Polyhexamethylene Biguanide Reduces High-Risk Human Papilloma Virus Viral Load in Cervical Cell Samples Derived from ThinPrep Pap Test.

Human papilloma virus (HPV) infection and its progression still represent a great medical challenge worldwide. Clinical evidence has demonstrated the beneficial effects of polyhexamethylene biguanide (PHMB) on HPV clinical manifestations; however, evidence of the effect of this molecule on HPV viral load is still lacking. In this in vitro study, 13 ThinPrep Papanicolaou (Pap) tests were treated with a PHMB solution (0.10 g/100 mL) for 2 h. We observed no cytological changes but a significant reduction in the viral load of high-risk (HR) HPV after PHMB treatment, also revealing a dose-dependent antiviral effect. In addition, by stratifying the obtained results according to HR-HPV genotype, we observed a significant reduction in the viral load of HPV 16, P2 (56, 59, 66), 31, and P3 (35, 39, 68) and a strong decrease in the viral load of HPV 45, 52, and P1 (33, 58). Overall, 85% of the analyzed cervical cell samples exhibited an improvement in HPV viral load after PHMB exposure, while only 15% remain unchanged. For the first time, the data from this pilot study support the activity of PHMB on a specific phase of the HPV viral lifecycle, the one regarding the newly generated virions, reducing viral load and thus blocking the infection of other cervical cells.

Safety and effectiveness of an antiseptic wound cleansing and irrigation solution containing polyhexamethylene biguanide.

OBJECTIVE: There is currently a wide range of cleansing and irrigation solutions available for wounds, many of which contain antimicrobial agents. The aim of this study was to assess the safety of HydroClean Solution (HARTMANN, Germany), a polyhexamethylene biguanide (PHMB)-containing irrigation solution, in a standard cytotoxicity assay, and to assess its effect in a three-dimensional (3D) full-thickness model of human skin. METHOD: A number of commercially available wound cleansing and irrigation solutions, including the PHMB-containing irrigation solution, were tested in a cytotoxicity assay using L929 mouse fibroblasts (ISO 10993-5:2009). The PHMB-containing irrigation solution was then assessed in an in vitro human keratinocyte-fibroblast 3D full-thickness wounded skin model to determine its effect on wound healing over six days. The effect of the PHMB-containing irrigation solution on tissue viability was measured using a lactate dehydrogenase (LDH) assay, and proinflammatory effects were measured using an interleukin-6 (IL-6) production assay. RESULTS: The PHMB-containing irrigation solution was shown to be equivalent to other commercially available cleansing and irrigation solutions when tested in the L929 fibroblast cytotoxicity assay. When assessed in the in vitro 3D human full-thickness wound healing model, the PHMB-containing irrigation solution treatment resulted in no difference in levels of LDH or IL-6 when compared with levels produced in control Dulbecco's phosphate-buffered saline cultures. There was, however, a pronounced tissue thickening of the skin model in the periwound region. CONCLUSION: The experimental data presented in this study support the conclusion that the PHMB-containing irrigation solution has a safety profile similar to other commercially available cleansing and irrigation solutions. Evidence also suggests that the PHMB-containing irrigation solution does not affect tissue viability or proinflammatory cytokine production, as evidenced by LDH levels or the production of IL-6 in a 3D human full-thickness wound healing model. The PHMB-containing irrigation solution stimulated new tissue growth in the periwound region of the skin model.

Ex vivo wound model on porcine skin for the evaluation of the antibiofilm activity of polyhexamethylene biguanide and ciprofloxacin.

This study aimed to standardize the use of an ex vivo wound model for the evaluation of compounds with antibiofilm activity. The in vitro susceptibility of Staphylococcus aureus ATCC 29213 and Pseudomonas aeruginosa ATCC 27853 to ciprofloxacin and polyhexamethylene biguanide (PHMB) was evaluated in planktonic and biofilm growth. The effects of ciprofloxacin and PHMB on biofilms grown on porcine skin explants were evaluated by colony-forming unit (CFU) counting and confocal microscopy. Minimum inhibitory concentrations (MICs) against S. aureus and P. aeruginosa were, respectively, 0.5 and 0.25 µg mL-1 for ciprofloxacin, and 0.78 and 6.25 µg mL-1 for PHMB. Minimum biofilm eradication concentrations (MBECs) against S. aureus and P. aeruginosa were, respectively, 2 and 8 µg mL-1 for ciprofloxacin, and 12.5 and >25 µg mL-1 for PHMB. Ciprofloxacin reduced (P < 0.05) log CFU counts of the biofilms grown ex vivo by 3 and 0.96 for S. aureus and P. aeruginosa, respectively, at MBEC, and by 0.58 and 8.12 against S. aureus and P. aeruginosa, respectively, at 2xMBEC. PHMB (100 µg/mL) reduced (P < 0.05) log CFU counts by 0.52 for S. aureus and 0.68 log for P. aeruginosa, leading to an overall decrease (P < 0.05) in biofilm biomass. The proposed methodology to evaluate the susceptibility of biofilms grown ex vivo led to reproducible and reliable results.

Clinical protective effects of polyhexamethylene biguanide hydrochloride (PHMB) against Vibrio parahaemolyticus causing translucent post-larvae disease (VpTPD) in Penaeus vannamei.

A new emerging disease called "translucent post-larvae disease" (TPD) in Penaeus vannamei, caused by a novel type of highly lethal Vibro parahaemolyticus (VpTPD), has become an urgent threat to the shrimp farming industry in China. In order to develop an effective disinfectant for the prevention and control of the VpTPD, the clinical protective effects of polyhexamethylene biguanide hydrochloride (PHMB) against VpTPD in Penaeus vannamei were investigated by carrying out an acute toxicity test of PHMB on post-larvae of P. vannamei and its effect of treatment test on VpTPD infection. The results showed that the median lethal concentration of disinfectant (LC50) values of PHMB to post-larvae of P. vannamei after treatment for 24 h, 48 h, 72 h, 96 h were 16.13 mg/L (14.18-18.57), 10.77 mg/L (9.93-11.72), 9.68 mg/L (8.53-11.64), 9.14 mg/L (7.70-10.99), respectively. In addition, a clinical trial showed that 1 mg/L PHMB showed a strong protective effect on the post-larvae of shrimp challenged with 101-104 CFU/ml of VpTPD. The relative percentage survival (RPS) of 1 mg/L PHMB on post-larvae of P. vannamei challenged with VpTPD at 101, 102, 103 and 104 CFU/ml were 63.65 %±6.81, 62.96 %±5.56, 60.00 %±3.75 and 66.67 %±3.75 at 96 hours post infection. The results highlight the clinical protective effects of the PHMB and therefor PHMB can be used as a preventive measure to control early TPD infection in shrimp culture. This study also provides valuable information for the prevention of other bacterial diseases in shrimp culture.

Polyhexamethylene biguanide and its antimicrobial role in wound healing: a narrative review.

A wound offers an ideal environment for the growth and proliferation of a variety of microorganisms which, in some cases, may lead to localised or even systemic infections that can be catastrophic for the patient; the development of biofilms exacerbates these infections. Over the past few decades, there has been a progressive development of antimicrobial resistance (AMR) in microorganisms across the board in healthcare sectors. Such resistant microorganisms have arisen primarily due to the misuse and overuse of antimicrobial treatments, and the subsequent ability of microorganisms to rapidly change and mutate as a defence mechanism against treatment (e.g., antibiotics). These resistant microorganisms are now at such a level that they are of grave concern to the World Health Organization (WHO), and are one of the leading causes of illness and mortality in the 21st century. Treatment of such infections becomes imperative but presents a significant challenge for the clinician in that treatment must be effective but not add to the development of new microbes with AMR. The strategy of antimicrobial stewardship (AMS) has stemmed from the need to counteract these resistant microorganisms and requires that current antimicrobial treatments be used wisely to prevent amplification of AMR. It also requires new, improved or alternative methods of treatment that will not worsen the situation. Thus, any antimicrobial treatment should be effective while not causing further development of resistance. Some antiseptics fall into this category and, in particular, polyhexamethylene hydrochloride biguanide (PHMB) has certain characteristics that make it an ideal solution to this problem of AMR, specifically within wound care applications. PHMB is a broad-spectrum antimicrobial that kills bacteria, fungi, parasites and certain viruses with a high therapeutic index, and is widely used in clinics, homes and industry. It has been used for many years and has not been shown to cause development of resistance; it is safe (non-cytotoxic), not causing damage to newly growing wound tissue. Importantly there is substantial evidence for its effective use in wound care applications, providing a sound basis for evidence-based practice. This review presents the evidence for the use of PHMB treatments in wound care and its alignment with AMS for the prevention and treatment of wound infection.

Effectiveness of a polyhexamethylene biguanide-containing wound cleansing solution using experimental biofilm models.

OBJECTIVE: Antiseptics are widely used in wound management to prevent or treat wound infections, and have been shown to have antibiofilm efficacy. The objective of this study was to assess the effectiveness of a polyhexamethylene biguanide (PHMB)-containing wound cleansing and irrigation solution on model biofilm of pathogens known to cause wound infections compared with a number of other antimicrobial wound cleansing and irrigation solutions. METHOD: Staphylococcus aureus and Pseudomonas aeruginosa single-species biofilms were cultured using microtitre plate and Centers for Disease Control and Prevention (CDC) biofilm reactor methods. Following a 24-hour incubation period, the biofilms were rinsed to remove planktonic microorganisms and then challenged with wound cleansing and irrigation solutions. Following incubation of the biofilms with a variety of concentrations of the test solutions (50%, 75% or 100%) for 20, 30, 40, 50 or 60 minutes, remaining viable organisms from the treated biofilms were quantified. RESULTS: The six antimicrobial wound cleansing and irrigation solutions used were all effective in eradicating Staphylococcus aureus biofilm bacteria in both test models. However, the results were more variable for the more tolerant Pseudomonas aeruginosa biofilm. Only one of the six solutions (sea salt and oxychlorite/NaOCl-containing solution) was able to eradicate Pseudomonas aeruginosa biofilm using the microtitre plate assay. Of the six solutions, three (a solution containing PHMB and poloxamer 188 surfactant, a solution containing hypochlorous acid (HOCl) and a solution containing NaOCl/HOCl) showed increasing levels of eradication of Pseudomonas aeruginosa biofilm microorganisms with increasing concentration and exposure time. Using the CDC biofilm reactor model, all six cleansing and irrigation solutions, except for the solution containing HOCl, were able to eradicate Pseudomonas aeruginosa biofilms such that no viable microorganisms were recovered. CONCLUSION: This study demonstrated that a PHMB-containing wound cleansing and irrigation solution was as effective as other antimicrobial wound irrigation solutions for antibiofilm efficacy. Together with the low toxicity, good safety profile and absence of any reported acquisition of bacterial resistance to PHMB, the antibiofilm effectiveness data support the alignment of this cleansing and irrigation solution with antimicrobial stewardship (AMS) strategies.

Cost-Effectiveness of PHMB & betaine wound bed preparation compared with standard care in venous leg ulcers: A cost-utility analysis in the United Kingdom.

BACKGROUND: Wounds cost £8.3 billion per year in the United Kingdom (UK) annually. Venous leg ulcers (VLUs) account for 15% of wounds and can be complicated to heal, increasing nurse visits and resource costs. Recent wound bed preparation consensus recommends wound cleansing and biofilm disrupting agents. However, inert cleansers such as tap water or saline are inexpensive, an evaluation of evidence is required to justify the higher upfront costs of treatment with active cleansers. We undertook a cost-effectiveness analysis of the use of a biofilm disrupting and cleansing solution and gel, Prontosan® Solution and Gel X, (PSGX) (B Braun Medical), as compared to the standard practice of using saline solution, for treating VLUs. METHODS: A Markov model was parameterised to one-year costs and health-related quality of life consequences of treating chronic VLUs with PSGX versus saline solution. Costs are viewed from a UK healthcare payer perspective, include routine care and management of complications. A systematic literature search was performed to inform the clinical parameters of the economic model. Deterministic univariate sensitivity analysis (DSA) and probabilistic sensitivity analysis (PSA) were undertaken. RESULTS: For PSGX an Incremental Net Monetary Benefit (INMB) of £1,129.65 to £1,042.39 per patient (with a Maximum Willingness to Pay of £30k and £20k per QALY respectively), of which cost savings are £867.87 and 0.0087 quality-adjusted life years (QALYs) gain per patient. PSA indicates a 99.3% probability of PSGX being cost-effective over saline. CONCLUSIONS: PSGX for the treatment of VLUs is dominant compared with saline solution in the UK with expected cost-savings within a year and improved patient outcomes.

Use of low-frequency contact ultrasonic debridement with and without polyhexamethylene biguanide in hard-to-heal leg ulcers: an RCT.

OBJECTIVE: The primary aim of this research was to investigate the combination effect of polyhexamethylene biguanide (PHMB) and low-frequency contact ultrasonic debridement (LFCUD) on the bacterial load in hard-to-heal wounds in adults, compared with ultrasonic debridement alone. Secondary outcomes included wound healing, quality of life (QoL) and pain scores. METHOD: In this single-blinded, randomised, controlled trial participants were randomised to two groups. All participants received LFCUD weekly for six weeks, plus six weeks of weekly follow-up. The intervention group received an additional 15-minute topical application of PHMB post-LFCUD, at each dressing change and in a sustained dressing product. The control group received non-antimicrobial products and the wounds were cleansed with clean water or saline. Wound swabs were taken from all wounds for microbiological analysis at weeks 1, 3, 6 and 12. RESULTS: A total of 50 participants took part. The intervention group (n=25) had a lower bacterial load at week 12 compared with the control group (n=25) (p<0.001). There was no difference in complete wound healing between the groups (p=0.47) or wound-related QoL (p=0.15). However, more wounds deteriorated in the control group (44%) compared with the intervention group (8%, p=0.01). A higher proportion of wounds reduced in size in the intervention group (61% versus 12%, p=0.019). Pain was lower in the intervention group at week six, compared with controls (p=0.04). CONCLUSION: LFCUD without the addition of an antimicrobial agent such as PHMB, cannot be recommended. Further research requires longer follow-up time and would benefit from being powered sufficiently to test the effects of multiple covariates.

Antibiofilm Properties of Antiseptic Agents Used on Pseudomonas aeruginosa Isolated from Diabetic Foot Ulcers.

In diabetic foot ulcers (DFUs), biofilm formation is a major challenge that promotes wound chronicity and delays healing. Antiseptics have been proposed to combat biofilms in the management of DFUs. However, there is limited evidence on the activity of these agents against biofilms, and there are questions as to which agents have the best efficiency. Here, we evaluated the antibiofilm activity of sodium hypochlorite, polyvinylpyrrolidoneIodine (PVPI), polyhexamethylenebiguanide (PHMB) and octenidine against Pseudomonas aeruginosa strains using static and dynamic systems in a chronic-wound-like medium (CWM) that mimics the chronic wound environment. Using Antibiofilmogram®, a technology assessing the ability of antiseptics to reduce the initial phase of biofilm formation, we observed the significant activity of antiseptics against biofilm formation by P. aeruginosa (at 1:40 to 1:8 dilutions). Moreover, 1:100 to 1:3 dilutions of the different antiseptics reduced mature biofilms formed after 72 h by 10-log, although higher concentrations were needed in CWM (1:40 to 1:2). Finally, in the BioFlux200TM model, after biofilm debridement, sodium hypochlorite and PHMB were the most effective antiseptics. In conclusion, our study showed that among the four antiseptics tested, sodium hypochlorite demonstrated the best antibiofilm activity against P. aeruginosa biofilms and represents an alternative in the management of DFUs.

The Saccharomyces cerevisiae Ncw2 protein works on the chitin/ß-glucan organisation of the cell wall.

The NCW2 gene was recently described as encoding a GPI-bounded protein that assists in the re-modelling of the Saccharomyces cerevisiae cell wall (CW) and in the repair of damage caused by the polyhexamethylene biguanide (PHMB) polymer to the cell wall. Its absence produces a re-organization of the CW structure that result in resistance to lysis by glucanase. Hence, the present study aimed to extend the analysis of the Ncw2 protein (Ncw2p) to determine its physiological role in the yeast cell surface. The results showed that Ncw2p is transported to the cell surface upon O-mannosylation mediated by the Pmt1p-Pmt2p enzyme complex. It co-localises with the yeast bud scars, a region in cell surface formed by chitin deposition. Once there, Ncw2p enables correct chitin/ß-glucan structuring during the exponential growth. The increase in molecular mass by hyper-mannosylation coincides with the increasing in chitin deposition, and leads to glucanase resistance. Treatment of the yeast cells with PHMB produced the same biological effects observed for the passage from exponential to stationary growth phase. This might be a possible mechanism of yeast protection against cationic biocides. In conclusion, we propose that Ncw2p takes part in the mechanism involved in the control of cell surface rigidity by aiding on the linkage between chitin and glucan layers in the modelling of the cell wall during cell growth.

Effect of Polyhexamethylene Biguanide in Combination with Undecylenamidopropyl Betaine or PslG on Biofilm Clearance.

Hospital-acquired infection is a great challenge for clinical treatment due to pathogens' biofilm formation and their antibiotic resistance. Here, we investigate the effect of antiseptic agent polyhexamethylene biguanide (PHMB) and undecylenamidopropyl betaine (UB) against biofilms of four pathogens that are often found in hospitals, including Gram-negative bacteria Pseudomonas aeruginosa and Escherichia coli, Gram-positive bacteria Staphylococcus aureus, and pathogenic fungus, Candida albicans. We show that 0.02% PHMB, which is 10-fold lower than the concentration of commercial products, has a strong inhibitory effect on the growth, initial attachment, and biofilm formation of all tested pathogens. PHMB can also disrupt the preformed biofilms of these pathogens. In contrast, 0.1% UB exhibits a mild inhibitory effect on biofilm formation of the four pathogens. This concentration inhibits the growth of S. aureus and C. albicans yet has no growth effect on P. aeruginosa or E. coli. UB only slightly enhances the anti-biofilm efficacy of PHMB on P. aeruginosa biofilms. However, pretreatment with PslG, a glycosyl hydrolase that can efficiently inhibit and disrupt P. aeruginosa biofilm, highly enhances the clearance effect of PHMB on P. aeruginosa biofilms. Meanwhile, PslG can also disassemble the preformed biofilms of the other three pathogens within 30 min to a similar extent as UB treatment for 24 h.

A new ophthalmic formulation containing antiseptics and dexpanthenol: In vitro antimicrobial activity and effects on corneal and conjunctival epithelial cells.

Antibiotic resistance is increasing even in ocular pathogens, therefore the interest towards antiseptics in Ophthalmology is growing. The aim of this study was to analyze the in vitro antimicrobial efficacy and the in vitro effects of an ophthalmic formulation containing hexamidine diisethionate 0.05%, polyhexamethylene biguanide (PHMB) 0.0001% disodium edetate (EDTA) 0.01%, dexpanthenol 5% and polyvinyl alcohol 1.25% (Keratosept, Bruschettini, Genova, Italy) on cultured human corneal and conjunctival cells. The in vitro antimicrobial activity was tested on Staphylococcus aureus, Methicillin-Resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa, Streptococcus pneumoniae, Streptococcus pyogenes and Streptococcus mitis. For each microbial strain 10 µL of a 0.5 MacFarland standardized bacterial inoculum were incubated at 25 °C with 100 µL of ophthalmic solution for up to 6 h. After different periods of time, samples were inoculated on blood agar with 5% sheep blood. Moreover, a 0.5 MacFarland bacterial inoculum was seeded in triplicate on Mueller-Hinton Agar or on Mueller-Hinton Fastidious Agar; then a cellulose disc soaked with 50 µL of ophthalmic solution was applied on the surface of agar and plates were incubated for 18 h at 37 °C, in order to evaluate the inhibition of bacterial growth around the disc. Human corneal and conjunctival epithelial cells in vitro were incubated for 5, 10 and 15 min with Keratosept or its components. The cytotoxicity was assessed through the release of cytoplasmic enzyme lactate dehydrogenase (LDH) into the medium immediately after exposure to the drugs; the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was performed to evaluate the metabolic cell activity. Our results show that Keratosept ophthalmic solution gave an average logarithmic (log) reduction of bacterial load of 2.14 ± 0.35 within 6 h of exposure (p-value < 0.05 versus control saline solution). On agar plates, all microbial strains, excluding P. Aeruginosa, showed an inhibition zone of growth around the Keratosept-soaked discs. Keratosept and its components after 5 and 10 min did not show any cytotoxic effect on cultured corneal and conjunctival cells, and only after 15 min a significant reduction of cell viability and an increase of cytotoxicity compared to control (vehicle) was seen; dexpanthenol 5% and polyvinyl alcohol accelerated the wounding of corneal cells in vitro. In conclusion, Keratosept showed good antimicrobial activity on the tested strains; the ophthalmic solution and its components were safe and non-toxic for the corneal and conjunctival epithelial cells for 5 and 10 min at the concentrations analyzed, and dexpanthenol 5% and polyvinyl alcohol promoted the wounding of corneal cells.

Wound bed preparation: a case series using polyhexanide and betaine solution and gel-a UK perspective.

OBJECTIVE: The burden of wound care within the NHS is estimated at a cost of £5.3 billion per year and is set to rise annually by 30%. This case series describes the results of using polyhexanide (PHMB) and betaine wound irrigation solution and gels (Prontosan, B.Braun Medical Ltd., UK) across the UK in hard-to-heal (also described as chronic) wounds up to 20 years' duration, with an observation period of greater than one month. Over half of the hard-to-heal wounds were healed and vast improvements to all other wounds were observed. Improvements to wound bed condition were reported as early as two days after commencing initial treatment, with decreases in malodour, exudate, slough and pain reported across the case series. In addition to wound bed improvements, a reduction in dressing change frequency of 55% was observed in hard-to-heal wounds under the new treatment regime.

Re-evaluation of polihexanide use in wound antisepsis in order to clarify ambiguities of two animal studies.

OBJECTIVE: Due to classification of the agent polihexanide (PHMB) in category 2 'may cause cancer' by the Committee for Risk Assessment of the European Chemicals Agency in 2011, the users of wound antiseptics may be highly confused. In 2017, this statement was updated, defining PHMB up to 0.1% as a preservative safe in all cosmetic products. In the interest of patient safety, a scientific clarification of the potential carcinogenicity of PHMB is necessary. METHODS: A multidisciplinary team (MDT) of microbiologists, surgeons, dermatologists and biochemists conducted a benefit-risk assessment to clarify the hazard of antiseptic use of PHMB. RESULTS: In two animal studies, from which the assessment of a carcinogenic risk was derived, PHMB was administered orally over two years in extremely high concentrations far above the NO(A)EL (no-observed-(adverse-) effect level) in rats and mice. Feeding in the NO(A)EL range resulted in no abnormal effects. In one male in the highest dose group of 4000ppm PHMB, an adenocarcinoma was found, which the author attributed to chronic inflammation of the colon with systemic atypical exposure. The increasing incidence of hemangiosarcomas highly probably resulted from increased endothelial proliferation, triggered by the exceedingly high dosage fed, because PHMB is not genotoxic and there is no evidence for epigenetic effects. DISCUSSION: It is well known that PHMB is not absorbed when applied topically. Considering the absence of genotoxicity and epigenetic effects together with the interpretation of the animal studies, it is the consensus of the multidisciplinary experts that a carcinogenic risk from PHMB-use for wound antisepsis can be ruled out. CONCLUSION: On this basis and considering their effectiveness, tolerability and clinical evidence, the indications for PHMB based wound antiseptics are justified.

Keratomycosis due to Tintelnotia destructans refractory to common therapy treated successfully with systemic and local terbinafine in combination with polyhexamethylene biguanide.

PURPOSE: To report on a wearer of rigid gas-permeable contact lenses with a keratomycosis due to Tintelnotia-a new genus of Phaeosphaeriaceae-treated with terbinafine and polyhexamethylene biguanide. METHODS: Chart review of a patient with fungal keratitis treated additionally with systemic and topical terbinafine 0.25% after symptoms increased under conventional antimycotic therapy with voriconazole. Antifungal susceptibility had been tested in vitro. RESULTS: After starting an additional treatment with systemic and topical terbinafine, the severe corneal infection was sufficiently resolved. The drug was well tolerated without any neurological, dermatological or gastroenterological problems. Terbinafine revealed a marked in vitro antifungal activity of 0.12 µg/ml. The fungus was identified as Tintelnotia destructans. CONCLUSIONS: Terbinafine might be considered as a therapeutic option in severe cases of fungal keratitis refractory to common antifungal therapy.

Polyhexamethylene biguanide and chloroquine induce programmed cell death in Acanthamoeba castellanii.

Several chemotherapeutic drugs have been described as amoebicidal agents acting against Acanthamoeba trophozoites and cysts. However, the underlying mechanism of action is poorly characterized. Here, we describe programmed cell death (PCD) in A. castellanii induced by polyhexamethylene biguanide (PHMB) and chloroquine. We used four types of amoebicidal agents including 0.02% PHMB, 0.02% chlorhexidine digluconate, 100 µM chloroquine, and 100 µM 2,6-dichlorobenzonitrile to kill Acanthamoeba trophozoites and cysts. Exposure to PHMB and chloroquine induced cell shrinkage and membrane blebbing in Acanthamoeba, observed microscopically. Externalization of phosphatidyl serine on the membranes of Acanthamoeba was detected by annexin V staining. Apoptotic cell death of Acanthamoeba by PHMB and chloroquine was confirmed by FACS analysis. Nuclear fragmentation of Acanthamoeba was demonstrated by DAPI staining. PHMB induced PCD in trophozoites and cysts, and chloroquine induced PCD in cysts. These findings are discussed to establish the most effective treatment for Acanthamoeba-induced keratitis.

Effect of poly-hexamethylene biguanide hydrochloride (PHMB) treated non-sterile medical gloves upon the transmission of Streptococcus pyogenes, carbapenem-resistant E. coli, MRSA and Klebsiella pneumoniae from contact surfaces.

BACKGROUND: Reduction of accidental contamination of the near-patient environment has potential to reduce acquisition of healthcare-associated infection(s). Although medical gloves should be removed when soiled or touching the environment, compliance is variable. The use of antimicrobial-impregnated medical gloves could reduce the horizontal-transfer of bacterial contamination between surfaces. AIM: Determine the activity of antimicrobial-impregnated gloves against common hospital pathogens: Streptococcus pyogenes, carbapenem-resistant E.coli (CREC), MRSA and ESBL-producing Klebsiella pneumoniae. METHODS: Fingerpads (~1cm2) of PHMB-treated and untreated gloves were inoculated with 10 µL (~104 colony-forming-units [cfu]) of test-bacteria prepared in heavy-soiling (0.5%BSA), blood or distilled-water (no-soiling) and sampled after 0.25, 1, 10 or 15 min contact-time. Donor surfaces (~1cm2 computer-keys) contaminated with wet/dry inoculum were touched with the fingerpad of treated/untreated gloves and subsequently pressed onto recipient (uncontaminated) computer-keys. RESULTS: Approximately 4.50log10cfu of all bacteria persisted after 15 min on untreated gloves regardless of soil-type. In the absence of soiling, PHMB-treated gloves reduced surface-contamination by ~4.5log10cfu (>99.99%) within 10 min of contact-time but only ~2.5log10 (>99.9%) and ~1.0log10 reduction respectively when heavy-soiling or blood was present. Gloves became highly-contaminated (~4.52log10-4.91log10cfu) when handling recently-contaminated computer-keys. Untreated gloves contaminated "recipient" surfaces (~4.5log10cfu) while PHMB-treated gloves transferred fewer bacteria (2.4-3.6log10cfu). When surface contamination was dry, PHMB gloves transferred fewer bacteria (0.3-0.6log10cfu) to "recipient" surfaces than untreated gloves (1.0-1.9log10; P < 0.05). CONCLUSIONS: Antimicrobial-impregnated gloves may be useful in preventing dissemination of organisms in the near-patient environment during routine care. However they are not a substitute for appropriate hand-hygiene procedures.

Effect of antiseptic irrigation on infection rates of traumatic soft tissue wounds: a longitudinal cohort study.

OBJECTIVE: Acute traumatic wounds are contaminated with bacteria and therefore an infection risk. Antiseptic wound irrigation before surgical intervention is routinely performed for contaminated wounds. However, a broad variety of different irrigation solutions are in use. The aim of this retrospective, non-randomised, controlled longitudinal cohort study was to assess the preventive effect of four different irrigation solutions before surgical treatment, on wound infection in traumatic soft tissue wounds. METHOD: Over a period of three decades, the prophylactic application of wound irrigation was studied in patients with contaminated traumatic wounds requiring surgical treatment, with or without primary wound closure. The main outcome measure was development of wound infection. From 1974-1983, either 0.04 % polihexanide (PHMB), 1 % povidone-iodine (PVP-I), 4 % hydrogen peroxide, or undiluted Ringer's solution were concurrently in use. From 1984-1996, only 0.04 % PHMB or 1 % PVP-I were applied. From 1997, 0.04 % PHMB was used until the end of the study period in 2005. RESULTS: The combined rate for superficial and deep wound infection was 1.7 % in the 0.04 % PHMB group (n=3264), 4.8 % in the 1 % PVP-I group (n=2552), 5.9 % in the Ringer's group (n=645), and 11.7 % in the 4 % hydrogen peroxide group (n=643). Compared with all other treatment arms, PHMB showed the highest efficacy in preventing infection in traumatic soft tissue wounds (p<0.001). However, compared with PVP-I, the difference was only significant for superficial infections. CONCLUSION: The large patient numbers in this study demonstrated a robust superiority of 0.04 % PHMB to prevent infection in traumatic soft tissue wounds. These retrospective results may further provide important information as the basis for power calculations for the urgently needed prospective clinical trials in the evolving field of wound antisepsis.

Preparation of nano cationic liposome as carrier membrane for polyhexamethylene biguanide chloride through various methods utilizing higher antibacterial activities with low cell toxicity.

This study suggested successful encapsulation of polyhexamethylene biguanide chloride (PHMB) into nano cationic liposome as a biocompatible antibacterial agent with less cytotoxicity and higher activities. Phosphatidylcholine, cholesterol and stearylamine were used to prepare nano cationic liposome using thin film hydration method along with sonication or homogeniser. Sonication was more effective in PHMB loaded nano cationic liposome preparation with smaller size (34 nm). FTIR, 1H NMR and XRD analyses were used to confirm the encapsulation of PHMB into nano cationic liposome. PHMB inclusion in nano cationic liposome was beneficial for increased antibacterial activity against Staphylococcus aureus and Escherichia coli. PHMB-loaded cationic liposome enables to deliver high concentrations of the antibacterial agent into the infectious cell. The cytotoxicity of PHMB entrapped in positively charged liposome was prominently reduced showing no significant visible detrimental effect on normal primary human skin fibroblast cell lines morphology confirming the effective role of cationic liposome encapsulation. Comparing with PHMB alone, encapsulation of PHMB in nano cationic liposome resulted in significant increase in cell viability from 2.4 to 63%.

Effectiveness of a polyhexanide irrigation solution on methicillin-resistant Staphylococcus aureus biofilms in a porcine wound model.

Irrigation and removal of necrotic debris can be beneficial for proper healing. It is becoming increasingly evident that wounds colonized with biofilm forming bacteria, such as Staphylococcus aureus (SA), can be more difficult to eradicate. Here we report our findings of the effects of an irrigation solution containing propyl-betaine and polyhexanide (PHMB) on methicillin-resistant Staphylococcus aureus (MRSA) biofilms in a porcine wound model. Thirty-nine deep partial thickness wounds were created with six wounds assigned to one of six treatment groups: (i) PHMB, (ii) Ringer's solution, (iii) hypochlorous acid/sodium hypochlorite, (iv) sterile water, (v) octenidine dihydrochloride, and (vi) octenilin. Wounds were inoculated with MRSA and covered with a polyurethane dressing for 24 hours to allow biofilm formation. The dressings were then removed and the wounds were irrigated twice daily for 3 days with the appropriate solution. MRSA from four wounds were recovered from each treatment group at 3 days and 6 days hours after initial treatment. Irrigation of wounds with the PHMB solution resulted in 97·85% and 99·64% reductions of MRSA at the respective 3 days and 6 days assessment times when compared to the untreated group. Both of these reductions were statistically significant compared to all other treatment groups (P values <0·05).