The potential of therapeutic hyperthermia to eradicate Staphylococcus aureus bacteria; an in vitro study.

Staphylococcus aureus is one of the most common infectious agents, causing morbidity and mortality worldwide. Most pathogenic bacteria are classified in the group of mesophilic bacteria and the optimal growth temperature of these bacteria changes between 33 and 41 °C. Increased temperature can inhibit bacterial growth and mobility, which in turn, can trigger autolysis and cause cell wall damage. Hyperthermia treatment is defined as a heat-mediated treatment method applied using temperatures higher than body temperature. Nowadays, this treatment method is used especially in the treatment of tumours. Hyperthermia treatment is divided into two groups: mild hyperthermia and ablative or high-temperature hyperthermia. Mild hyperthermia is a therapeutic technique in which tumour tissue is heated above body temperature to produce a physiological or biological effect but is often not aimed at directly causing significant cell death. The goal of this method is to achieve temperatures of 40-45 °C in human tissues for up to 2 h. Hyperthermia can be used in the treatment of infections caused by such bacterial pathogens. In addition, using hyperthermia in combination with antimicrobial drugs may result in synergistic effects and reduce resistance issues. In our study, we used two different temperature levels (37 °C and 45 °C). We assessed growth inhibition, some virulence factors, alteration colony morphologies, and antimicrobial susceptibility for several antibiotics with three methods (Kirby-Bauer, E-test and broth microdilution) under hyperthermia. In the study, we observed that hyperthermia affected the urease enzyme, antibiotic sensitivity levels showed synergy with hyperthermia, and changes occurred in colony diameters and affected bacterial growth. We hypothesise that hyperthermia might be a new therapeutic option for infectious diseases as a sole agent or in combination with different antimicrobials.

In Vitro and Ex Vivo Investigation of the Antibacterial Effects of Methylene Blue against Methicillin-Resistant Staphylococcus aureus.

Methylene blue (MB) is a water-soluble dye that has a number of medical applications. Methicillin-resistant Staphylococcus aureus (MRSA) was selected as a subject for research due to the numerous serious clinical diseases it might cause and because there is a significant global resistance challenge. Our main goal was to determine and analyze the antibacterial effects of MB against S. aureus both in vitro and ex vivo to enhance treatment options. A total of 104 MRSA isolates recovered from various clinical specimens were included in this study. Minimum inhibitory concentration (MIC) values of MB against MRSA isolates were determined by the agar dilution method. One randomly selected MRSA isolate and a methicillin-susceptible S. aureus strain (S. aureus ATCC 25923) were employed for further evaluation of the antibacterial effects of MB in in vitro and ex vivo time-kill assays. A disc diffusion method-based MB + antibiotic synergy assay was performed to analyze the subinhibitory effects of MB on ten isolates. MICs of MB against 104 MRSA isolates, detected by the agar dilution method, ranged between 16 and 64 µg/mL. MB concentrations of 4 and 16 µg/mL showed a bactericidal effect at 24 h in the ex vivo time-kill assays and in vitro time-kill assays, respectively. We observed a significant synergy between cefoxitin and methylene blue at a concentration of 1-2 µg/mL in two (20%) test isolates. Employing MB, which has well-defined pharmacokinetics, bioavailability, and safety profiles, for the treatment of MRSA infections and nasal decolonization could be a good strategy.

In vitro effects of deferoxamine on antibiotic susceptibility in Gram-negative bacteria.

BACKGROUND: Iron is a vital element for the growth of bacteria. Bacteria use several strategies to scavenge iron, such as siderophores, which are thought to be important virulence components. The mammalian host uses various iron-binding substances to make iron unavailable for bacterial uptake. Deferoxamine (DFO) is a semi-synthetic iron chelator that has been licensed for medical use. Iron chelators like DFO may provide an alternative therapeutic technique for treating Gram-negative bacteria infections, which frequently display multidrug resistance. OBJECTIVES: We assumed that iron deprivation or interactions with the cell membrane caused by DFO or increased siderophore synthesis may cause the inhibition or inactivation of proteins and enzymes necessary for critical processes in bacteria. Additionally, we proposed that these bacterial alterations might be the origin of synergistic interactions between DFO and several antibiotics. MATERIAL AND METHODS: To test this hypothesis, we used disc diffusion, broth microdilution and checkerboard synergy testing methods on combinations of DFO with ceftriaxone, cefepime, meropenem, amikacin, levofloxacin, and tigecycline, respectively, in a total of 55 isolates (Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii, and Proteus mirabilis strains - 11 isolates for each genus). RESULTS: No synergistic or antagonistic interactions were observed between DFO and the tested antibiotics in the E. coli, K. pneumoniae, P. aeruginosa, and A. baumannii isolates, while the addition of DFO significantly increased the inhibition zone diameters of cefepime, amikacin, meropenem, tigecycline, and levofloxacin in P. mirabilis isolates. According to the checkerboard synergy results, a synergistic interaction was found between DFO and tigecycline, cefepime and amikacin for P. mirabilis isolates. CONCLUSIONS: Among the investigated bacteria, a synergy between antibiotics and DFO was only discovered against P. mirabilis. We do not believe that this entirely disproves our hypothesis, though. The production of siderophores triggered by the increased metabolic activity of actively proliferating bacteria at the infection site may provide better results. Therefore, expanding these investigations and developing infection models through animal testing would be advantageous.

Effects of deferoxamine on intrinsic colistin resistance of Proteus mirabilis.

Proteus mirabilis is a common pathogen, which is responsible for urinary tract infections. Iron is a critical element necessary for both humans and pathogens to maintain their biological functions, and iron limitation via chelator agents may be useful in the treatment of infections. The present study aimed to investigate the synergistic interactions between the iron chelator agent deferoxamine (DFO) and the antibacterial drug colistin. The minimum inhibitory concentration (MIC) values of DFO and colistin for P. mirabilis isolates were determined by broth microdilution. The checkerboard technique was used to examine the potential synergy between DFO and colistin. Furthermore, time-kill assays were used for the confirmation of synergy detected by the checkerboard assay, as well as for determining bacteriostatic and bactericidal interactions throughout a 24-h period. As expected, all P. mirabilis isolates were resistant to colistin. DFO did not inhibit P. mirabilis growth when used alone, even at very high doses (10 µg ml-1). Notably, when in combination with DFO, the MIC values of colistin were markedly reduced, and the checkerboard assay results showed synergy between colistin and DFO for all isolates. In addition, in time-kill assays, colistin + DFO exhibited synergistic activity against all strains at most time intervals and concentrations tested. Colistin + DFO showed bactericidal activity at colistin concentrations of 1xMIC and 2xMIC, although a degree of re-growth was observed in one of the strains at 12-24 h. These findings indicated that DFO has the potential for use as an adjunct to colistin through iron sequestration, thus providing synergistic activity to an antibiotic that would not normally be considered a treatment option against P. mirabilis. In vivo experiments in the future may provide useful information on the efficacy of DFO/colistin since these models effectively reflect physiological parameters.

An epidemiologic analysis of Candida spp. urinary infections in intensive care unit / Uma análise epidemiológica das infecções urinárias por Candida spp. em unidade de terapia intensiva / Un análisis epidemiológico de las infecciones urinarias por Candida spp. en la unidad de cuidados intensivos

Background and objectives: The finding of Candida species in urine is an usual finding and is called candiduria. There is an increase in the frequency of urinary tract infections (UTI) caused by Candida especially in critically ill patients. This study aimed to determine the epidemiological, clinical, and mycological characteristics of Candida urinary infections in intensive care unit (ICU) and antifungal susceptibilities. Methods: Urine cultures of 394 ICU patients with clinical suspicion of UTI were evaluated. After 24-48 hours of incubation, colonies appeared to grow as yeast, were morphologically examined by Gram staining. Candida strains that grew 104 ≥ CFU/mL in urine cultures were accepted as candiduria. The susceptibilities of the Candida strains to amphotericin B, itraconazole, fluconazole, voriconazole, flucytosine, and caspofungin were investigated with broth microdilution method. Results: The distribution of the isolated 100 urinary Candida strains were as, 54 Candida albicans, 34 C. glabrata, 7 C. tropicalis, 2 C. kefyr, 2 C. lusitaniae, and 1 as C. parapsilosis. Among 100 Candida species isolated in our study susceptibility rates of amphotericin B, flucytosine, caspofungin, fluconazole, itraconazole, and voriconazole were 100%, 100%, 91%, 23%, 13%, 25.8%, respectively. Conclusion: Accurate identification of Candida spp., as well as the investigating the antifungal susceptibility, will be beneficial in terms of the effectiveness of the treatment and the prevention of resistance development.(AU)

Justificativa e objetivos: O achado de espécies de Candida na urina é um achado comum e é chamado de candidúria. Há um aumento na frequência de infecções do trato urinário (ITU) causadas por Candida, principalmente em pacientes críticos. Este estudo teve como objetivo determinar as características epidemiológicas, clínicas e micológicas das infecções urinárias por Candida em unidade de terapia intensiva (UTI) e a susceptibilidade aos antifúngicos. Métodos: Foram avaliadas culturas de urina de 394 pacientes de UTI com suspeita clínica de ITU. Após 24-48 horas de incubação, as colônias pareceram crescer como leveduras, foram morfologicamente examinadas por coloração de Gram. As cepas de Candida que cresceram ≥104 UFC/mL em culturas de urina foram aceitas como candidúria. As suscetibilidades das cepas de Candida à anfotericina B, itraconazol, fluconazol, voriconazol, flucitosina e caspofungina foram investigadas com o método de microdiluição em caldo. Resultados: A distribuição das cepas 100 isoladas de Candida urinária foi de 54 Candida albicans, 34 C. glabrata, 7 C. tropicalis, 2 C. kefyr, 2 C. lusitaniae e 1 como C. parapsilosis. Entre 100 espécies de Candida isoladas em nosso estudo, as taxas de susceptibilidade de anfotericina B, flucitosina, caspofungina, fluconazol, itraconazol e voriconazol foram de 100%, 100%, 91%, 23%, 13%, 25,8%, respectivamente. Conclusão: A identificação precisa de Candida spp., bem como a investigação da susceptibilidade aos antifúngicos, será benéfica em termos de eficácia do tratamento e prevenção do desenvolvimento de resistência.(AU)

Justificación y objetivos: El hallazgo de especies de Candida en la orina es un hallazgo habitual y se denomina candiduria. Hay un aumento en la frecuencia de infecciones del tracto urinario (ITU) causadas por Candida, especialmente en pacientes críticamente enfermos. Este estudio tuvo como objetivo determinar las características epidemiológicas, clínicas y micológicas de las infecciones urinarias por Candida en la unidad de cuidados intensivos (UCI) y la susceptibilidad antifúngica. Métodos: Se evaluaron urocultivos de 394 pacientes de UCI con sospecha clínica de ITU. Después de 24-48 horas de incubación, las colonias parecían crecer como levadura, se examinaron morfológicamente mediante tinción de Gram. Las cepas de Candida que crecieron 104 ≥ UFC / ml en urocultivos se aceptaron como candiduria. Las susceptibilidades de las cepas de Candida a la anfotericina B, itraconazol, fluconazol, voriconazol, flucitosina y caspofungina se investigaron con el método de microdilución en caldo. Resultados: La distribución de las cepas 100 urinarias aisladas de Candida fue de, 54 C. albicans, 34 C. glabrata, 7 C. tropicalis, 2 C. kefyr, 2 C. lusitaniae y 1 como C. parapsilosis. Entre las 100 especies de Candida aisladas en nuestro estudio, las tasas de susceptibilidad de anfotericina B, flucitosina, caspofungina, fluconazol, itraconazol y voriconazol fueron 100%, 100%, 91%, 23%, 13%, 25,8%, respectivamente. Conclusión: La identificación precisa de Candida spp., así como la investigación de la susceptibilidad antifúngica, será beneficiosa en términos de la eficacia del tratamiento y la prevención del desarrollo de resistencias.(AU)

Rhodotorula fungemia in a patient with acute lymphoblastic leukemia / Fungemia de Rhodotorula em paciente com leucemia linfoblástica aguda / Fungemia por Rhodotorula en un paciente con leucemia linfoblástica aguda

Objectives: Rhodotorula is an environmental yeast that belongs to Basidiomycota Phylum. Rhodotorula species are ubiquitous in nature, can be found in soil and freshwater. Immunocompromised patients can develop Rhodotorulosis due to wide-ranging exposure to Rhodotorula in the hospital environment. Case Discussion: The patient was a 3-year-old male with a diagnosis of Pro B-Acute Lymphoblastic Leukemia (ALL). He was admitted to the hospital with complaints of malaise, fatigue, weight loss, and diarrhea between courses of chemotherapy. Rhodotorula was isolated from the patient's blood culture obtained during the elevation of temperature. After 14 days of amphotericin B treatment, clinical situation of the patient was improved and he was discharged. Conclusion: Rhodotorula spp. as a rare yet emerging pathogen, often presents as fever of unknown etiology resistant to antibacterial treatment and can be associated with fungemia and other severe complications.(AU)

Objetivos: Rhodotorula é uma levedura ambiental que pertence ao filo Basidiomycota. As espécies de Rhodotorula são onipresentes na natureza, podem ser encontradas no solo e na água doce. Pacientes imunocomprometidos podem desenvolver rodotorulose devido à ampla exposição a Rhodotorula no ambiente hospitalar. Descrição do caso: O paciente era uma criança de 3 anos de idade com diagnóstico de Leucemia Linfoblástica Aguda Pro B (LLA). O paciente deu entrada no hospital com queixas de mal-estar, cansaço, perda de peso e diarreia entre os ciclos de quimioterapia. A Rhodotorula foi isolada da hemocultura do paciente obtida durante a elevação da temperatura. Após 14 dias de tratamento com anfotericina B, a situação clínica do paciente melhorou e o paciente recebeu alta. Conclusão: Rhodotorula spp. como um patógeno raro, porém emergente, frequentemente se apresenta como febre de etiologia desconhecida resistente ao tratamento antibacteriano e pode estar associada a fungemia e outras complicações graves.(AU)

Objetivos: Rhodotorula es una levadura ambiental que pertenece al filo Basidiomycota. Las especies de Rhodotorula son ubicuas en la naturaleza, se pueden encontrar en el suelo y en agua dulce. Los pacientes inmunodeprimidos pueden desarrollar Rhodotorulosis debido a una amplia exposición a Rhodotorula en el entorno hospitalario. Descripción del caso: El paciente era un niño de 3 años con diagnóstico de leucemia linfoblástica aguda Pro B (LLA). El paciente ingresó en el hospital con quejas de malestar, fatiga, pérdida de peso y diarrea entre ciclos de quimioterapia. Se aisló Rhodotorula del hemocultivo del paciente que se obtuvo durante la elevación de la temperatura. Después de 14 días de tratamiento con anfotericina B, la situación clínica del paciente mejoró y fue dado de alta. Conclusión: Rhodotorula spp. como patógeno poco común pero emergente, a menudo se presenta como fiebre de etiología desconocida resistente al tratamiento antibacteriano y puede asociarse con fungemia y otras complicaciones graves.(AU)

Sphingomonas paucimobilis Outbreak a Pediatric Hematology-Oncology Hospital: Epidemiological Investigation and Literature Review of an Emerging Healthcare-Associated Infection.

Sphingomonas paucimobilis is an aerobic, non-fermentative, opportunistic Gram-negative bacillus found in water systems. This study was conducted to analyze concurrent S. paucimobilis bacteremia cases and treatment outcomes, potential outbreak sources, and antimicrobial resistance profiles. This ambidirectional cohort study was conducted in a 30-bed pediatric hematology-oncology hospital. The patients' ages ranged from 1 to 17 years, with a median age of 5 years. Environmental sampling was applied to investigate the outbreak source. Bacterial identification and antimicrobial susceptibility tests of the isolated bacteria were performed using the disk diffusion method and Vitek®2 automated system. S. paucimobilis was detected in 181 blood culture samples from 51 patients over 2 years and was isolated from hot tap water. Acute lymphoblastic leukemia (ALL) was diagnosed for 66% of patients, and two of our patients with ALL died due to S. paucimobilis sepsis. S. paucimobilis isolates are susceptible to carbapenems and quinolones. Surveillance and epidemic control should be performed for hospital-acquired infectious agents such as S. paucimobilis. In additon, water distribution systems should be checked for colonizing agents at regular intervals.

In vitro activity of hyperthermia on swarming motility and antimicrobial susceptibility profiles of Proteus mirabilis isolates.

AIM: Swarming motility is a virulence factor for Proteus mirabilis and is a coordinated multicellular movement of bacteria. In this study, we investigated the inhibitory effect of hyperthermia on bacterial swarming motility and antimicrobial resistance. METHODS: Thirty-one P. mirabilis isolates were included in the study. Seven inoculated agar plates were incubated inside incubators with increasing temperature levels: at 36 °C (control) and 40-45 °C. On the next day, inhibition of swarming was evaluated and minimum paralyzing temperature (MPT) values were determined. An antimicrobial susceptibility test (antibiogram) is performed by exposing bacteria to increasing concentrations of antibiotics, in vitro. Thus, we used the Kirby-Bauer disk diffusion test as a screening method to analyze the antibiogram profiles of the isolates at 36 °C and 42 °C. Finally, a time-kill assay was performed to analyze the killing effect of hyperthermia (42 °C) on planktonic bacteria, in combination with the antibiotic meropenem at the first and third hours. A Wilcoxon signed-rank test was used to compare the killing effects of meropenem, hyperthermia and their combinations. RESULTS: The median MPT value was determined as 44 °C. In the disk diffusion assay, susceptibility development was observed in 94% of isolates for at least one antibiotic. In the time-kill assay, we observed a significant killing effect of hyperthermia in combination with meropenem. Under the microscope, we observed the formation of spherical cells by the effect of heat. CONCLUSION: We conclude that these findings might be useful when employing the hyperthermia method to treat infectious diseases caused by P. mirabilis in the future.

Investigation of an outbreak of Elizabethkingia meningoseptica on a pediatric intensive care unit.

Objective: This paper reports an Elizabethkingia meningoseptica outbreak on a pediatric intensive care unit with emphasis on investigation of outbreak source, infection control interventions, patient characteristics and comparative antimicrobial susceptibility results. Methods: This was an ambidirectional cohort study conducted in a university hospital 20-bed pediatric intensive care unit. Patient ages ranged from 4 to 11 months, with a median age of 9 months. 83% of the patients had severe underlying conditions. Samples from staff and environmental surfaces were obtained to identify a common source of infection. Antimicrobial susceptibility tests of isolated bacteria were done using the disk diffusion method and the Vitek®2 automated system. Results: Environmental surveillance revealed contamination of the water reservoirs of two different mechanical ventilators. In-vitro antimicrobial susceptibility testing results with two different methods (Vitek®2 and disk diffusion) were coherent for most of the investigated antibiotics, but without coherence for ciprofloxacin and levofloxacin. Resistance was found to the relatively new antibiotics ceftaroline and ceftazidime-avibactam. Conclusions: E. meningoseptica is a significant cause of nosocomial infections, with high mortality especially in children. Investigation of the outbreak source and continuation of intensive infection control precautions are vital to handle E. meningoseptica outbreaks in PICUs. Using quinolones according to testing results of automated AST systems may lead to inadequate treatment and foster the selection of resistant strains.

Investigation of heteroresistant vancomycin intermediate Staphylococcus aureus among MRSA isolates.

INTRODUCTION: Heteroresistant vancomycin intermediate Staphylococcus aureus (hVISA) testing is recommended when therapeutic failure is suspected in the clinics. In our research, we aimed to investigate the prevalence of hVISA among methicilline-resistant S. aureus (MRSA) isolates in our university hospital and compared three methods for detection of hVISA. METHODOLOGY: One hundred MRSA clinical isolates were collected in our medical microbiology laboratory between 01.04.2018 and 01.10.2019. For screening of hVISA, we used two screening agar plates and used one commercial medium; brain heart infusion agar (BHI) plates containing 4 µg/mL vancomycin and 16 g/Lt casein (BHIA-VC; Satola's test), BHI agar plates containing 4 µg/mLvancomycin (BHIAV), and commercially obtained vancomycin resistant Enterococci (VRE) agar for detetection of hVISA. Colonies which could grow on plates were counted manually at 24th and 48th hours. RESULTS: Among 100 MRSA isolates, 43 (43%) were found as hVISA using Satola's test. BHIAV and VRE agar screening test results were found 70% and 4%, respectively. Finally, at the step, MIC values of 20 (47%) hVISA isolates reduced to 2 µg/mL after sub culturing for the gradient test. CONCLUSIONS: We found higher rates of hVISA comparing other studies in Turkey. Both VRE agar and BHIAV screening test failed to detect hVISA properly. Meropenem in combination with vancomycin inhibited the growth of 90% hVISA isolates in our study.