Biofilm Screening Research Articles

Isolation and screening of high biofilm producing lactic acid bacteria, and exploration of its effects on the microbial hazard in corn straw silage

Silage is a well-established method for preserving feed. However, the preparation process still poses several potential microbial hazards. Lactic acid bacteria exhibiting a biofilm phenotype are considered the most advanced 'fourth-generation probiotics' due to their significant potential in enhancing fermentation quality. In this study, a strain of high-biofilm-producing lactic acid bacteria (HBP-LAB) was successfully isolated from silage samples using the crystal violet method and designated as Lactiplantibacillus plantarum S23Y. This strain was subsequently used as an inoculant in corn straw for experimental purposes. The results indicated that it effectively reduced dry matter loss caused by microorganisms, thereby enhancing the retention of dry matter in silage. Following aerobic exposure, this strain was able to maintain the population of Lactobacillus and the concentration of lactic acid, which significantly decreased the likelihood of yeast-induced aerobic spoilage and improved the aerobic stability of the silage. However, it is important to note that this HBP-LAB did not have a significant impact on antibiotic resistance genes (ARGs) or mobile genetic elements (MGEs) in the silage. In conclusion, using S23Y as a representative strain, we have demonstrated that HBP-LAB can enhance the fermentation quality of silage to a certain extent and mitigate the detrimental effects of microorganisms. The findings of this study provide valuable insights for the application of lactic acid bacteria with a biofilm phenotype in silage fermentation.

Multi-trait efficiency and interactivity of bacterial consortia used to enhance plant performance under water stress conditions

Water stress is a major limiting factor for agricultural production under current and projected climate change scenarios. As a sustainable strategy, plant growth-promoting bacterial consortia have been used to reduce plant water stress. However, few studies have examined the effects of stress on multi-trait efficiency and interactivity of bacterial species. In this study, we used several in-vitro experiments, plant assays and greenhouse trials to investigate the effects of stress and bacterial consortia on 1-aminocyclopropane-1-carboxylic acid deaminase (ACCD) activities, indole-3-acetic acid (IAA) production and plant growth-promoting traits (Phosphate-solubilization, starch hydrolysis, siderophores and ammonium production). We further assessed biofilm formation and the chemotactic behaviour in response to ACC. A total of fifteen ACCD rhizobacteria with multiple growth-promoting traits from the dominant plant species from the hyperseasonal Aripo Savannas were screened in this study. Five of the isolates were further analyzed based on their ACCD activities and were tested in single and dual consortium to assess their abilities in promoting growth under simulated drought stress (−0.35 MPa) and chemically induced ACC conditions (0.03 mM). Our findings showed that bacteria which produce high concentrations of IAA affected the isolates’ ability to promote growth under stress, irrespective of microbial combination with ACCD activity above the minimal threshold of 20 nmol α-ketobutyrate mg−1 h−1. Biofilm production with co-culture interaction varied greatly across treatments, however, the general trend showed an increase in biofilm under stress induce conditions. The best performing co-culture, UWIGT-83 and UWIGT-120 (Burkholderia sp.) showed enhanced growth in germination assays and in greenhouse trials with Capsicum chinense (Moruga red hot peppers) under drought stress, when compared to non-inoculated treatments. The findings highlight the importance of testing interactivity of bacterial species with multiple growth promoting traits under stress conditions; and proposed the use of ACC growth media as a novel biofilm screening method for selecting potential stress plant growth-promoting bacteria. Better screening strategies for appropriate plant growth-promoting bacteria may narrow the inconsistency observed between laboratory and field trials.

Antimicrobials discovery against Staphylococcus aureus by high throughput screening of drug library

To develop antimicrobials against Staphylococcus aureus by high throughput screening of drug library. The type of this study is experimental research. The clinical isolates of S. aureus were collected from the sputum samples of respiratory inpatient department of the Third Xiangya Hospital of Central South University. The anti-planktonic cells growth inhibition activity of FDA-approved drugs library (including 1 573 molecules) was assessed by building a planktonic cells screening platform; The biofilm inhibitory effect of the FDA-approved drugs was detected by building a biofilm screening platform combined with crystal violet staining; Minimal inhibitory concentrations of the selected hits were determined by broth microdilution assay. Finally, the cytotoxicity of the selected hits was detected by CCK-8 assay. The results showed that 218 hits were exhibited effective growth inhibitory effects against S. aureus by setting the concentrations of the molecules in the FDA-approved library to 100 μmol/L. These selected molecules are mainly anti-infective drugs, accounting for 118 hits; Followed by anti-cancer drugs, anti-inflammatory/-immune drugs, neurological drugs, cardiovascular drugs, endocrine drugs, and metabolic disease drugs, which accounts for 40, 19, 12, 9, 8, and 3 hits; Other unclassified drugs accounts for 9 hits. The top 10 hits exhibiting anti-planktonic cells activity against S. aureus were mainly including antitumor drugs, followed by neurological drugs and unclassified drugs like vitamin K3 with the inhibition rate of 99.65%-100%. Similarly, the top 10 hits showing biofilm inhibitory effects against S. aureus were also mainly including antitumor drugs, followed by neurological drugs and anti-inflammatory/-immune drugs with the inhibition rate of 50.22%-92.95%. The minimal inhibitory concentration (MIC) of the 51 hits by second round screening was determined by micro-dilution assay, which mainly include the antitumor drugs, cardiovascular drugs, endocrine drugs, anti-inflammatory/-immune drugs, metabolic disease drugs, neurological drugs and other unclassified drugs accounted for 22, 5, 3, 9, 2, 5 and 5 hits, respectively, with the MICs of 1.56-50 μmol/L, 6.25-25 μmol/L, 6.25-25 μmol/L, 0.2-50 μmol/L, 25-50 μmol/L, 1.56-50 μmol/L and 0.1-12.5 μmol/L, respectively. In conclusion, the minimum inhibitory concentrations of small molecules screened through high-throughput assay are at the level of micromolar with strong drug development potential and high modifiability. The high effective anti-planktonic cells and anti-biofilm activity by these molecules are expected to provide new ideas for the development of new antimicrobials against S. aureus.

A Simple and Reproducible Stereomicroscopic Method to Assess Fungal Biofilms: Application to Antifungal Susceptibility Testing.

Candida albicans, a well-known opportunistic pathogen, is a major cause of human fungal infections. Biofilm formation is considered an important pathogenesis factor. Biofilms are less sensitive to antibiotics and immune responses, allowing them to colonize and persist in host niches. Biofilm screening is important in the identification of anti-biofilm drugs. However, developing nations, with limited financial resources, often do not have access to advanced scientific equipment. Here, we describe an in vitro, protocol using common materials and simple equipment to evaluate static microbial biofilms.

Antibiotic effectiveness on biofilm-producing Escherichia coli isolated from catheterized patients

Biofilm is one of the factors that facilitate the occurrence of catheter-associated urinary tract infection (CAUTI). Escherichia coli is reported as one of the most dominant bacteria that have virulence factors including biofilm formation. Uropathogenic E. coli (UPEC) shows increasing resistance to several antibiotics. Examination of the antibiotic sensitivity on the biofilm-producing E. coli and its activity on biofilm formation are important for selecting high effectiveness antibiotics which is beneficial for the management of CAUTI patients. A total of 35 E. coli isolates were recultured in the medium of LB agar and blood agar. The isolates were evaluated the sensitivity based on their MIC value to several antibiotics. In addition, the antibiofilm activity of the antibiotics based on their MBIC value was also evaluated. The data obtained were analyzed both descriptively and analytically. Almost the E. coli isolates have good sensitivity to meropenem antibiotics, amoxicillin-clavulanic acid, and Fosfomycin. However, among the evaluated antibiotics, only fosfomycin that showed antibiofilm activity. The different in terms of the resistance phenotype between the urinary isolates and the catheter isolates was observed. However, there were no significantly differences in the MIC value (pMIC=0.522) and the MBIC value (pMBIC = 0.523). In conclusion, the alternatives of antibiotic therapy for the planktonic bacteria are amoxicillin-clavulanic acid and fosfomycin, while for the biofilm bacteria is fosfomycin. A biofilm screening examination on the catheter to improve the effectiveness of therapy management for CAUTI patients is recommended.

Bioremediation on a chip: A portable microfluidic device for efficient screening of bacterial biofilm with polycyclic aromatic hydrocarbon removal capacity.

Polycyclic aromatic hydrocarbons (PAHs) are pollutants of critical environmental and public health concern and their elimination from contaminated sites is significant for the environment. Biodegradation studies have demonstrated the ability of bacteria in biofilm conformation to enhance the biodegradation of pollutants. In this study, we used our newly developed microfluidic platform to explore biofilm development, properties, and applications of fluid flow, as a new technique for screening PAHs-degrading biofilms. The optimization and evaluation of the flow condition in the microchannels were performed through computational fluid dynamics (CFD). The formation of biofilms by PAHs-degrading bacteria Pseudomonas sp. P26 and Gordonia sp. H19, as pure cultures and co-culture, was obtained in the developed microchips. The removal efficiencies of acenaphthene, fluoranthene and pyrene were determined by HPLC. All the biofilms formed in the microchips removed all tested PAHs, with the higher removal percentages observed with the Pseudomonas sp. P26 biofilm (57.4% of acenaphthene, 40.9% of fluoranthene, and 28.9% of pyrene). Pseudomonas sp. P26 biofilm removed these compounds more efficiently than planktonic cultures. This work proved that the conformation of biofilms enhances the removal rate. It also provided a new tool to rapid and low-cost screen for effective pollutant-degrading biofilms.

QPCR screening for Yersinia ruckeri clonal complex 1 against a background of putatively avirulent strains in Norwegian aquaculture.

Although a number of genetically diverse Yersinia ruckeri strains are present in Norwegian aquaculture environments, most if not all outbreaks of yersiniosis in Atlantic salmon in Norway are associated with a single specific genetic lineage of serotype O1, termed clonal complex 1. To investigate the presence and spread of virulent and putatively avirulent strains in Norwegian salmon farms, PCR assays specific for Y. ruckeri (species level) and Y. ruckeri clonal complex 1 were developed. Following extensive screening of water and biofilm, the widespread prevalence of putatively avirulent Y. ruckeri strains was confirmed in freshwater salmon hatcheries, while Y. ruckeri clonal complex 1 was found in fewer farms. The formalin‐killed bacterin yersiniosis vaccine was detected in environmental samples by both PCR assays for several weeks post‐vaccination. It is thus important to interpret results from recently vaccinated fish with great care. Moreover, field studies and laboratory trials confirmed that stressful management procedures may result in increased shedding of Y. ruckeri by sub‐clinically infected fish. Analysis of sea water sampled throughout thermal delousing procedures proved effective for detection of Y. ruckeri in sub‐clinically infected populations.

Regulatory role of CsuR (YiaU) in determination of cell surface properties of Escherichia coli K-12.

Genomic SELEX screening was performed to identify the binding sites of YiaU, an uncharacterized LysR family transcription factor, on the Escherichia coli K-12 genome. Five high-affinity binding targets of YiaU were identified, all of which were involved in the structures of the bacterial cell surface such as outer and inner membrane proteins, and lipopolysaccharides. Detailed in vitro and in vivo analyses suggest that YiaU activates these target genes. To gain insight into the effects of YiaU in vivo on physiological properties, we used phenotype microarrays, biofilm screening assays and the sensitivity against serum complement analysed using a yiaU deletion mutant or YiaU expression strain. Together, these results suggest that the YiaU regulon confers resistance to some antibiotics, and increases biofilm formation and complement sensitivity. We propose renaming YiaU as CsuR (regulator of cell surface).

Isolation of Biofilm Producing Bacteria from Stool Samples and Their Antibiogram

Aim: Bacterial biofilm formation is a menacing attribute that has been linked to a rise in antibiotic resistance. The aim of this study was to isolate biofilm-producing bacteria from stool samples and their antibiogram. Study Design: The study involved laboratory research, statistical analysis and interpretation of the data. Place and Duration of Study: The research was carried out in three (3) hospital facilities. They are University of Port Harcourt Teaching Hospital (UPTH), Meridian Hospital D/line branch (MRD1), and Meridian Hospital Ikoku branch (MDR2). Specimens were gathered within three (3) months, and analyses were performed. Methodology: 45 stool specimens were collected from the three (3) hospitals. The specimens were correctly labelled with the sampling date and time. Standard microbiological procedures were performed on the collected specimens, including plate counts, identification, biofilm screening, sensitivity testing, extended spectrum beta lactamase phenotypic screening, and molecular characterization of the isolates. Results: The total heterotrophic bacterial counts ranged from 6.2 to 8.2 x107 cfu/g, total coliform counts ranged from 3.2 to 4.1 x106 cfu/g and faecal coliform counts ranged from 1.3 to 1.4 x105 cfu/g. There was no significant difference at (p≤0.05) in the total heterotrophic bacterial counts, total and faecal coliform counts between the hospitals sampled. A total of Thirty-two (32) bacterial isolates were identified in stool specimens, with 20 (62.5%) of them being biofilm producers. Staphylococcus 30%, 35% Escherichia coli, 25% Enterococcus and 10% Bacillus species were detected among biofilm bacteria. Biofilm isolates showed a variety of susceptibility patterns and antibiotic resistance was found in biofilm bacteria. Most bacterial intestinal tract infections from patients and hospitals investigated can be treated with ofloxacin, Gentamycin, Imipenem, and Nitrofurantoin. TET A and CTX-M genes, were reported in Bacillus subtilis and Escherichia coli biofilm bacteria as possible genes that could confer antibiotic resistance.The existence of the icaD and papC genes in Bacillus subtilis and Escherichia coli has been discovered to be probable determinants that impart biofilm forming abilities, according to genomic studies. Conclusion: The existence of biofilm-producing bacteria in patients’ stools, as well as their antibiotic resistance, was observed in this study. Ceftazidime (third generation cephalosporin) resistance was found in both biofilm and non-biofilm bacteria. This research reveals that ofloxacin, Gentamycin, Imipenem, and Nitrofurantoin are the drugs of choice for bacterial intestinal tract infections caused by Escherichia coli, Staphylococcus, Enterococcus, and Bacillus species. As a result, proper infection control techniques and therapeutic recommendations for proven infections should be swiftly implemented.

In situ graphene-modified carbon microelectrode array biosensor for biofilm impedance analysis

Microbial films, formed by the microorganisms to adapt to their living environment, are closely related to the drug resistance and persistent infection of certain diseases such as gingivitis. Impedimetric biosensors could provide an effective means to study the biofilm formation and screening antibiofilm drugs, their performance is closely dependent on the interface materials used thereon. Here, we report a sensitive biosensor platform for this purpose by integration of graphene with carbon microelectrode arrays (C-MEA). A C-MEA was obtained on a silicon substrate through photolithography and subsequent pyrolysis. Characterized by scanning electron microscopy (SEM), Raman spectra, X-ray photoelectron spectroscopy (XPS), transmission electron microscopy (TEM) and electrochemical, we verified that the C-MEAs could evolve into an electrochemically in situ graphene-modified form, GC-MEAs. Using GC-MEA, the growth of biofilm of S. mutans was successfully monitored. Compared to C-MEAs and conventional macrosized counterparts, significant impedance signal amplification and wider detectable frequency range were achieved on the GC-MEAs. Besides, we demonstrated the impedance sensing technology presents high specificity towards the interface-stacked biofilm. The increase in impedimetric signal on the GC-MEAs was 1.4∼2.1 times stronger than that of the C-MEAs and 12.2–13.9 times that of the macrosized electrodes with biofilm growth at a detection frequency of 10.0 kHz. In the last part, the destruction of biofilms grown on the GC-MEAs by using cetylpyridinium chloride was readily monitored by the GC-MEAs senor. In general, graphene-based microelectrode arrays show promise for developing high-throughput drug screening platforms for dealing with detrimental biofilms.

Antibiogram of Biofilm Producing Bacteria Isolated from Urine of Patients in Three Hospitals in Port Harcourt, Rivers State

Aim: The aim of this study was to determine the antibiogram of biofilm producing bacteria isolated from urine of patients in three hospitals in Port Harcourt, Rivers State.
 Study Design: The study employs statistical analysis of the data and interpretation
 Place and Duration of Study: The study was conducted at three (3) hospitals; University of Port Harcourt Teaching Hospital (UPTH), Meridian Hospital D / line branch (MRD1) and Meridian Hospital Ikoku branch, all located in Port Harcourt, Rivers State. Sample collection was for three (3) months, analysis was carried out daily and it lasted for six (6) months.
 Methodology: A total of Forty-five (45) urine samples were collected for a period of three (3) months from the three (3) hospitals. The samples were labelled properly, according to date and time of collection. The collected samples were subjected to standard microbiological procedures which includes standard plate counts, identification, biofilm screening, sensitivity testing using Kirby-Bauer disk diffusion method, Phenotypic screening of extended spectrum beta lactamase and molecular characterization of the isolates
 Results: The results of the bacterial population of urine samples from the hospitals showed that the total heterotrophic bacterial counts for Meridian Hospital D/line (MRD1), Meridian Hospital Ikoku (MRD2) and University of Port Harcourt Teaching Hospital (UPTH) ranged from 4.93 - 6.30 x107cfu/ml. The Total coliform count ranged from 1.89-3.04 x106cfu/ml for Meridian Hospital D/line (MRD1), Meridian Hospital Ikoku (MRD2) and University of Port Harcourt Teaching Hospital (UPTH). Total faecal coliform counts ranged from 0.78-1.11 x105CFU/ml for Meridian Hospital D/line (MRD1), Meridian Hospital Ikoku (MRD2) and University of Port Harcourt Teaching Hospital (UPTH). A total of fifty-eight (58) bacterial isolates were isolated from urine of patients and 36(62.1%) isolates were identified as biofilm producers. The biofilm bacteria identified were 17.2% Staphylococcus,6.9% E. coli, 10.3% Pseudomonas, 6.9% Proteus ,10.3% Bacillus and 10.3% Enterococcus species. Biofilm forming ability of bacteria is considered a virulent factor and it is implicated to being a possible cause of increased resistance to most antibiotics. Varying susceptibility pattern was observed among biofilm isolates. Biofilm bacteria were resistant to several groups of antibiotics. Ofloxacin, Gentamycin, Imipenem and Nitrofurantoin can be used as drug of interest for most bacterial biofilm urinary tract infections. CTX-M and TET A gene were identified in the biofilm bacteria in this study to be possible factors that confer resistance to antibiotics. The presence of icaD and papC gene in the isolates whose genome were studied have been found to be possible factors that confers biofilm producing ability. This study indicates the emergence and rapid spread of biofilm producing bacteria and their resistance to antibiotics. Therefore, strict infection control practices as well as therapeutic guidance for confirmed infections should be rapidly initiated.

Extraction and Screening of Biofilm producing Bacterial isolates in Short-and Long-term Catheters

Biofilm is defined as microbial-derived sessile communities enclosed with extra polymeric substances and characterized by cells that are irreversibly attached to a surface. Biofilm bacteria are a serious threat to public health as biofilm formation of bacteria in indwelling medical devices especially in urinary catheters causes persistent infections resistant to treatment with antimicrobial agents. The present study focused to screen the biofilm-producing bacteria from the indwelling urinary catheters. Both short-term and long-term urinary catheters were collected from the catheterized patients admitted in the national and a private hospital. Bacterial population removed from the catheters were tested for biofilm production by widely used three methods such as Tube method, Congo Red Agar method, and Tissue Culture Plate method. In this preliminary screening, the major biofilm producer identified in the present study is E.Coli in all types of catheters. Further, biofilm-producing bacteria were predominantly detected in long-term catheters than short-term catheters. In addition, the identification of more than one bacterial strain in long-term catheters revealed that bacterial diversity increases with the duration of catheterization. The results of the present study revealed that long-term urinary catheters have the potential for survival and diverse biofilm-producing bacteria.

In-Vitro Evaluation of Biofilm and Hemolysis activity of Candida albicans Isolated from Oral Cavity

Candida albicans is a member of the healthy human microflora, colonizing several niches in the body and can cause opportunistic infection under host debilitated and immunocompromised condition. The present study aimed to investigate the in-vitro hemolytic activity of C. albicans isolated from oral cavity and screen biofilm through three different methods. During the study, 200 oral rinse samples from general human population were analyzed in microbiology laboratory of Central Campus of Technology, Tribhuvan University, Hattisar, Dharan. Nepal. Candida albicans were isolated and identified by conventional microbiological procedures. The hemolytic activity was evaluated through two different Sabouraud dextrose broth media (SDB) containing 7% defibrinated human blood, one supplemented with 3% glucose (SDBwG) and the other without glucose (SDBwoG). The biofilm formation was screened through congo red agar, tube method and tissue culture plate method. In this present study, 42 (21%) isolates of Candida albicans were isolated from 200 oral rinse samples. Isolated Candida albicans exhibited mean hemolysis activity of 28.66% on human blood SDB without glucose and 43.55% on human blood SDB with 3% glucose. Tissue culture plate method was considered sensitive, specific and accurate method for quantitative screening of biofilm in comparison to tube method and congo red agar method. This research concluded that Candida albicans exhibited greater hemolytic activity in human blood with glucose (SDBwG) than in human blood without glucose (SDBwoG). This finding explains that an increased blood glucose concentration may contribute to increased hemolysis activity of Candida albicans that could play pathogenic role for inducing infection like oral candidiasis in debilitated host like diabetic patients. Tissue culture plate method can accurately screen biofilms than tube and congo red agar method.
 Int. J. Appl. Sci. Biotechnol. Vol 8(4): 394-399

Identification of biofilm-producing microorganism’s and drug susceptibility pattern from diabetic foot ulcer patients at Puducherry

Diabetes mellitus is a significant health problem worldwide that affects approximately 171 million people; severe complications lead to the development of diabetic foot ulcers. Diabetic ulcer infections are mainly polymicrobial in nature and multidrug-resistant (MDR), which is capable of forming a biofilm, which is the important virulence factor results in treatment failure. The main objectives of this study to investigate the etiologic agents of diabetic foot infections, their antimicrobial resistance and biofilm formation. A total of 200 patient samples were taken from diabetic foot ulcer patients between September 2015 and February 2016. Isolation and identification of microorganism were made according to standard microbiological procedures. Antibiotic Susceptibility testing performed by Kirby Bauer disc diffusion method and the biofilm production was performed by the tube method and Congo Red Method. Out of 200 samples processed, 110 (55%) were polymicrobial, 50 (25%) monomicrobial and 40(20 %)culture Sterile. The most common organism isolated were 82(39%) Pseudomonas aeruginosa,45(21%) Staphylococcus aureus, 48(23%) Candida sp followed by others. Biofilm production was seen in 112 (53%) of the isolates. Antimicrobial drug resistance was higher among 92(82%) biofilm producers than non-biofilm 20(18%) producing microorganisms. Organisms isolated from chronic diabetic foot ulcers cases were multidrug-resistant and biofilm producers. Our study shows the importance of biofilm screening with the usual antibiogram, as a routine technique in diabetic foot ulcers patients for effective treatment.

The Influence of Biofilms on Carbapenem Susceptibility and Patient Outcome in Device Associated K. pneumoniae Infections: Insights Into Phenotype vs Genome-Wide Analysis and Correlation.

Klebsiella pneumoniae is one of the leading causes of nosocomial infections. Carbapenem-resistant K. pneumoniae are on the rise globally. The biofilm forming ability of K. pneumoniae further complicates patient management. There is still a knowledge gap on the association of biofilm formation with patient outcome and carbapenem susceptibility, which is investigated in present study. K. pneumoniae isolates from patients admitted in critical care units with catheters and ventilators were included. K. pneumoniae (n = 72) were subjected to 96-well plate biofilm formation assay followed by MBEC assay for subset of strong biofilm formers. Whole genome sequencing and a core genome phylogenetic analysis in comparison with global isolates were performed. Phenotypic analyses showed a positive correlation between biofilm formation and carbapenem resistance. Planktonic cells observed to be susceptible in vitro exhibited higher MICs in biofilm structure, hence MICs cannot be extrapolated for treatment. The biofilm forming ability had a significant association with morbidity/mortality. Infections by stronger biofilm forming pathogens significantly (p < 0.05) resulted in fewer “average days alive” for the patient (3.33 days) in comparison to those negative for biofilms (11.33 days). Phylogenetic analysis including global isolates revealed clear association of sequence types with genes for biofilm formation and carbapenem resistance. Known hypervirulent clone-ST23 with wcaG, magA, rmpA, rmpA2, and wzc with lack of mutation for hyper-capsulation might be poor biofilm formers. ST15, ST16, ST307, and ST258 (reported global high-risk clones) were wcaJ negative indicating the high potential of biofilm forming capacity. Genes wabG and treC for CPS, bcsA and pgaC for adhesins, luxS for quorum sensing were common in all clades in addition to genes for aerobactin (iutA), allantoin (allS), type I and III fimbriae (fimA, fimH, and mrkD) and pili (pilQ and ecpA). This study is the first of its kind to compare genetic features of antimicrobial resistance with a spectrum covering most of the genetic factors for K. pneumoniae biofilm. These results highlight the importance of biofilm screening to effectively manage nosocomial infections by K. pneumoniae. Further, data obtained on epidemiology and associations of biofilm and resistance genetic factors will serve to enhance our understanding on biofilm mechanisms in K. pneumoniae.

Synthesis and Biological Evaluation of 4/5‐Aroyl‐2‐aminoimidazoles as Microbial Biofilm Inhibitors

AbstractWe had earlier used Oroidin, a 2‐aminoimidazole‐pyrrole natural microbial biofilm inhibitor, as a structural prototype to develop its 4/5‐acyl modified form as an anti‐biofilm lead for further optimization. Herein, as a part of our efforts towards that direction, we report the design, synthesis, as well as anti‐bacterial, anti‐biofilm, and fungal adhesion inhibition activities of various 4/5‐aroyl‐2‐aminoimidazoles. These simplified Oroidin analogues are moderately active against bacterial biofilms and attest to the need for an aliphatic acyl linker that was part of the original 4/5‐acyl modified Oroidin lead. From our studies, protonation propensity of the 2‐aminoimidazole group of the series emerges as an important pre‐requisite for potent antibiofilm activity. Also, the fungal biofilm screening studies identified analogue 8 b as an active molecule.

A high throughput method to investigate nanoparticle entrapment efficiencies in biofilms

The commercial use of nanoparticles has increased in recent years due to their unique characteristics, including high surface area, modifiable shape and surface charge and size-dependent properties. Consequently, a greater number of nanomaterials are now being released into the environment and inevitably interact with the natural ecosystem. Bacterial biofilms have the potential to capture and retain nanoparticles, however the factors determining the specific nanoparticle entrapment efficiencies of biofilms are not yet fully understood. Based on fluorescent intensity measurements we developed a simple and straightforward method that allowed the entrapment of different silica nanoparticles by two Pseudomonas strains to be quantified. It was determined that, regardless of nanoparticle size or surface functionalisation, Pseudomonas putida biofilms showed enhanced entrapment efficiencies compared to Pseudomonas fluorescens biofilms. It was also noted that both biofilms showed a higher entrapment capacity towards positively charged NPs. The method developed has the potential to be utilized for high throughput biofilm screening studies in order to develop a new understating of the relationship between nanoparticle characteristics and its uptake by bacterial biofilms.

Insights into the Microbiological Safety of Wooden Cutting Boards Used for Meat Processing in Hong Kong's Wet Markets: A Focus on Food-Contact Surfaces, Cross-Contamination and the Efficacy of Traditional Hygiene Practices.

Hong Kong’s wet markets play a crucial role in the country’s supply of safe, fresh meat to satisfy the dietary needs of its population. Whilst food safety regulations have been introduced over the past few years to maintain the microbial safety of foods sold from these wet markets, it remains unclear whether the hygiene maintenance that is performed on the wooden cutting boards used for meat-processing is effective. In fact, hygiene maintenance may often be overlooked, and hygiene standards may be insufficient. If so, this may lead to the spread of harmful pathogens through cross-contamination, thereby causing severe risks to public health. The aim of this study was to determine the level of microbial transfer between wooden cutting boards and swine meat of various qualities, using 16S metagenomic sequencing, strain identification and biofilm screening of isolated strains. The results established that: (a) the traditional hygiene practices used for cleaning wooden cutting boards in Hong Kong’s wet markets expose the surfaces to potentially harmful microorganisms; (b) the processing of microbially contaminated meat on cutting boards cleaned using traditional practices leads to cross-contamination; and (c) several potentially pathogenic microorganisms found on the cutting boards have good biofilm-forming abilities. These results reinforce the need to review the traditional methods used to clean wooden cutting boards after the processing of raw meat in Hong Kong’ wet markets so as to prevent cross-contamination events. The establishment of proper hygiene protocols may reduce the spread of disease-causing microorganisms (including antibiotic-resistant microorganisms) in food-processing environments.

In-Vitro Biofilm Formation and Antimicrobial Resistance of Escherichia coli in Diabetic and Nondiabetic Patients.

Background Diabetic patients are more susceptible to urinary tract infection compared to nondiabetic patients, Escherichia coli being the most common uropathogen causing UTI. Unreasonable and incorrect antibiotic prescription for UTI in these patients may induce the development of antibiotic-resistant urinary pathogens resulting in delayed recovery and longer hospitalization. In addition to these, biofilm forming capacity of the pathogen may worsen the problem. The main aim of this cross-sectional study (conducted from March to September 2015) is to detect the biofilm forming capacity of UTI causing micro-organisms and compare the antibiotic resistance pattern of Escherichia coli, the most common cause of UTI, which will help the physician in choosing the best antibiotic. Method Total of 1,099 clean-catch mid stream urine (CCMSU) was processed by standard microbiological technique; 182 were from the diabetic group and 917 nondiabetic. Following identification, all isolates were subjected to antibiotic susceptibility testing using modified Kirby-Bauer disc diffusion method. In-vitro biofilm forming capacity of the isolates were detected by Microtitre plate method. The data were analyzed using SPSS software 16. Result Urinary tract infection was found to be significantly higher in diabetic patients (42.9%) compared to nondiabetic patients (17.4%) with Escherichia coli as the most common uropathogen in both diabetic and nondiabetic groups. Similarly, UTI was more common in elderly population (29.5%). Imipenem, nitrofurantoin and amikacin were found to be the most effective drug for uropathogenic E. coli in both diabetic and nondiabetic patients, whereas amoxicillin, ciprofloxacin, and cotrimoxazole were least effective. Of the total bacterial isolates, 43.3% showed positive results for in-vitro biofilm production by the Microtitre plate method. A significantly higher resistance rate was observed among biofilm producing E. coli for quinolones, cotrimoxazole, and third generation cephalosporin ceftriaxone. Most of the biofilm producers (79.5%) were found to be MDR (p-value 0.015). ConclusionElderly populations with diabetes are at a higher risk of UTI. Higher biofilm production and resistance to in-use antimicrobial agents in this study render its inefficacy for empirical treatment and point out the importance of biofilm screening to ensure the effective management of infection.

Combined Yeast Biofilm Screening – Characterization and Validation of Yeast Related Biofilms in a Brewing Environment with Combined Cultivation and Specific Real-time PCR Screening of Selected Indicator Species

Microbial spoilage of alcohol-free and low-alcohol beers, beer-mixed beverages, and soft drinks is most commonly caused by yeast. Yeast-related biofilms are therefore a serious problem in the production of these beverages. Fast detection of developing biofilms is a key factor to prevent subsequent spoilage of the product. For fast yeast detection, a new specific medium was developed and combined with real-time polymerase chain reaction (PCR) detection of characteristic beverage spoiling yeast species. The medium is based on MYPG broth (malt extract, yeast extract, peptone, glucose broth) with resazurin as a redox indicator for cell activity. The growth and biofilm potential of representative strains of commonly present beverage spoilage yeast species was evaluated using the developed medium. A novel real-time PCR detection system for Rhodotorula mucilaginosa, an early biofilm colonizer, was designed and successfully validated. Two field tests of the medium in combination with real-time PCR were performed. One test showed a differentiated hygienic status on a filler, and the other test tracked the contamination source of Saccharomyces cerevisiae var. diastaticus. Biofilm relevance of the strain set was proven. The modified MYPG proved to be highly sensitive when detecting yeasts. The detection of the selected target species directly in the medium was compatible and can provide detailed hygienic profiles when combined with additional information on the target species. This provides a fast detection method for yeast-related biofilms in brewery environments, differentiated hygienic monitoring, and makes it possible to troubleshoot contamination incidents.