Successful control of Serratia marcescens outbreak in a neonatal unit of a tertiary-care hospital in Spain.

OBJECTIVE: Serratia marcescens is a Gram-negative bacterium that is found in hospital environments and commonly associated with outbreaks in neonatal units. One S. marcescens isolate was detected from a bloodstream culture from a neonate in our hospital that was followed by an outbreak. The aim of this study was to describe the molecular epidemiology of a S. marcescens outbreak in the neonatal unit. METHODS: In order to investigate the outbreak, weekly surveillance rectal swabs were submitted for culture from all patients admitted in this unit from August to September 2018. Environmental samples were obtained from potential sources in September 2018. Typing of isolates was performed by pulsed field gel electrophoresis (PFGE). In addition, we studied the in vitro activity of chlorhexidine against S. marcescens. RESULTS: During this period, 146 infants were hospitalised in our neonatal unit, of which 16 patients had a S. marcescens-positive sample. A total of 36 environmental surveillance samples were collected, and one sample from a stethoscope from an incubator of a colonized baby was positive for S. marcescens. All the 18 isolates, including the isolate from the stethoscope, belonged to a single PFGE cluster. We found that very low concentrations of chlorhexidine, even with application times close to 0 achieved significant reductions in the amount of S. marcescens. CONCLUSION: A unique clone of S. marcescens caused this outbreak, including isolates from patients and from one stethoscope. The outbreak was controlled with the early implementation of specific control measures.

Successful control of Serratia marcescens outbreak in a neonatal unit of a tertiary-care hospital in Spain / Control exitoso de un brote de Serratia marcescens en la unidad neonatal de un hospital terciario en España

ObjectiveSerratia marcescens is a Gram-negative bacterium that is found in hospital environments and commonly associated with outbreaks in neonatal units. One S. marcescens isolate was detected from a bloodstream culture from a neonate in our hospital that was followed by an outbreak. The aim of this study was to describe the molecular epidemiology of a S. marcescens outbreak in the neonatal unit.MethodsIn order to investigate the outbreak, weekly surveillance rectal swabs were submitted for culture from all patients admitted in this unit from August to September 2018. Environmental samples were obtained from potential sources in September 2018. Typing of isolates was performed by pulsed field gel electrophoresis (PFGE). In addition, we studied the in vitro activity of chlorhexidine against S. marcescens.ResultsDuring this period, 146 infants were hospitalised in our neonatal unit, of which 16 patients had a S. marcescens-positive sample. A total of 36 environmental surveillance samples were collected, and one sample from a stethoscope from an incubator of a colonized baby was positive for S. marcescens. All the 18 isolates, including the isolate from the stethoscope, belonged to a single PFGE cluster. We found that very low concentrations of chlorhexidine, even with application times close to 0 achieved significant reductions in the amount of S. marcescens.ConclusionA unique clone of S. marcescens caused this outbreak, including isolates from patients and from one stethoscope. The outbreak was controlled with the early implementation of specific control measures.

ObjetivoSerratia marcescens(S. marcescens) es una bacteria gramnegativa que se encuentra en ambientes hospitalarios y comúnmente aparece asociada a brotes en unidades neonatales. En agosto de 2018 se detectó un aislado de esta bacteria a partir de un hemocultivo de un paciente neonatal en nuestro hospital. El objetivo de este estudio fue describir la epidemiología molecular del brote de S. marcescens en la unidad neonatal.MétodosCon el fin de investigar el brote, se enviaron semanalmente para cultivo muestras rectales de todos los pacientes ingresados en estas unidades de agosto a septiembre de 2018. Asimismo, se obtuvieron muestras ambientales de potenciales orígenes del brote en septiembre de 2018. Los aislados se genotipificaron mediante electroforesis de campo pulsado (PFGE).ResultadosDurante este período, 146 lactantes fueron hospitalizados en nuestra unidad neonatal, de los cuales 16 pacientes tenían una muestra positiva para S. marcescens. Se recogieron un total de 36 muestras de vigilancia ambiental. Una muestra de un estetoscopio de una incubadora de un bebé colonizado resultó positiva para S. marcescens. Los 18 aislamientos, incluido el aislado de la muestra ambiental, pertenecían a un solo patrón de PFGE. Se realizaron experimentos in vitro con clorhexidina y se encontró que concentraciones muy bajas incluso con tiempos de aplicación cercanos a 0 lograban reducciones significativas en la cantidad de S. marcescens.ConclusiónEstos datos confirmaron un único clon de S. marcescens en la unidad neonatal con aislamientos tanto de pacientes como ambientales. La implementación temprana de medidas de control específicas fue eficaz para limitar la transmisión nosocomial.

In Vitro Effect of Photodynamic Therapy with Different Lights and Combined or Uncombined with Chlorhexidine on Candida spp.

Candidiasis is very common and complicated to treat in some cases due to increased resistance to antifungals. Antimicrobial photodynamic therapy (aPDT) is a promising alternative treatment. It is based on the principle that light of a specific wavelength activates a photosensitizer molecule resulting in the generation of reactive oxygen species that are able to kill pathogens. The aim here is the in vitro photoinactivation of three strains of Candida spp., Candida albicans ATCC 10231, Candida parapsilosis ATCC 22019 and Candida krusei ATCC 6258, using aPDT with different sources of irradiation and the photosensitizer methylene blue (MB), alone or in combination with chlorhexidine (CHX). Irradiation was carried out at a fluence of 18 J/cm2 with a light-emitting diode (LED) lamp emitting in red (625 nm) or a white metal halide lamp (WMH) that emits at broad-spectrum white light (420-700 nm). After the photodynamic treatment, the antimicrobial effect is evaluated by counting colony forming units (CFU). MB-aPDT produces a 6 log10 reduction in the number of CFU/100 µL of Candida spp., and the combination with CHX enhances the effect of photoinactivation (effect achieved with lower concentration of MB). Both lamps have similar efficiencies, but the WMH lamp is slightly more efficient. This work opens the doors to a possible clinical application of the combination for resistant or persistent forms of Candida infections.

Photodynamic Therapy Combined with Antibiotics or Antifungals against Microorganisms That Cause Skin and Soft Tissue Infections: A Planktonic and Biofilm Approach to Overcome Resistances.

The present review covers combination approaches of antimicrobial photodynamic therapy (aPDT) plus antibiotics or antifungals to attack bacteria and fungi in vitro (both planktonic and biofilm forms) focused on those microorganisms that cause infections in skin and soft tissues. The combination can prevent failure in the fight against these microorganisms: antimicrobial drugs can increase the susceptibility of microorganisms to aPDT and prevent the possibility of regrowth of those that were not inactivated during the irradiation; meanwhile, aPDT is effective regardless of the resistance pattern of the strain and their use does not contribute to the selection of antimicrobial resistance. Additive or synergistic antimicrobial effects in vitro are evaluated and the best combinations are presented. The use of combined treatment of aPDT with antimicrobials could help overcome the difficulty of fighting high level of resistance microorganisms and, as it is a multi-target approach, it could make the selection of resistant microorganisms more difficult.

Chalcogenide nanoparticles and organic photosensitizers for synergetic antimicrobial photodynamic therapy.

Synergistic antimicrobial effects were observed for copper sulfide (CuS) nanoparticles together with indocyanine green (ICG) in the elimination of wild type pathogenic bacteria (Staphylococcus aureus ATCC 29213 and Pseudomonas aeruginosa ATCC 27853) and also opportunistic fungal infective yeast (Candida albicans ATCC 10231). Furthermore, large antibacterial effects were observed for clinical isolates of Methicillin-resistant S. aureus (MRSA) PFGE strain-type USA300. This efficient antimicrobial action was attributed to the combined extra- and intracellular generation of reactive oxygen species upon light irradiation. Instead of the use of visible-light for the activation of common photosensitizers, both ICG and CuS nanoparticles can be activated in the near infrared (NIR)-region of the electromagnetic spectrum and therefore, superior tissue penetration would be expected in a potential elimination of pathogenic microorganisms not only on the skin but also in the soft tissue. In the different bacteria studied a 3-log reduction in the bacterial counts was achieved after only 6 min of NIR irradiation and treatment with ICG or CuS alone at concentrations of 40 and 160 µg mL-1, respectively. A maximum bactericidal effect against S. aureus and USA300 strains was obtained for the combination of both photosensitizers at the same concentration. Regarding P. aeruginosa, a 4-log reduction in the CFU was observed for the combination of CuS and ICG at various concentrations. In Candida albicans the combination of both ICG and CuS and light irradiation showed an antimicrobial dose-dependent effect with the reduction of at least 3-log in the cell counts for the combination of ICG + CuS at reduced concentrations. The observed antimicrobial effect was solely attributed to a photodynamic effect and any photothermal effect was avoided to discard any potential thermal injury in a potential clinical application. The generation of reactive oxygen species upon near infrared-light irradiation for those photosensitizers used was measured either alone or in combination. The cytocompatibility of the proposed materials at the doses used in photodynamic therapy was also demonstrated in human dermal fibroblasts and keratinocytes by cell culturing and flow cytometry studies.

Successful control of Serratia marcescens outbreak in a neonatal unit of a tertiary-care hospital in Spain.

OBJECTIVE: Serratia marcescens is a Gram-negative bacterium that is found in hospital environments and commonly associated with outbreaks in neonatal units. One S. marcescens isolate was detected from a bloodstream culture from a neonate in our hospital that was followed by an outbreak. The aim of this study was to describe the molecular epidemiology of a S. marcescens outbreak in the neonatal unit. METHODS: In order to investigate the outbreak, weekly surveillance rectal swabs were submitted for culture from all patients admitted in this unit from August to September 2018. Environmental samples were obtained from potential sources in September 2018. Typing of isolates was performed by pulsed field gel electrophoresis (PFGE). In addition, we studied the in vitro activity of chlorhexidine against S. marcescens. RESULTS: During this period, 146 infants were hospitalised in our neonatal unit, of which 16 patients had a S. marcescens-positive sample. A total of 36 environmental surveillance samples were collected, and one sample from a stethoscope from an incubator of a colonized baby was positive for S. marcescens. All the 18 isolates, including the isolate from the stethoscope, belonged to a single PFGE cluster. We found that very low concentrations of chlorhexidine, even with application times close to 0 achieved significant reductions in the amount of S. marcescens. CONCLUSION: A unique clone of S. marcescens caused this outbreak, including isolates from patients and from one stethoscope. The outbreak was controlled with the early implementation of specific control measures.

Comparison of Antibacterial Activity and Wound Healing in a Superficial Abrasion Mouse Model of Staphylococcus aureus Skin Infection Using Photodynamic Therapy Based on Methylene Blue or Mupirocin or Both.

Background: Antibiotic resistance and impaired wound healing are major concerns in S. aureus superficial skin infections, and new therapies are needed. Antimicrobial photodynamic therapy (aPDT) is a new therapeutic approach for infections, but it also improves healing in many wound models. Objective: To compare the antimicrobial activity and the effects on wound healing of aPDT based on Methylene Blue (MB-aPDT) with mupirocin treatment, either alone or in combination, in superficial skin wounds of S. aureus-infected mice. Additionally, to evaluate the clinical, microbiological, and cosmetic effects on wound healing. Materials and Methods: A superficial skin infection model of S. aureus was established in SKH-1 mice. Infected wounds were treated with MB-aPDT, MB-aPDT with a daily topical mupirocin or only with mupirocin. No treatment was carried out in control animals. Daily clinical and microbiological examinations were performed until complete clinical wound healing. Histopathological studies and statistical analysis were performed at the end of the study. Results: MB-aPDT treatment induced the best wound healing compared to mupirocin alone or to mupirocin plus MB-aPDT. Superficial contraction at 24 h and a greater reduction in size at 48 h, quicker detachment of the crust, less scaling, and absence of scars were observed. Histopathological studies correlated with clinical and gross findings. By contrast, mupirocin showed the highest logaritmic reduction of S. aureus. Conclusions: MB-aPDT and mupirocin treatments are effective in a murine superficial skin infection model of S. aureus. One session of MB-aPDT was the best option for clinical wound healing and cosmetic results. The addition of mupirocin to MB-aPDT treatment improved antimicrobial activity; however, it did not enhance wound healing. No synergistic antibacterial effects were detected.

Broad-Spectrum Photo-Antimicrobial Polymers Based on Cationic Polystyrene and Rose Bengal.

New strategies to fight bacteria and fungi are necessary in view of the problem of iatrogenic and nosocomial infections combined with the growing threat of increased antimicrobial resistance. Recently, our group has prepared and described two new readily available materials based on the combination of Rose Bengal (singlet oxygen photosensitizer) and commercially available cationic polystyrene (macroporous resin Amberlite® IRA 900 or gel-type resin IRA 400). These materials showed high efficacy in the antimicrobial photodynamic inactivation (aPDI) of Pseudomonas aeruginosa. Here, we present the photobactericidal effect of these polymers against an extended group of pathogens like Escherichia coli, Enterococcus faecalis, Staphylococcus aureus, and the opportunistic yeast Candida albicans using green light. The most interesting finding is that the studied materials are able to reduce the population of both Gram-positive and Gram-negative bacteria with good activity, although, for C. albicans, in a moderate manner. In view of the results achieved and especially considering the inexpensiveness of these two types of photoactive polymers, we believe that they could be used as the starting point for the development of coatings for self-disinfecting surfaces.

A cost-effective combination of Rose Bengal and off-the-shelf cationic polystyrene for the photodynamic inactivation of Pseudomonas aeruginosa.

Two new photoactive materials have been prepared, characterized and tested against Pseudomonas aeruginosa bacteria (planktonic suspension). The synthesis of the polymeric photosensitizers can be made at a multigram scale, in few minutes, starting from inexpensive and readily available materials, such as Rose Bengal (photosensitizer) and ion exchange resins Amberlite® IRA 900 (macroporous) or IRA 400 (gel-type) as cationic polystyrene supports. The most notable feature of these systems is their notable bactericidal activity in the dark (4-5 log10 CFU/mL reduction of the population of P. aeruginosa) which becomes enhanced upon irradiation with visible light (to reach a total reduction of 8 log10 CFU/mL for the macroporous polymer at a fluence of 120 J/cm2 using green light of 515 nm).

Photodynamic therapy using methylene blue, combined or not with gentamicin, against Staphylococcus aureus and Pseudomonas aeruginosa.

Antimicrobial photodynamic therapy (a-PDT), combined or not with antibiotics, constitutes a promising therapy for superficial infections caused by bacteria implicated in multidrug resistance processes. We compared the efficacy of aPDT using the photosensitizer methylene blue (MB), combined or not with the antibiotic gentamicin (GN), against Staphylococcus aureus and Pseudomonas aeruginosa. Different concentrations of MB (0.03-7000 µg/mL), with or without GN (1-20 µg/mL), were added to planktonic cultures or biofilms and the samples irradiated with a LED lamp (λ 625 nm, 7 mW/cm2, 18 J/cm2). The number of viable bacteria in the samples and in corresponding nonirradiated controls was quantified by counting colony-forming units to evaluate the individual effects of MB, GN, and irradiation. MB-aPDT resulted in significant bacterial photoinactivation. The combination of GN and MB-aPDT exerted a synergistic bactericidal effect against planktonic cultures of S. aureus and P. aeruginosa. This combination did not significantly alter the photoinactivating effect of MB against S. aureus biofilms, but exerted a positive bactericidal effect against P. aeruginosa biofilms. These results underscore the need for further clinical studies of this therapeutic combination for the management of difficult-to-treat skin and mucous infections, especially those caused by P. aeruginosa.

A combination of photodynamic therapy and antimicrobial compounds to treat skin and mucosal infections: a systematic review.

BACKGROUND: Antimicrobial photodynamic therapy (aPDT) is a growing approach to treat skin and mucosal infections. Despite its effectiveness, investigators have explored whether aPDT can be further combined with antibiotics and antifungal drugs. OBJECTIVE: To systematically assess the in vivo studies on the effectiveness of combinations of aPTD plus antimicrobials in the treatment of cutaneous and mucosal infections. MATERIALS AND METHODS: Searches were performed in four databases (PubMed, EMBASE, Cochrane library databases, ClinicaTrials.gov) until July 2018. The pooled information was evaluated according to the PRISMA guidelines. RESULTS: 11 full-text articles were finally evaluated and included. The best aPDT combinations involved 5-aminolevulinic acid or phenothiazinium dye-based aPDT. In general, the combination shows benefits such as reducing treatment times, lowering drug dosages, decreasing drug toxicity, improving patient compliance and diminishing the risk of developing resistance. The mechanism of action may be that first aPDT damages the microbial cell wall or membrane, which allows better penetration of the antimicrobial drug. LIMITATIONS: The number of studies was low, the protocols used were heterogeneous, and there was a lack of clinical trials. CONCLUSIONS: The additive or synergistic effect of aPDT combined with antimicrobials could be promising to manage skin and mucosal infections, helping to overcome the microbial drug resistance.

Antimicrobial effects of photodynamic therapy.

The microorganisms that cause infections are increasing their resistance to antibiotics. In this context, alternative treatments are necessary. The antimicrobial photodynamic therapy (aPDT) is a therapeutic modality based on photosensitizing molecules that end up generating reactive oxygen species that induce the destruction of the target cells when are irradiated with light of a suitable wavelength and at a proper dose. The cells targeted by aPDT are all types of microorganisms (bacteria, fungi and parasites) including viruses and has been proven effective against representative members of all of them. In the field of dermatology, aPDT has been tested with promising results in different infections such as chronic ulcers, acne, onychomycosis and other cutaneous mycoses, as well as in leishmaniasis. Therefore, it is presented as a possible treatment option against the agents that cause skin and/or mucous infections.

Antimicrobial photodynamic activity of Rose Bengal, alone or in combination with Gentamicin, against planktonic and biofilm Staphylococcus aureus.

Antimicrobial photodynamic therapy (aPDT) could constitute an alternative therapy to antibiotics especially against superficial infections caused by bacteria involved in multidrug resistance processes. The aim of this study is to compare the efficacy of aPDT using the photosensitizer rose bengal (RB), combined or uncombined with gentamicin (GN), against Staphylococcus aureus. Different concentrations of RB (ranging from 0.03 to 64 µg/ml) were added to S. aureus in water suspensions or forming biofilms in the absence or presence of GN (1-40 µg/ml) and the samples were irradiated (18 or 37 J/cm2). The number of viable bacteria was quantified by counting colony-forming units. RB-aPDT shows significant photoactivity. The combination of GN and RB-aPDT exerts a synergistic bactericidal effect against planktonic S. aureus. On the other hand, a synergistic effect is observed only when the maximum concentration tested of RB and GN was used in biofilm. According to these result the use of RB-aPDT alone or in combination with GN could be implemented against S. aureus.

Bactericidal Effect of Photodynamic Therapy, Alone or in Combination with Mupirocin or Linezolid, on Staphylococcus aureus.

Antibiotic treatments frequently fail due to the development of antibiotic resistance, underscoring the need for new treatment strategies. Antimicrobial photodynamic therapy (aPDT) could constitute an alternative therapy. In bacterial suspensions of Staphylococcus aureus, which is commonly implicated in cutaneous and mucosal infections, we evaluated the in vitro efficacy of aPDT, using the photosensitizing agents rose bengal (RB) or methylene blue (MB), alone or combined with the antibiotics mupirocin (MU) or linezolid (LN). RB or MB, at concentrations ranging from 0.03 to 10 µg/ml, were added to S. aureus ATCC 29213 suspensions containing >108 cells/ml, in the absence or presence of MU or LN (1 or 10 µg/ml). Suspensions were irradiated with a white metal halide (λ 420-700 nm) or light-emitting diode lamp (λ 515 and λ 625 nm), and the number of viable bacteria quantified by counting colony-forming units (CFU) on blood agar. Addition of either antibiotic had no significant effect on the number of CFU/ml. By contrast, RB-aPDT and MB-aPDT effectively inactivated S. aureus, as evidenced by a 6 log10 reduction in bacterial growth. In the presence of MU or LN, the same 6 log10 reduction was observed in response to aPDT, but was achieved using significantly lower concentrations of the photosensitizers RB or MB. In conclusion, the combination of MU or LN and RB/MB-aPDT appears to exert a synergistic bactericidal effect against S. aureus in vitro.

Comparative effect of photodynamic therapy on separated or mixed cultures of Streptococcus mutans and Streptococcus sanguinis.

Antimicrobial photodynamic therapy (aPDT) has shown to exert a bactericidal effect against Streptococcus sanguinis and Streptococcus mutans. However, this efficacy has been reported for either type of bacteria separately. Bacterial suspensions of both strains, separately or together, were treated with concentrations of methylene blue (MB) and rose bengal (RB). Suspensions were irradiated with a light-emitting diode lamp (λ center at 625nm for MB and λ center at 515nm for RB) using a fluence of 18J/cm2. RB-aPDT at concentrations of 0.16-0.62 and 0.16-0.31µg/mL, and MB-aPDT at concentrations of 0.62-1.25 and 0.31-1.25µg/mL inhibited the growth of S. mutans and S. sanguinis respectively by 6 log10. In suspensions of both strains together, the same 6 log10 reduction in bacterial growth was achieved using the same concentrations of each photosensiziser. In conclusion, RB-aPDT and MB-aPDT appear to exert the same bactericidal effect against suspensions of S. sanguinis and S. mutans either for single strain treatment or for samples constituted by both bacteria mixed together. RB shows to be slightly more efficient than MB.

Staphylococcus pseudintermedius Human Infection Cases in Spain: Dog-to-Human Transmission.

Staphylococcus pseudintermedius is an opportunistic pathogen that has been identified as infectious agent or colonizer mainly in dogs. S. pseudintermedius has been also detected in humans and more specifically in people in contact with dogs. In this study, the possible S. pseudintermedius pet-to-human transmission was analyzed in four clinical human cases. Two patients were dog owners and S. pseudintermedius was also detected as colonizer in these healthy animals. S. pseudintermedius isolates from patients and dogs of the same household showed identical pulsed-field gel electrophoresis patterns, sequence types (STs), and antimicrobial resistance phenotypes and genotypes, and were methicillin susceptible. Resistance to erythromycin, clindamycin, tetracycline, trimetoprim-sulfamethoxazole, and/or ciprofloxacin was identified among S. pseudintermedius strains. The lineages ST241 and the new ST521 were detected in the strains of the two dog-owner patients, respectively. The strains from the other two patients presented two new STs, ST719 and ST720. To our knowledge, this is the first description of human infections caused by S. pseudintermedius in Spain.

Superior performance of macroporous over gel type polystyrene as a support for the development of photo-bactericidal materials.

A hexanuclear molybdenum cluster [Mo6I8Ac6]2- (1) has been ionically bound onto macroporous (Pmp) and gel-type (Pgel) resins and their performance as materials for the photodynamic inactivation of microorganisms has been studied. It has been found that 1@Pmp in combination with light is able to reduce 99.999999% of the population of Gram-positive Staphylococcus aureus whereas the activity of 1@Pgel is limited to a 99.99% reduction at the same light dose. The same trend is observed with Gram-negative Pseudomonas aeruginosa. A comprehensive study of both materials has been performed using confocal laser scanning microscopy, thermogravimetric analysis, nitrogen porosimetry, steady state and time resolved fluorometries and diffuse reflectance spectroscopy. The photochemical generation of singlet oxygen (1O2) has been assessed using 9,10-dimethylanthracene as a trap for this reactive oxygen species. It can be concluded that the nature of the polymeric support is of paramount importance for the development of surfaces with bactericidal properties.

A photobleaching resistant polymer supported hexanuclear molybdenum iodide cluster for photocatalytic oxygenations and photodynamic inactivation of Staphylococcus aureus.

The ability of a hexanuclear molybdenum cluster, [Mo6I8(CH3COO)6]2-, to carry out, upon irradiation, singlet oxygen mediated photocatalytic oxygenation reactions has been tested. This complex has been also supported on a solid polymeric matrix, yielding an immobilized photosensitizer with remarkable photostability and recyclability. Preliminary studies of antibacterial photoinactivation of Staphylococcus aureus have been conducted, with positive results.