The Matrix Revisited: Opening Night for the Pel Polysaccharide Across Eubacterial Kingdoms

Bacteria synthesize and export adhesive macromolecules to enable biofilm formation. These macromolecules, collectively called the biofilm matrix, are structurally varied and often unique to specific bacterial species or subspecies. This heterogeneity in matrix utilization makes it difficult to facilitate direct comparison between biofilm formation mechanisms of different bacterial species. Despite this, some matrix components, in particular the polysaccharides poly-β-1,6- N-acetyl-glucosamine (PNAG) and bacterial cellulose, are utilized by many Gram-negative species for biofilm formation. However, there is a very narrow distribution of these components across Gram-positive organisms, whose biofilm matrix determinants remain largely undiscovered. We found that a genetic locus required for the production of a biofilm matrix component of P. aeruginosa, the Pel polysaccharide, is widespread in Gram-negative bacteria and that there is a variant form of this cluster present in many Gram-positive bacterial species. We demonstrated that this locus is required for biofilm formation by Bacillus cereus ATCC 10987, produces a polysaccharide that is similar to Pel, and is post-translationally regulated by cyclic-3′,5′-dimeric-guanosine monophosphate (c-di-GMP) in a manner identical to P. aeruginosa. However, while the proposed mechanism for Pel production appears remarkably similar between B. cereus and P. aeruginosa, we identified several key differences between Gram-negative and Gram-positive Pel biosynthetic components in other monoderms. In particular, 4 different architectural subtypes of the c-di-GMP-binding component PelD were identified, including 1 found only in Streptococci that has entirely lost the c-di-GMP recognition domain. These observations highlight how existing multi-component bacterial machines can be subtly tweaked to adapt to the unique physiology and regulatory mechanisms of Gram-positive organisms. Collectively, our analyses suggest that the Pel biosynthetic locus is one of the most phylogenetically widespread biofilm matrix determinants in bacteria, and that its mechanism of production and regulation is extraordinarily conserved across the majority of organisms that possess it.

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Enzymes from carbohydrase group destroy biofilm matrix of gram-positive and gram-negative bacteria

Medical alphabet ◽

10.33667/2078-5631-2019-4-34(409)-40-45 ◽

2020 ◽

Vol 4 (34) ◽

pp. 40-45

Author(s):

Yu. M. Romanova ◽

A. V. Tutelyan ◽

A. P. Sinitsyn ◽

V. M. Pisarev ◽

N. V. Alekseeva ◽

...

Keyword(s):

Exposure Time ◽

Gram Negative Bacteria ◽

Gram Positive ◽

Gram Negative ◽

Abiotic Surfaces ◽

The Matrix ◽

Biofilm Matrix

The effect of enzymes carbohydrases exopolysaccharide matrix of biofilms formed by gram-positive and gram-negative bacteria on abiotic surfaces was studied. The ability of a mixture of carbohydrase enzymes (hydrolases and lyases) to completely destroy the matrix of biofilms formed by gram-positive and gram-negative bacteria causing health-associated infections has been confirmed. An optimal mixture of carbohydrases possesed high anti-biofilm activity even when employed in relatively small concentrations and at a brief exposure time.

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Evaluation of MicroScan Bacterial Identification Panels for Low-Resource Settings

Diagnostics ◽

10.3390/diagnostics11020349 ◽

2021 ◽

Vol 11 (2) ◽

pp. 349

Author(s):

Sien Ombelet ◽

Alessandra Natale ◽

Jean-Baptiste Ronat ◽

Olivier Vandenberg ◽

Liselotte Hardy ◽

...

Keyword(s):

Bacterial Species ◽

Ease Of Use ◽

Bacterial Identification ◽

Gram Positive ◽

Gram Negative ◽

Low Resource Settings ◽

Low Resource ◽

Bacillus Spp ◽

Gram Negative Organisms ◽

Instructions For Use

Bacterial identification is challenging in low-resource settings (LRS). We evaluated the MicroScan identification panels (Beckman Coulter, Brea, CA, USA) as part of Médecins Sans Frontières’ Mini-lab Project. The MicroScan Dried Overnight Positive ID Type 3 (PID3) panels for Gram-positive organisms and Dried Overnight Negative ID Type 2 (NID2) panels for Gram-negative organisms were assessed with 367 clinical isolates from LRS. Robustness was studied by inoculating Gram-negative species on the Gram-positive panel and vice versa. The ease of use of the panels and readability of the instructions for use (IFU) were evaluated. Of species represented in the MicroScan database, 94.6% (185/195) of Gram-negative and 85.9% (110/128) of Gram-positive isolates were correctly identified up to species level. Of species not represented in the database (e.g., Streptococcus suis and Bacillus spp.), 53.1% out of 49 isolates were incorrectly identified as non-related bacterial species. Testing of Gram-positive isolates on Gram-negative panels and vice versa (n = 144) resulted in incorrect identifications for 38.2% of tested isolates. The readability level of the IFU was considered too high for LRS. Inoculation of the panels was favorably evaluated, whereas the visual reading of the panels was considered error-prone. In conclusion, the accuracy of the MicroScan identification panels was excellent for Gram-negative species and good for Gram-positive species. Improvements in stability, robustness, and ease of use have been identified to assure adaptation to LRS constraints.

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The Development of an Effective Bacterial Single-Cell Lysis Method Suitable for Whole Genome Amplification in Microfluidic Platforms

Micromachines ◽

10.3390/mi9080367 ◽

2018 ◽

Vol 9 (8) ◽

pp. 367 ◽

Cited By ~ 6

Author(s):

Yuguang Liu ◽

Dirk Schulze-Makuch ◽

Jean-Pierre de Vera ◽

Charles Cockell ◽

Thomas Leya ◽

...

Keyword(s):

Single Cell ◽

Single Cells ◽

Bacterial Species ◽

Cell Manipulation ◽

Microfluidic Platforms ◽

Gram Positive ◽

Gram Negative ◽

Genome Amplification ◽

Single Cell Sequencing ◽

Wide Range

Single-cell sequencing is a powerful technology that provides the capability of analyzing a single cell within a population. This technology is mostly coupled with microfluidic systems for controlled cell manipulation and precise fluid handling to shed light on the genomes of a wide range of cells. So far, single-cell sequencing has been focused mostly on human cells due to the ease of lysing the cells for genome amplification. The major challenges that bacterial species pose to genome amplification from single cells include the rigid bacterial cell walls and the need for an effective lysis protocol compatible with microfluidic platforms. In this work, we present a lysis protocol that can be used to extract genomic DNA from both gram-positive and gram-negative species without interfering with the amplification chemistry. Corynebacterium glutamicum was chosen as a typical gram-positive model and Nostoc sp. as a gram-negative model due to major challenges reported in previous studies. Our protocol is based on thermal and chemical lysis. We consider 80% of single-cell replicates that lead to >5 ng DNA after amplification as successful attempts. The protocol was directly applied to Gloeocapsa sp. and the single cells of the eukaryotic Sphaerocystis sp. and achieved a 100% success rate.

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Evaluating Flinders Technology Associates card for transporting bacterial isolates and retrieval of bacterial DNA after various storage conditions

Veterinary World ◽

10.14202/vetworld.2020.2243-2251 ◽

2020 ◽

Vol 13 (10) ◽

pp. 2243-2251

Author(s):

Azhar G. Shalaby ◽

Neveen R. Bakry ◽

Abeer A. E. Mohamed ◽

Ashraf A. Khalil

Keyword(s):

Nucleic Acid ◽

Bacterial Species ◽

Storage Conditions ◽

Animal Origin ◽

Cell Detection ◽

Bacterial Cells ◽

Gram Positive ◽

Bacterial Dna ◽

Gram Negative ◽

Fta Cards

Background and Aim: Flinders Technology Associates (FTA) cards simplify sample storage, transport, and extraction by reducing cost and time for diagnosis. This study evaluated the FTA suitability for safe transport and storage of Gram-positive and Gram-negative bacterial cells of animal origin on its liquid culture form and from organ impression smears (tissues) under the same routine condition of microbiological laboratory along with detecting their nucleic acid over different storage conditions. Materials and Methods: Increase in bacterial count from 104 to 107 (colony-forming units/mL) of 78 isolates representing seven bacterial species was applied onto cards. FTA cards were grouped and inoculated by these bacteria and then stored at different conditions of 24-27°C, 4°C, and –20°C for 24 h, for 2 weeks, for 1 and 3 month storage, respectively. Bacteriological examination was done, after which bacterial DNA was identified using specific primers for each bacterial type and detected by polymerase chain reaction (PCR). Results: The total percentage of recovered bacteria from FTA cards was 66.7% at 24-27–C for 24 h, the detection limit was 100% in Gram-positive species, while it was 57.4% in Gram-negative ones. Regarding viable cell detection from organ impression smears, it was successful under the previous conditions. No live bacterial cells were observed by bacteriological isolation rather than only at 24-27°C for 24 h storage. All bacterial DNA were sufficiently confirmed by the PCR technique at different conditions. Conclusion: Overall, the FTA card method was observed to be a valid tool for nucleic acid purification for bacteria of animal origin in the form of culture or organ smears regardless of its Gram type and is used for a short time only 24 h for storage and transport of live bacteria specifically Gram-positive type. Moreover, the bacterial nucleic acid was intact after storage in –20°C for 3 months and was PCR amplifiable.

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Diversity profiling of xenic cultures of Dientamoeba fragilis following systematic antibiotic treatment and prospects for genome sequencing

Parasitology ◽

10.1017/s0031182019001173 ◽

2019 ◽

Vol 147 (1) ◽

pp. 29-38

Author(s):

Rory Gough ◽

Joel Barratt ◽

Damien Stark ◽

John Ellis

Keyword(s):

Antibiotic Treatment ◽

Genome Sequencing ◽

Ribosomal Dna ◽

Bacterial Species ◽

Nutritional Requirement ◽

Gram Negative Bacteria ◽

Gram Positive ◽

Gram Negative ◽

Dientamoeba Fragilis ◽

The Impact

AbstractThe presence of bacterial DNA in Dientamoeba fragilis DNA extracts from culture poses a substantial challenge to sequencing the D. fragilis genome. However, elimination of bacteria from D. fragilis cultures has proven difficult in the past, presumably due to its dependence on some unknown prokaryote/s. This study explored options for removal of bacteria from D. fragilis cultures and for the generation of genome sequence data from D. fragilis. DNA was extracted from human faecal samples and xenic D. fragilis cultures. Extracts were subjected to 16S ribosomal DNA bacterial diversity profiling. Xenic D. fragilis cultures were then subject to antibiotic treatment regimens that systematically removed bacterial species depending on their membrane structure (Gram-positive or Gram-negative) and aerobic requirements. The impact of these treatments on cultures was assessed by 16S amplicon sequencing. Prior to antibiotic treatment, the cultures were dominated by Gram-negative bacteria. Addition of meropenem to cultures eliminated anaerobic Gram-negative bacteria, but it also led to protozoan death after 5 days incubation. The seeding of meropenem resistant Klebsiella pneumoniae strain KPC-2 into cultures before treatment by meropenem prevented death of D. fragilis cells beyond this 5 day period, suggesting that one or more species of Gram-negative bacteria may be an essential nutritional requirement for D. fragilis. Gram-positive cells were completely eliminated using vancomycin without affecting trophozoite growth. Finally, this study shows that genome sequencing of D. fragilis is feasible following bacterial elimination from cultures as the result of the major advances occurring in bioinformatics. We provide evidence on this fact by successfully sequencing the D. fragilis 28S large ribosomal DNA subunit gene using culture-derived DNA.

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Galactose Metabolism Plays a Crucial Role in Biofilm Formation by Bacillus subtilis

mBio ◽

10.1128/mbio.00184-12 ◽

2012 ◽

Vol 3 (4) ◽

Cited By ~ 83

Author(s):

Yunrong Chai ◽

Pascale B. Beauregard ◽

Hera Vlamakis ◽

Richard Losick ◽

Roberto Kolter

Keyword(s):

Bacillus Subtilis ◽

Biofilm Formation ◽

Carbon Sources ◽

Galactose Metabolism ◽

Content Type ◽

Leloir Pathway ◽

The Matrix ◽

Living Organisms ◽

Galactose Metabolites ◽

Biofilm Matrix

ABSTRACTGalactose is a common monosaccharide that can be utilized by all living organisms via the activities of three main enzymes that make up the Leloir pathway: GalK, GalT, and GalE. InBacillus subtilis, the absence of GalE causes sensitivity to exogenous galactose, leading to rapid cell lysis. This effect can be attributed to the accumulation of toxic galactose metabolites, since thegalEmutant is blocked in the final step of galactose catabolism. In a screen for suppressor mutants restoring viability to agalEnull mutant in the presence of galactose, we identified mutations insinR, which is the major biofilm repressor gene. These mutations caused an increase in the production of the exopolysaccharide (EPS) component of the biofilm matrix. We propose that UDP-galactose is the toxic galactose metabolite and that it is used in the synthesis of EPS. Thus, EPS production can function as a shunt mechanism for this toxic molecule. Additionally, we demonstrated that galactose metabolism genes play an essential role inB. subtilisbiofilm formation and that the expressions of both thegalandepsgenes are interrelated. Finally, we propose thatB. subtilisand other members of theBacillusgenus may have evolved to utilize naturally occurring polymers of galactose, such as galactan, as carbon sources.IMPORTANCEBacteria switch from unicellular to multicellular states by producing extracellular matrices that contain exopolysaccharides. In such aggregates, known as biofilms, bacteria are more resistant to antibiotics. This makes biofilms a serious problem in clinical settings. The resilience of biofilms makes them very useful in industrial settings. Thus, understanding the production of biofilm matrices is an important problem in microbiology. In studying the synthesis of the biofilm matrix ofBacillus subtilis, we provide further understanding of a long-standing microbiological observation that certain mutants defective in the utilization of galactose became sensitive to it. In this work, we show that the toxicity observed before was because cells were grown under conditions that were not propitious to produce the exopolysaccharide component of the matrix. When cells are grown under conditions that favor matrix production, the toxicity of galactose is relieved. This allowed us to demonstrate that galactose metabolism is essential for the synthesis of the extracellular matrix.

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A novel compound from the marine bacteriumBacillus pumilusS6-15 inhibits biofilm formation in Gram-positive and Gram-negative species

Biofouling ◽

10.1080/08927014.2011.586127 ◽

2011 ◽

Vol 27 (5) ◽

pp. 519-528 ◽

Cited By ~ 67

Author(s):

Chari Nithya ◽

Muthu Gokila Devi ◽

Shunmugiah Karutha Pandian

Keyword(s):

Biofilm Formation ◽

Gram Positive ◽

Gram Negative

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Effect of Organochlorine Pesticides on Growth and Biological Activities of Bacterial Species Important to the Dairy Food Industry

Journal of Milk and Food Technology ◽

10.4315/0022-2747-39.2.90 ◽

1976 ◽

Vol 39 (2) ◽

pp. 90-94 ◽

Cited By ~ 1

Author(s):

MARY L. SANDFORD ◽

B. E. LANGLOIS

Keyword(s):

Food Industry ◽

Biological Activities ◽

Bacterial Species ◽

Inhibitory Effect ◽

Growth Patterns ◽

Gram Positive ◽

Dairy Food ◽

Gram Negative ◽

Generation Times ◽

Slight Inhibition

Three growth patterns (no effect, slight inhibition, or complete inhibition) were observed when bacterial species common to the dairy-food industry were grown in media containing 50 or 100 ppm DDT, dieldrin, or endrin. The pattern obtained appeared to depend on species and type and concentration of pesticide. All pesticides studied had a greater inhibitory effect on gram positive species than they had on gram negative species when grown in broth. Acid production by lactic acid bacteria was inhibited in broth plus 5 ppm chlordane or heptachlor but unaffected in skimmilk plus up to 100 ppm of these pesticides. Generation times for gram negative species grown in broth plus 10 ppm chlordane or heptachlor were similar to those obtained in controls. Growth of gram positive species was inhibited in broth plus 10 ppm of these pesticides but unaffected in skimmilk containing similar pesticide concentrations. Generation times for several gram negative species were increased by 10 ppm heptachlor in skimmilk.

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In Vitro Study of Antimicrobial Percutaneous Nephrostomy Catheters for Prevention of Renal Infections

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02596-16 ◽

2017 ◽

Vol 61 (6) ◽

Cited By ~ 2

Author(s):

Nylev Vargas-Cruz ◽

Ruth A. Reitzel ◽

Joel Rosenblatt ◽

Mohamed Jamal ◽

Ariel D. Szvalb ◽

...

Keyword(s):

Antimicrobial Activity ◽

Biofilm Formation ◽

In Vitro Study ◽

Percutaneous Nephrostomy ◽

Multidrug Resistant ◽

Gram Positive ◽

Gram Negative ◽

Content Type ◽

Fungal Organisms

ABSTRACT Percutaneous nephrostomy (PCN) catheters are the primary method for draining ureters obstructed by malignancy and preventing a decline of renal function. However, PCN catheter-related infections, such as pyelonephritis and urosepsis, remain a significant concern. Currently, no antimicrobial PCN catheters are available for preventing infection complications. Vascular catheters impregnated with minocycline-rifampin (M/R) and M/R with chlorhexidine coating (M/R plus CHD) have previously demonstrated antimicrobial activity. Therefore, in this study, we examined whether these combinations could be applied to PCN catheters and effectively inhibit biofilm formation by common uropathogens. An in vitro biofilm colonization model was used to assess the antimicrobial efficacy of M/R and M/R-plus-CHD PCN catheters against nine common multidrug-resistant Gram-positive and Gram-negative uropathogens as well as Candida glabrata and Candida albicans. Experimental catheters were also assessed for durability of antimicrobial activity for up 3 weeks. PCN catheters coated with M/R plus CHD completely inhibited biofilm formation for up to 3 weeks for all the organisms tested. The reduction in colonization compared to uncoated PCN catheters was significant for all Gram-positive, Gram-negative, and fungal organisms (P < 0.05). M/R-plus-CHD PCN catheters also produced significant reductions in biofilm colonization relative to M/R PCN catheters for Enterobacter spp., Escherichia coli, Pseudomonas aeruginosa, methicillin-resistant Staphylococcus aureus, vancomycin-resistant enterococci, C. glabrata, and C. albicans (P < 0.05). M/R-plus-CHD PCN catheters proved to be highly efficacious in preventing biofilm colonization when exposed to multidrug-resistant pathogens common in PCN catheter-associated pyelonephritis. M/R-plus-CHD PCN catheters warrant evaluation in a clinical setting to assess their ability to prevent clinically relevant nephrostomy infections.

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