myxococcus xanthus. Utilizing a predator–prey pairing that instead included the myxobacterium, Cystobacter ferrugineus, with Pseudomonas putida as prey, we observed surviving phenotypes capable of eluding predation. Comparative transcriptomics between P. putida unexposed to C. ferrugineus and the survivor phenotype suggested that increased expression of efflux pumps, genes associated with mucoid conversion, and various membrane features contribute to predator avoidance. Unique features observed from the survivor phenotype when compared to the parent P. putida include small colony variation, efflux-mediated antibiotic resistance, phenazine-1-carboxylic acid production, and increased mucoid conversion. These results demonstrate the utility of myxobacterial predator–prey models and provide insight into prey resistances in response to predatory stress that might contribute to the phenotypic diversity and structure of bacterial communities.

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Section: Resultsmentioning

confidence: 99%

Functional genomics study of Pseudomonas putida to determine traits associated with avoidance of a myxobacterial predator

Akbar

Stevens

2021

Sci Rep

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“…This modification of metabolic activities can limit effective detection and treatment of infecting P. aeruginosa , as illustrated by the emergence of highly resistant small colony variants (SCVs) and viable but non-culturable (VBNC) isolates [115–117].…”

Section: Phenotypical Signatures Of Cf-adapted P Aeruginosamentioning

confidence: 99%

“…This adjusted metabolism increases P. aeruginosa fitness in the CF environment, but it often results in a slowed growth in laboratory conditions in comparison to nonadapted isolates [7,8,12,13,18,107,110,114]. This modification of metabolic activities can limit effective detection and treatment of infecting P. aeruginosa, as illustrated by the emergence of highly resistant small colony variants (SCVs) and viable but non-culturable (VBNC) isolates [115][116][117].…”

Section: Metabolic Alterationsmentioning

confidence: 99%

From genotype to phenotype: adaptations of Pseudomonas aeruginosa to the cystic fibrosis environment

2021

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Pseudomonas aeruginosa is one of the main microbial species colonizing the lungs of cystic fibrosis patients and is responsible for the decline in respiratory function. Despite the hostile pulmonary environment, P. aeruginosa is able to establish chronic infections thanks to its strong adaptive capacity. Various longitudinal studies have attempted to compare the strains of early infection with the adapted strains of chronic infection. Thanks to new ‘-omics’ techniques, convergent genetic mutations, as well as transcriptomic and proteomic dysregulations have been identified. As a consequence of this evolution, the adapted strains of P. aeruginosa have particular phenotypes that promote persistent infection.

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“…Alginate-based formation of mucoid biofilms has previously been associated with P. aeruginosa avoidance of the protozoan grazer Rhynchomonas nasuta (Matz et al, 2002; Matz et al, 2004; Matz and Kjelleberg, 2005; Matz et al, 2005). Utilizing established carbazole assays on isolated alginate (Knutson and Jeanes, 1968; Al Ahmar et al, 2020), we sought to determine if the increased transcription of genes known to impact alginate production observed from the survivor phenotype resulted in increased production of alginate when compared to predator unexposed P. putida. Comparing isolated alginate from predation sensitive P. putida with the survivor phenotype, we observed that the survivor phenotype indeed produced more alginate when compared to P. putida not exposed to C. ferrugineus (Figure 4).…”

Section: Resultsmentioning

confidence: 99%

“…Precipitated alginate was suspended in 150 μl 0.9% (w/v) saline solution. Following established protocols (Knutson and Jeanes, 1968; Al Ahmar et al, 2020), a 50 μL aliquot of the isolated alginate and a dilution series of standard alginic acid (Sigma) were mixed with 200 μL of a solution of 25 mM sodium tetraborate in sulfuric acid and added in a 96-well plate. Next, the plate was heated for 10 min at 100 °C in an oven.…”

Section: Methodsmentioning

confidence: 99%

Predatory selection of mucoid, antibiotic resistantPseudomonas putidaphenotype by myxobacteriumCystobacter ferrugineus

Akbar

Stevens

2020

Preprint

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Predation contributes to the structure and diversity of microbial communities. Predatory myxobacteria are ubiquitous to a variety of microbial habitats and capably consume a broad diversity of microbial prey. Predator-prey experiments utilizing myxobacteria have provided details into predatory mechanisms and features that facilitate consumption of prey. However, prey resistance to myxobacterial predation remains underexplored, and prey resistances have been observed exclusively from predator-prey experiments that included the model myxobacterium Myxococcus xanthus. Utilizing a predator-prey pairing that instead included the myxobacterium, Cystobacter ferrugineus, with Pseudomonas putida as prey, we infrequently observed surviving phenotypes capable of eluding predation. Comparative transcriptomics between P. putida unexposed to C. ferrugineus and the survivor phenotype suggested that increased expression of efflux pumps, genes associated with mucoid conversion, and various membrane features contribute to predator avoidance. The P. putida survivor phenotype was confirmed to be resistant to the antibiotics kanamycin, gentamicin, and tetracycline and to produce more alginate than predator-unexposed P. putida. Unique features observed from the survivor phenotype including small colony variation, efflux-mediated antibiotic resistance, and increased mucoid conversion overlap with traits associated with Pseudomonas aeruginosa predator avoidance and pathogenicity. The survivor phenotype also benefited from increased predator resistance during subsequent predation assays. These results demonstrate the utility of myxobacterial predator-prey models and provide insight into prey resistances in response to predatory stress might contribute to the phenotypic diversity and structure of bacterial communities.

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Culture of Small Colony Variant of <em>Pseudomonas aeruginosa</em> and Quantitation of its Alginate

Cited by 5 publications

References 0 publications

Functional genomics study of Pseudomonas putida to determine traits associated with avoidance of a myxobacterial predator

Functional genomics study of Pseudomonas putida to determine traits associated with avoidance of a myxobacterial predator

From genotype to phenotype: adaptations of Pseudomonas aeruginosa to the cystic fibrosis environment

Predatory selection of mucoid, antibiotic resistantPseudomonas putidaphenotype by myxobacteriumCystobacter ferrugineus

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