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14 pages, 4337 KiB

Open AccessArticle

Killing of a Multispecies Biofilm Using Gram-Negative and Gram-Positive Targeted Antibiotic Released from High Purity Calcium Sulfate Beads

by Kelly Moore, Anthony Li, Niraj Gupta, Tripti Thapa Gupta, Craig Delury, Sean S. Aiken, Phillip A. Laycock and Paul Stoodley

Microorganisms 2023, 11(9), 2296; https://doi.org/10.3390/microorganisms11092296 - 12 Sep 2023

Cited by 1 | Viewed by 1602

Abstract

Background: Multispecies biofilm orthopedic infections are more challenging to treat than mono-species infections. In this in-vitro study, we aimed to determine if a multispecies biofilm, consisting of Gram positive and negative species with different antibiotic susceptibilities could be treated more effectively using high [...] Read more.

Background: Multispecies biofilm orthopedic infections are more challenging to treat than mono-species infections. In this in-vitro study, we aimed to determine if a multispecies biofilm, consisting of Gram positive and negative species with different antibiotic susceptibilities could be treated more effectively using high purity antibiotic-loaded calcium sulfate beads (HP-ALCSB) containing vancomycin (VAN) and tobramycin (TOB) in combination than alone. Methods: Three sets of species pairs from bioluminescent strains of Pseudomonas aeruginosa (PA) and Staphylococcus aureus (SA) and clinical isolates, Enterococcus faecalis (EF) and Enterobacter cloacae were screened for compatibility. PA + EF developed intermixed biofilms with similar cell concentrations and so were grown on 316L stainless steel coupons for 72 h or as 24 h agar lawn biofilms and then treated with HP-ALCSBs with single or combination antibiotics and assessed by viable count or bioluminescence and light imaging to distinguish each species. Replica plating was used to assess viability. Results: The VAN + TOB bead significantly reduced the PA + EF biofilm CFU and reduced the concentration of surviving antibiotic tolerant variants by 50% compared to single antibiotics. Conclusions: The combination of Gram-negative and positive targeted antibiotics released from HP-ALCSBs may be more effective in treating multispecies biofilms than monotherapy alone. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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10 pages, 1821 KiB

Open AccessCommunication

Effect of pH, Norepinephrine and Glucose on Metabolic and Biofilm Activity of Uropathogenic Microorganisms

by Nadezhda Ignatova, Alina Abidullina, Olga Streltsova, Vadim Elagin and Vladislav Kamensky

Microorganisms 2023, 11(4), 862; https://doi.org/10.3390/microorganisms11040862 - 28 Mar 2023

Cited by 3 | Viewed by 2263

Abstract

Urinary tract infection (UTIs) aremainly caused by a number of anatomical and physiological dysfunctions, but there are also some iatrogenic factors, including the use of certain medications, that contribute to the development of UTIs. The virulence of bacteria that colonize the urinary tract [...] Read more.

Urinary tract infection (UTIs) aremainly caused by a number of anatomical and physiological dysfunctions, but there are also some iatrogenic factors, including the use of certain medications, that contribute to the development of UTIs. The virulence of bacteria that colonize the urinary tract may be modified by pH and by the presence of soluble substances in urine, such as norepinephrine (NE) and glucose. In this work, we studied the influence of NE and glucose across a range of pHs (5, 7, 8) on the biomass, matrix production and metabolism of uropathogenic strains of Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Staphylococcus aureus and Enterococcus faecalis. We used Congo red and gentian violet to stain the extracellular matrix and biomass, respectively, of biofilms. The optical density of staining of the biofilms was measured using a multichannel spectrophotometer. The metabolic activity was analyzed by MTT assay. It was shown that NE and glucose stimulate biomass production both in the Gram-negative and Gram-positive uropathogens. The metabolic activity in the presence of glucose was higher at pH 5 for E. coli (in 4.0 ± 0.1 times), Ps. aeruginosa (in 8.2 ± 0.2 times) and Kl. pneumoniae (in 4.1 ± 0.2 times). Matrix production of Kl. pneumoniae increased under NE (in 8.2 ± 0.2 times) and in the presence of glucose (in 1.5 ± 0.3 times). Thus, NE and glucose in urine may lead to persistent UTI under patient stress and in the case of metabolic glucose disorders. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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16 pages, 1685 KiB

Open AccessArticle

Genomic Diversity, Antimicrobial Susceptibility, and Biofilm Formation of Clinical Acinetobacter baumannii Isolates from Horses

by Johanna Rühl-Teichner, Lisa Jacobmeyer, Ursula Leidner, Torsten Semmler and Christa Ewers

Microorganisms 2023, 11(3), 556; https://doi.org/10.3390/microorganisms11030556 - 22 Feb 2023

Cited by 1 | Viewed by 2067

Abstract

Acinetobacter (A.) baumannii is an opportunistic pathogen that causes severe infections in humans and animals, including horses. The occurrence of dominant international clones (ICs), frequent multidrug resistance, and the capability to form biofilms are considered major factors in the successful spread of A. baumannii [...] Read more.

Acinetobacter (A.) baumannii is an opportunistic pathogen that causes severe infections in humans and animals, including horses. The occurrence of dominant international clones (ICs), frequent multidrug resistance, and the capability to form biofilms are considered major factors in the successful spread of A. baumannii in human and veterinary clinical environments. Since little is known about A. baumannii isolates from horses, we studied 78 equine A. baumannii isolates obtained from clinical samples between 2008 and 2020 for their antimicrobial resistance (AMR), clonal distribution, biofilm-associated genes (BAGs), and biofilm-forming capability. Based on whole-genome sequence analyses, ICs, multilocus (ML) and core-genome ML sequence types (STs), and AMR genes were determined. Antimicrobial susceptibility testing was performed by microbroth dilution. A crystal violet assay was used for biofilm quantification. Almost 37.2% of the isolates were assigned to IC1 (10.3%), IC2 (20.5%), and IC3 (6.4%). Overall, the isolates revealed high genomic diversity. We identified 51 different STs, including 22 novel STs (ST1723–ST1744), and 34 variants of the intrinsic oxacillinase (OXA), including 8 novel variants (OXA-970 to OXA-977). All isolates were resistant to ampicillin, amoxicillin/clavulanic acid, cephalexin, cefpodoxime, and nitrofurantoin. IC1-IC3 isolates were also resistant to gentamicin, enrofloxacin, marbofloxacin, tetracycline, and trimethoprim/sulfamethoxazole. All isolates were susceptible to imipenem. Thirty-one multidrug-resistant (MDR) isolates mainly accumulated in the IC1-IC3 groups. In general, these isolates showed less biofilm formation (IC1 = 25.0%, IC2 = 18.4%, IC3 = 15.0%) than the group of non-IC1-IC3 isolates (58.4%). Isolates belonging to the same ICs/STs revealed identical BAG patterns. BAG blp1 was absent in all isolates, whereas bfmR and pgaA were present in all isolates. At the level of the IC groups, the AMR status was negatively correlated with the isolates’ ability to form a biofilm. A considerable portion of equine A. baumannii isolates revealed ICs/STs that are globally present in humans. Both an MDR phenotype and the capability to form biofilms might lead to therapeutic failures in equine medicine, particularly due to the limited availability of licensed drugs. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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11 pages, 2171 KiB

Open AccessArticle

Comparison between Symptomatic and Asymptomatic Mice after Clostridioides difficile Infection Reveals Novel Inflammatory Pathways and Contributing Microbiota

by Ahmed AbdelKhalek and Sanjeev K. Narayanan

Microorganisms 2022, 10(12), 2380; https://doi.org/10.3390/microorganisms10122380 - 30 Nov 2022

Cited by 4 | Viewed by 2413

Abstract

Clostridioides difficile causes the highest number of nosocomial infections. Currently, treatment options for C. difficile infection (CDI) are very limited, resulting in poor treatment outcomes and high recurrence rates. Although the disease caused by CDI is inflammatory in nature, the role of inflammation [...] Read more.

Clostridioides difficile causes the highest number of nosocomial infections. Currently, treatment options for C. difficile infection (CDI) are very limited, resulting in poor treatment outcomes and high recurrence rates. Although the disease caused by CDI is inflammatory in nature, the role of inflammation in the development of CDI symptoms is contradictory and not completely understood. Hence, the use of anti-inflammatory medication is debatable in CDI. In the current study, we evaluated the genetic and microbiome profiles of mice after infection with C. difficile. These mice were categorized based on the severity of CDI and the results were viewed accordingly. Our results indicate that certain genes are upregulated in severe CDI more than in the moderate case. These include oncostatin-M (OSM), matrix metalloprotease 8 (MMP8), triggering receptor expressed on myeloid cells 1 (Trem-1), and dual oxidase 2 (Duox2). We also investigated the microbiome composition of CDI mice before and after infecting with C. difficile. The results show that C. difficile abundance is not indicative of diseases severity. Certain bacterial species (e.g., Citrobacter) were enriched while others (e.g., Turicibacter) were absent in severe CDI. This study identifies novel inflammatory pathways and bacterial species with a potential role in determining the severity of CDI. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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16 pages, 5266 KiB

Open AccessArticle

Characterization of Mixed-Species Biofilms Formed by Four Gut Microbiota

by Tao Xu, Yue Xiao, Hongchao Wang, Jinlin Zhu, Yuankun Lee, Jianxin Zhao, Wenwei Lu and Hao Zhang

Microorganisms 2022, 10(12), 2332; https://doi.org/10.3390/microorganisms10122332 - 25 Nov 2022

Cited by 6 | Viewed by 2363

Abstract

In natural settings, approximately 40–80% of bacteria exist as biofilms, most of which are mixed-species biofilms. Previous studies have typically focused on single- or dual-species biofilms. To expand the field of study on gut biofilms, we found a group of gut microbiota that [...] Read more.

In natural settings, approximately 40–80% of bacteria exist as biofilms, most of which are mixed-species biofilms. Previous studies have typically focused on single- or dual-species biofilms. To expand the field of study on gut biofilms, we found a group of gut microbiota that can form biofilms well in vitro: Bifidobacterium longum subsp. infantis, Enterococcus faecalis, Bacteroides ovatus, and Lactobacillus gasseri. The increase in biomass and bio-volume of the mixed-species biofilm was confirmed via crystal violet staining, field emission scanning electron microscopy, and confocal laser scanning microscopy, revealing a strong synergistic relationship in these communities, with B. longum being the key biofilm-contributing species. This interaction may be related to changes in the cell number, biofilm-related genes, and metabolic activities. After quantifying the cell number using quantitative polymerase chain reaction, B. longum and L. gasseri were found to be the dominant flora in the mixed-species biofilm. In addition, this study analyzed biological properties of mixed-species biofilms, such as antibiotic resistance, cell metabolic activity, and concentration of water-insoluble polysaccharides. Compared with single-species biofilms, mixed-species biofilms had higher metabolic activity, more extracellular matrix, and greater antibiotic resistance. From these results, we can see that the formation of biofilms is a self-protection mechanism of gut microbiota, and the formation of mixed-species biofilms can greatly improve the survival rate of different strains. Finally, this study is a preliminary exploration of the biological characteristics of gut biofilms, and the molecular mechanisms underlying the formation of biofilms warrant further research. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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12 pages, 2362 KiB

Open AccessArticle

The Effect of the Gallbladder Environment during Chronic Infection on Salmonella Persister Cell Formation

by Juan F. González, Regan Hitt, Baileigh Laipply and John S. Gunn

Microorganisms 2022, 10(11), 2276; https://doi.org/10.3390/microorganisms10112276 - 16 Nov 2022

Cited by 3 | Viewed by 2474

Abstract

Typhoid fever is caused by Salmonella enterica serovar Typhi (S. Typhi). Around 3–5% of individuals infected become chronic carriers, with the gallbladder (GB) as the predominant site of persistence. Gallstones (GS) aid in the development and maintenance of GB carriage, serving as [...] Read more.

Typhoid fever is caused by Salmonella enterica serovar Typhi (S. Typhi). Around 3–5% of individuals infected become chronic carriers, with the gallbladder (GB) as the predominant site of persistence. Gallstones (GS) aid in the development and maintenance of GB carriage, serving as a substrate to which Salmonellae attach and form a biofilm. This biofilm matrix protects bacteria from the host immune system and environmental stress. This shielded environment is an ideal place for the development of persister cells, a transient phenotype of a subset of cells within a population that allows survival after antibiotic treatment. Persisters can also arise in response to harsh environments such as the GB. Here we investigate if GB conditions affect the number of persisters in a Salmonella population. To simulate the chronic GB environment, we cultured biofilms in cholesterol-coated 96-well plates in the presence of ox or human bile. We then treated planktonic or biofilm Salmonella cultures with high concentrations of different antibiotics. This study suggests that biofilms provide a niche for persister cells, but GB conditions either play no role or have a negative influence on persister formation, especially after kanamycin treatment. The antibiotic target was important, as antimicrobials directed against DNA replication or the cell wall had no effect on persister cell formation. Interestingly, repeated treatment with ciprofloxacin increased the percentage of S. Typhimurium persisters in a biofilm, but this increase was abolished by GB conditions. On the other hand, repeated ciprofloxacin treatment of S. Typhi biofilms in GB conditions slightly increased the fraction of persisters. Thus, while the harsh conditions in the GB would be thought to give rise to increased persisters, therefore contributing to the development of chronic carriage, these data suggest persister cell formation is dampened in this environment. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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13 pages, 1895 KiB

Open AccessArticle

Candida albicans Promotes the Antimicrobial Tolerance of Escherichia coli in a Cross-Kingdom Dual-Species Biofilm

by Shintaro Eshima, Sanae Kurakado, Yasuhiko Matsumoto, Takayuki Kudo and Takashi Sugita

Microorganisms 2022, 10(11), 2179; https://doi.org/10.3390/microorganisms10112179 - 3 Nov 2022

Cited by 9 | Viewed by 1824

Abstract

Cross-kingdom multi-species biofilms consisting of fungi and bacteria are often resistant to antimicrobial treatment, leading to persistent infections. We evaluated whether the presence of Candida albicans affects the antibacterial tolerance of Escherichia coli in dual-species biofilms and explored the underlying mechanism. We found [...] Read more.

Cross-kingdom multi-species biofilms consisting of fungi and bacteria are often resistant to antimicrobial treatment, leading to persistent infections. We evaluated whether the presence of Candida albicans affects the antibacterial tolerance of Escherichia coli in dual-species biofilms and explored the underlying mechanism. We found that the survival of E. coli in the presence of antibacterial drugs was higher in dual-species biofilms compared to single-species biofilms. This tolerance-inducing effect was observed in E. coli biofilms that were treated with a C. albicans culture supernatant. To explore the antibacterial tolerance-inducing factor contained in the culture supernatant and identify the tolerance mechanism, a heated supernatant, a supernatant treated with lyticase, DNase, and proteinase K, or a supernatant added to a drug efflux pump inhibitor were used. However, the tolerance-inducing activity was not lost, indicating the existence of some other mechanisms. Ultrafiltration revealed that the material responsible for tolerance-inducing activity was <10 kDa in size. This factor has not yet been identified and needs further studies to understand the mechanisms of action of this small molecule precisely. Nevertheless, we provide experimental evidence that Candida culture supernatant induces E. coli antibacterial tolerance in biofilms. These findings will guide the development of new treatments for dual-species biofilm infections. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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13 pages, 13073 KiB

Open AccessArticle

Determining the Antibiofilm Efficacy of Oregano Gel in an Ex Vivo Model of Percutaneous Osseointegrated Implants

by Jemi Ong, Rose Godfrey, Brad Isaacson, Paul Pasquina and Dustin Williams

Microorganisms 2022, 10(11), 2133; https://doi.org/10.3390/microorganisms10112133 - 28 Oct 2022

Cited by 1 | Viewed by 1996

Abstract

Biofilm contamination is common in patients with percutaneous osseointegrated (OI) implants, leading to frequent infections, irritation, and discomfort. Reported infection rates soar up to 65% as the recalcitrant nature of biofilms complicates treatment. There is persistent need for therapies to manage biofilm burden. [...] Read more.

Biofilm contamination is common in patients with percutaneous osseointegrated (OI) implants, leading to frequent infections, irritation, and discomfort. Reported infection rates soar up to 65% as the recalcitrant nature of biofilms complicates treatment. There is persistent need for therapies to manage biofilm burden. In response, we formulated and tested oregano essential oil in a topical gel as a potential biofilm management therapy. We developed an ex vivo system based on an established ovine OI implant model with Staphylococcus aureus ATCC 6538 biofilms as initial inocula. Gel was administered to the samples across a period of five days. Samples were quantified and colony forming unit (CFU) counts were compared against a positive control (initial bacterial inocula without treatment). Significant biofilm reduction was observed in samples treated with oregano gel compared to controls, demonstrating the potential of an oregano oil-based gel as a biofilm management therapy at the skin-implant interface of percutaneous OI implants. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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17 pages, 2405 KiB

Open AccessArticle

Microbial Biofilms at Meat-Processing Plant as Possible Places of Bacteria Survival

by Yury Nikolaev, Yulia Yushina, Andrey Mardanov, Evgeniy Gruzdev, Ekaterina Tikhonova, Galina El-Registan, Aleksey Beletskiy, Anastasia Semenova, Elena Zaiko, Dagmara Bataeva and Ekaterina Polishchuk

Microorganisms 2022, 10(8), 1583; https://doi.org/10.3390/microorganisms10081583 - 6 Aug 2022

Cited by 8 | Viewed by 3008

Abstract

Biofilm contamination in food production threatens food quality and safety, and causes bacterial infections. Study of food biofilms (BF) is of great importance. The taxonomic composition and structural organization of five foods BF taken in different workshops of a meat-processing plant (Moscow, RF) [...] Read more.

Biofilm contamination in food production threatens food quality and safety, and causes bacterial infections. Study of food biofilms (BF) is of great importance. The taxonomic composition and structural organization of five foods BF taken in different workshops of a meat-processing plant (Moscow, RF) were studied. Samples were taken from the surface of technological equipment and premises. Metagenomic analysis showed both similarities in the presented microorganisms dominating in different samples, and unique families prevailing on certain objects were noted. The bacteria found belonged to 11 phyla (no archaea). The dominant ones were Actinobacteria, Bacteroidetes, Firmicutes, and Proteobacteria. The greatest diversity was in BFs taken from the cutting table of raw material. Biofilms’ bacteria may be the cause of meat, fish and dairy products spoilage possible representatives include Pseudomonas, Flavobacterium, Arcobacter, Vagococcus, Chryseobacterium, Carnobacterium, etc.). Opportunistic human and animal pathogens (possible representatives include Arcobacter, Corynebacterium, Kocuria, etc.) were also found. Electron-microscopic studies of BF thin sections revealed the following: (1) the diversity of cell morphotypes specific to multispecies BFs; (2) morphological similarity of cells in BFs from different samples, micro-colonial growth; (3) age heterogeneity of cells within the same microcolony (vegetative and autolyzed cells, resting forms); (4) heterogeneity of the polymer matrix chemical nature according to ruthenium red staining. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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16 pages, 3825 KiB

Open AccessArticle

Effects of Volatile Organic Compounds on Biofilms and Swimming Motility of Agrobacterium tumefaciens

by Daria E. Sidorova, Mariia I. Skripka, Inessa A. Khmel, Olga A. Koksharova and Vladimir A. Plyuta

Microorganisms 2022, 10(8), 1512; https://doi.org/10.3390/microorganisms10081512 - 26 Jul 2022

Cited by 5 | Viewed by 1968

Abstract

Volatile organic compounds (VOCs) emitted by bacteria play an important role in the interaction between microorganisms and other organisms. They can inhibit the growth of phytopathogenic microorganisms, modulate plant growth, and serve as infochemicals. Here, we investigated the effects of ketones, alcohols, and [...] Read more.

Volatile organic compounds (VOCs) emitted by bacteria play an important role in the interaction between microorganisms and other organisms. They can inhibit the growth of phytopathogenic microorganisms, modulate plant growth, and serve as infochemicals. Here, we investigated the effects of ketones, alcohols, and terpenes on the colony biofilms of plant pathogenic Agrobacterium tumefaciens strains and swimming motility, which can play an important role in the formation of biofilms. It was shown that 2-octanone had the greatest inhibitory effect on biofilm formation, acting in a small amount (38.7 g/m³). Ketone 2-butanone and unsaturated ketone β-ionone reduced the formation of biofilms at higher doses (145.2–580.6 and 387.1–1548.3 g/m³, respectively, up to 2.5–5 times). Isoamyl alcohol and 2-phenylethanol decreased the formation of biofilms at doses of 88.7 and 122.9 g/m³ by 1.7 and 5 times, respectively, with an increased effect at 177.4 and 245.9 g/m³, respectively. The agrobacteria cells in mature biofilms were more resistant to the action of ketones and alcohols. These VOCs also suppressed the swimming motility of agrobacteria; the radius of swimming zones decreased ~from 2 to 5 times. Terpenes (−)-limonene and (+)-α-pinene had no significant influence on the colony biofilms and swimming motility at the doses used. The results obtained represent new information about the effect of VOCs on biofilms and the motility of bacteria. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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11 pages, 2777 KiB

Open AccessArticle

Analysis of the Metabolic Response of Planktonic Cells and Biofilms of Klebsiella pneumoniae to Sublethal Disinfection with Sodium Hypochlorite Measured by NMR

by David Felipe Garcia Mendez, Julián Andrés Rengifo Herrera, Janeth Sanabria and Julien Wist

Microorganisms 2022, 10(7), 1323; https://doi.org/10.3390/microorganisms10071323 - 30 Jun 2022

Cited by 5 | Viewed by 2287

Abstract

Klebsiella pneumoniae is a pathogenic agent able to form biofilms on water storage tanks and pipe walls. This opportunistic pathogen can generate a thick layer as one of its essential virulence factors, enabling the bacteria to survive disinfection processes and thus develop drug [...] Read more.

Klebsiella pneumoniae is a pathogenic agent able to form biofilms on water storage tanks and pipe walls. This opportunistic pathogen can generate a thick layer as one of its essential virulence factors, enabling the bacteria to survive disinfection processes and thus develop drug resistance. Understanding the metabolic differences between biofilm and planktonic cells of the K. pneumoniae response to NaClO is key to developing strategies to control its spread. In this study, we performed an NMR metabolic profile analysis to compare the response to a sublethal concentration of sodium hypochlorite of biofilm and planktonic cells of K. pneumoniae cultured inside silicone tubing. Metabolic profiles revealed changes in the metabolism of planktonic cells after a contact time of 10 min with 7 mg L⁻¹ of sodium hypochlorite. A decrease in the production of metabolites such as lactate, acetate, ethanol, and succinate in this cell type was observed, thus indicating a disruption of glucose intake. In contrast, the biofilms displayed a high metabolic heterogeneity, and the treatment did not affect their metabolic signature. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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17 pages, 2957 KiB

Open AccessArticle

A Porcine Model for the Development and Testing of Preoperative Skin Preparations

by Hannah R. Duffy, Rose W. Godfrey, Dustin L. Williams and Nicholas N. Ashton

Microorganisms 2022, 10(5), 837; https://doi.org/10.3390/microorganisms10050837 - 19 Apr 2022

Cited by 2 | Viewed by 3465

Abstract

Clinical preoperative skin preparations (PSPs) do not eradicate skin flora dwelling in the deepest dermal regions. Survivors constitute a persistent infection risk. In search of solutions, we created a porcine model intended for PSP developmental testing. This model employed microbiological techniques sensitive to [...] Read more.

Clinical preoperative skin preparations (PSPs) do not eradicate skin flora dwelling in the deepest dermal regions. Survivors constitute a persistent infection risk. In search of solutions, we created a porcine model intended for PSP developmental testing. This model employed microbiological techniques sensitive to the deep-dwelling microbial flora as these microorganisms are frequently overlooked when using institutionally-entrenched testing methodologies. Clinical gold-standard PSPs were assessed. Ten Yorkshire pigs were divided into two groups: prepared with either povidone iodine (PVP-I) or chlorhexidine gluconate (CHG) PSP. Bioburdens were calculated on square, 4 cm by 4 cm, full-thickness skin samples homogenized in neutralizing media. Endogenous bioburden of porcine skin (3.3 log₁₀ CFU/cm²) was consistent with natural flora numbers in dry human skin. On-label PSP scrub kits with PVP-I (n = 39) or CHG (n = 40) failed the 2–3 log₁₀-reduction criteria established for PSPs by the Food and Drug Administration (FDA), resulting in a 1.46 log₁₀ and 0.58 log₁₀ reduction, respectively. Porcine dermal microbiota mirrored that of humans, displaying abundant staphylococcal species. Likewise, histological sections showed similarity in hair follicle depths and sebaceous glands (3.2 ± 0.7 mm). These shared characteristics and the considerable fraction of bacteria which survived clinical PSPs make this model useful for developmental work. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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12 pages, 2177 KiB

Open AccessArticle

Biofilm Growth on Simulated Fracture Fixation Plates Using a Customized CDC Biofilm Reactor for a Sheep Model of Biofilm-Related Infection

by Walker Kay, Connor Hunt, Lisa Nehring, Brian Barnum, Nicholas Ashton and Dustin Williams

Microorganisms 2022, 10(4), 759; https://doi.org/10.3390/microorganisms10040759 - 31 Mar 2022

Cited by 3 | Viewed by 2567

Abstract

Most animal models of infection utilize planktonic bacteria as initial inocula. However, this may not accurately mimic scenarios where bacteria in the biofilm phenotype contaminate a site at the point of injury. We developed a modified CDC biofilm reactor in which biofilms can [...] Read more.

Most animal models of infection utilize planktonic bacteria as initial inocula. However, this may not accurately mimic scenarios where bacteria in the biofilm phenotype contaminate a site at the point of injury. We developed a modified CDC biofilm reactor in which biofilms can be grown on the surface of simulated fracture fixation plates. Multiple reactor runs were performed and demonstrated that monomicrobial biofilms of a clinical strain of methicillin-resistant Staphylococcus aureus, S. aureus ATCC 6538, and Pseudomonas aeruginosa ATCC 27853 consistently developed on fixation plates. We also identified a method by which to successfully grow polymicrobial biofilms of S. aureus ATCC 6538 and P. aeruginosa ATCC 27853 on fixation plates. This customized reactor can be used to grow biofilms on simulated fracture fixation plates that can be inoculated in animal models of biofilm implant-related infection that, for example, mimic open fracture scenarios. The reactor provides a method for growing biofilms that can be used as initial inocula and potentially improve the testing and development of antibiofilm technologies. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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23 pages, 5206 KiB

Open AccessArticle

Can Aggregate-Associated Organisms Influence the Fouling in a SWRO Desalination Plant?

by Tamar Jamieson, Harriet Whiley, Jason R. Gascooke and Sophie C. Leterme

Microorganisms 2022, 10(4), 682; https://doi.org/10.3390/microorganisms10040682 - 22 Mar 2022

Cited by 1 | Viewed by 2405

Abstract

This pilot study investigates the formation of aggregates within a desalination plant, before and after pre-treatment, as well as their potential impact on fouling. The objective is to provide an understanding of the biofouling potential of the feed water within a seawater reverse [...] Read more.

This pilot study investigates the formation of aggregates within a desalination plant, before and after pre-treatment, as well as their potential impact on fouling. The objective is to provide an understanding of the biofouling potential of the feed water within a seawater reverse osmosis (SWRO) desalination plant, due to the limited removal of fouling precursors. The 16S and 18S rRNA was extracted from the water samples, and the aggregates and sequenced. Pre-treatment systems, within the plant remove < 5 µm precursors and organisms; however, smaller size particles progress through the plant, allowing for the formation of aggregates. These become hot spots for microbes, due to their nutrient gradients, facilitating the formation of niche environments, supporting the proliferation of those organisms. Aggregate-associated organisms are consistent with those identified on fouled SWRO membranes. This study examines, for the first time, the factors supporting the formation of aggregates within a desalination system, as well as their microbial communities and biofouling potential. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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10 pages, 5147 KiB

Open AccessCommunication

Mapping Bacterial Biofilm on Features of Orthopedic Implants In Vitro

by Kelly Moore, Niraj Gupta, Tripti Thapa Gupta, Khushi Patel, Jacob R. Brooks, Anne Sullivan, Alan S. Litsky and Paul Stoodley

Microorganisms 2022, 10(3), 586; https://doi.org/10.3390/microorganisms10030586 - 8 Mar 2022

Cited by 9 | Viewed by 4967

Abstract

Implant-associated infection is a major complication of orthopedic surgery. One of the most common organisms identified in periprosthetic joint infections is Staphylococcus aureus, a biofilm-forming pathogen. Orthopedic implants are composed of a variety of materials, such as titanium, polyethylene and stainless steel, which [...] Read more.

Implant-associated infection is a major complication of orthopedic surgery. One of the most common organisms identified in periprosthetic joint infections is Staphylococcus aureus, a biofilm-forming pathogen. Orthopedic implants are composed of a variety of materials, such as titanium, polyethylene and stainless steel, which are at risk for colonization by bacterial biofilms. Little is known about how larger surface features of orthopedic hardware (such as ridges, holes, edges, etc.) influence biofilm formation and attachment. To study how biofilms might form on actual components, we submerged multiple orthopedic implants of various shapes, sizes, roughness and material type in brain heart infusion broth inoculated with Staphylococcus aureus SAP231, a bioluminescent USA300 strain. Implants were incubated for 72 h with daily media exchanges. After incubation, implants were imaged using an in vitro imaging system (IVIS) and the metabolic signal produced by biofilms was quantified by image analysis. Scanning electron microscopy was then used to image different areas of the implants to complement the IVIS imaging. Rough surfaces had the greatest luminescence compared to edges or smooth surfaces on a single implant and across all implants when the images were merged. The luminescence of edges was also significantly greater than smooth surfaces. These data suggest implant roughness, as well as large-scale surface features, may be at greater risk of biofilm colonization. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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12 pages, 1404 KiB

Open AccessArticle

Antimicrobial Susceptibility of Persister Biofilm Cells of Bacillus cereus and Pseudomonas fluorescens

by Susana Fernandes, Inês B. Gomes, Sérgio F. Sousa and Manuel Simões

Microorganisms 2022, 10(1), 160; https://doi.org/10.3390/microorganisms10010160 - 13 Jan 2022

Cited by 19 | Viewed by 3242

Abstract

The present study evaluates the antimicrobial susceptibility of persister cells of Bacillus cereus and Pseudomonas fluorescens after their regrowth in suspension and as biofilms. Two conventional (benzalkonium chloride—BAC and peracetic acid—PAA) and two emerging biocides (glycolic acid—GA and glyoxal—GO) were selected for this [...] Read more.

The present study evaluates the antimicrobial susceptibility of persister cells of Bacillus cereus and Pseudomonas fluorescens after their regrowth in suspension and as biofilms. Two conventional (benzalkonium chloride—BAC and peracetic acid—PAA) and two emerging biocides (glycolic acid—GA and glyoxal—GO) were selected for this study. Persister cells resulted from biofilms subjected to a critical treatment using the selected biocides. All biocide treatments developed B. cereus persister cells, except PAA that effectively reduced the levels of vegetative cells and endospores. P. fluorescens persister cells comprise viable and viable but non-culturable cells. Afterwards, persister cells were regrown in suspension and in biofilms and were subjected to a second biocide treatment. In general, planktonic cultures of regrown persister cells in suspension lost their antimicrobial tolerance, for both bacteria. Regrown biofilms of persister cells had antimicrobial susceptibility close to those regrown biofilms of biocide-untreated cells, except for regrown biofilms of persister P. fluorescens after BAC treatment, which demonstrated increased antimicrobial tolerance. The most active biocide against persister cells was PAA, which did not promote changes in susceptibility after their regrowth. In conclusion, persister cells are ubiquitous within biofilms and survive after critical biocide treatment. The descendant planktonic and biofilms populations showed similar properties as the original ones. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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27 pages, 1529 KiB

Open AccessReview

Silver Nanoparticles: Bactericidal and Mechanistic Approach against Drug Resistant Pathogens

by Pragati Rajendra More, Santosh Pandit, Anna De Filippis, Gianluigi Franci, Ivan Mijakovic and Massimiliano Galdiero

Microorganisms 2023, 11(2), 369; https://doi.org/10.3390/microorganisms11020369 - 1 Feb 2023

Cited by 176 | Viewed by 12875

Abstract

This review highlights the different modes of synthesizing silver nanoparticles (AgNPs) from their elemental state to particle format and their mechanism of action against multidrug-resistant and biofilm-forming bacterial pathogens. Various studies have demonstrated that the AgNPs cause oxidative stress, protein dysfunction, membrane disruption, [...] Read more.

This review highlights the different modes of synthesizing silver nanoparticles (AgNPs) from their elemental state to particle format and their mechanism of action against multidrug-resistant and biofilm-forming bacterial pathogens. Various studies have demonstrated that the AgNPs cause oxidative stress, protein dysfunction, membrane disruption, and DNA damage in bacteria, ultimately leading to bacterial death. AgNPs have also been found to alter the adhesion of bacterial cells to prevent biofilm formation. The benefits of using AgNPs in medicine are, to some extent, counter-weighted by their toxic effect on humans and the environment. In this review, we have compiled recent studies demonstrating the antibacterial activity of AgNPs, and we are discussing the known mechanisms of action of AgNPs against bacterial pathogens. Ongoing clinical trials involving AgNPs are briefly presented. A particular focus is placed on the mechanism of interaction of AgNPs with bacterial biofilms, which are a significant pathogenicity determinant. A brief overview of the use of AgNPs in other medical applications (e.g., diagnostics, promotion of wound healing) and the non-medical sectors is presented. Finally, current drawbacks and limitations of AgNPs use in medicine are discussed, and perspectives for the improved future use of functionalized AgNPs in medical applications are presented. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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14 pages, 1331 KiB

Open AccessReview

Biofilm Survival Strategies in Chronic Wounds

by Ida Clement Thaarup, Anne Kristine Servais Iversen, Mads Lichtenberg, Thomas Bjarnsholt and Tim Holm Jakobsen

Microorganisms 2022, 10(4), 775; https://doi.org/10.3390/microorganisms10040775 - 5 Apr 2022

Cited by 33 | Viewed by 5188

Abstract

Bacterial biofilms residing in chronic wounds are thought to have numerous survival strategies, making them extremely difficult to eradicate and resulting in long-term infections. However, much of our knowledge regarding biofilm persistence stems from in vitro models and experiments performed in vivo in [...] Read more.

Bacterial biofilms residing in chronic wounds are thought to have numerous survival strategies, making them extremely difficult to eradicate and resulting in long-term infections. However, much of our knowledge regarding biofilm persistence stems from in vitro models and experiments performed in vivo in animal models. While the knowledge obtained from such experiments is highly valuable, its direct translation to the human clinical setting should be undertaken with caution. In this review, we highlight knowledge obtained from human clinical samples in different aspects of biofilm survival strategies. These strategies have been divided into segments of the following attributes: altered transcriptomic profiles, spatial distribution, the production of extracellular polymeric substances, an altered microenvironment, inter-and intra-species interactions, and heterogeneity in the bacterial population. While all these attributes are speculated to contribute to the enhanced persistence of biofilms in chronic wounds, only some of them have been demonstrated to exist in human wounds. Some of the attributes have been observed in other clinical diseases while others have only been observed in vitro. Here, we have strived to clarify the limitations of the current knowledge in regard to this specific topic, without ignoring important in vitro and in vivo observations. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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11 pages, 9094 KiB

Open AccessReview

Bacterial Biofilms Utilize an Underlying Extracellular DNA Matrix Structure That Can Be Targeted for Biofilm Resolution

by Steven D. Goodman and Lauren O. Bakaletz

Microorganisms 2022, 10(2), 466; https://doi.org/10.3390/microorganisms10020466 - 18 Feb 2022

Cited by 22 | Viewed by 3823

Abstract

Bacterial biofilms contribute significantly to the antibiotic resistance, pathogenesis, chronicity and recurrence of bacterial infections. Critical to the stability and survival of extant biofilms is the extracellular DNA (eDNA)-dependent matrix which shields the resident bacteria from hostile environments, allows a sessile metabolic state, [...] Read more.

Bacterial biofilms contribute significantly to the antibiotic resistance, pathogenesis, chronicity and recurrence of bacterial infections. Critical to the stability and survival of extant biofilms is the extracellular DNA (eDNA)-dependent matrix which shields the resident bacteria from hostile environments, allows a sessile metabolic state, but also encourages productive interactions with biofilm-inclusive bacteria. Given the importance of the eDNA, approaches to this area of research have been to target not just the eDNA, but also the additional constituent structural components which appear to be widespread. Chief among these is a ubiquitous two-member family of bacterial nucleoid associated proteins (the DNABII proteins) responsible for providing structural integrity to the eDNA and thereby the biofilm. Moreover, this resultant novel eDNA-rich secondary structure can also be targeted for disruption. Here, we provide an overview of both what is known about the eDNA-dependent matrix, as well as the resultant means that have resulted in biofilm resolution. Results obtained to date have been highly supportive of continued development of DNABII-targeted approaches, which is encouraging given the great global need for improved methods to medically manage, or ideally prevent biofilm-dependent infections, which remains a highly prevalent burden worldwide. Full article

(This article belongs to the Special Issue Biofilm Formation and Survival Strategies)

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