Inhaler Steroid Use Changes Oral and Airway Bacterial and Fungal Microbiome Profile in Asthma Patients.

INTRODUCTION: The full spectrum of bacterial and fungal species in adult asthma and the effect of inhaled corticosteroid use is not well described. The aim was to collect mouthwash and induced sputum samples from newly diagnosed asthma patients in the pretreatment period and in chronic asthma patients while undergoing regular maintenance inhaled corticosteroid therapy, in order to demonstrate the bacterial and fungal microbiome profile. METHODS: The study included 28 asthmatic patients on inhaler steroid therapy, 25 steroid-naive asthmatics, and 24 healthy controls. Genomic DNA was isolated from induced sputum and mouthwash samples. Analyses were performed using bacterial primers selected from the 16S rRNA region for the bacterial genome and "panfungal" primers selected from the 5.8S rRNA region for the fungal genome. RESULTS: Dominant genera in mouthwash samples of steroid-naive asthmatics were Neisseria, Haemophilus, and Rothia. The oral microbiota of asthmatic patients on inhaler steroid treatment included Neisseria, Rothia, and Veillonella species. Abundant genera in induced sputum samples of steroid-naive asthma patients were Actinomyces, Granulicatella, Fusobacterium, Peptostreptococcus, and Atopobium. Sputum microbiota of asthma patients taking inhaler steroids were dominated by Prevotella and Porphyromonas. Mucor plumbeus and Malassezia restricta species were abundant in the airways of steroid-naive asthma patients. Choanephora infundibulifera and Malassezia restricta became dominant in asthma patients taking inhaled steroids. CONCLUSION: The oral and airway microbiota consist of different bacterial and fungal communities in healthy and asthmatic patients. Inhaler steroid use may influence the composition of the oral and airway microbiota.

[Comparison of the Results of Galactomannan and Beta Glucan Antigen Tests in the Suspicion of Invasive Pulmonary Aspergillosis]. / Invaziv Pulmoner Aspergilloz Süpheli Hastalarda Galaktomannan ve Beta Glukan Antijen Testlerinin Karsilastirilmasi.

Opportunistic fungal infections are an important cause of morbidity and mortality in immunocompromised patients. Invasive aspergillosis (IA) has an important place among these infections with ~ 250.000 cases annually. Reducing the mortality rate due to invasive aspergillosis is possible with early diagnosis and treatment of the disease. Because of the low sensitivity in microscopic examination, the time consuming of culture growth, and the difficulties in distinguishing colonization/infection, serological methods are frequently used in the diagnosis of invasive aspergillosis. The aim of this study was to determine the diagnostic performance of galactomannan and beta glucan tests for the diagnosis of invasive pulmonary aspergillosis (IPA). Sixty patients, followed up with the suspicion of invasive pulmonary aspergillosis in Gazi University Hospital were included in the study. The clinical classification of the patients was made according to the revised European Organization for Research and Treatment of Cancer and the Mycoses Study Group (EORTC/MSG) criteria. A total of 10 patients were classified as probable invasive aspergillosis and 20 patients were classified as possible invasive fungal disease. Demographic data of the patients and various risk factors were recorded. One hundred and thirty serum and nine bronchoalveolar lavage (BAL) fluid samples were studied with Plateliaᵀᴹ Aspergillus Ag (Bio-Rad, France), Dynamiker Aspergillus Galactomannan and Dynamiker Fungus (1-3)-beta-D-Glucan (Dynamiker, China) kits. Sensitivity and specificity values were calculated according to U.S. Food and Drug Administration (FDA) approved Plateliaᵀᴹ Aspergillus Ag test. According to this study, the most important risk factors in the development of IPA were the use of steroids and immunomodulatory drugs. The sensitivity of the galactomannan test in the probable group was 77.8%, the specificity was 96.7%, the sensitivity of the beta glucan test was 61.1%, and the specificity was 92.6%. When these two tests were evaluated together, it was observed that the sensitivity in the probable group increased to 83.3% and the specificity decreased to 89.3%. The combined use of galactomannan and beta glucan tests increases the diagnostic sensitivity. Although the presence of prolonged neutropenia is an important risk factor for IA, the use of steroids and immunomodulatory drugs should be kept in mind in non-neutropenic patients.

Frequency of azole resistance in clinical and environmental strains of Aspergillus fumigatus in Turkey: a multicentre study.

OBJECTIVES: Aspergillus fumigatus causes several diseases in humans and azole resistance in A. fumigatus strains is an important issue. The aim of this multicentre epidemiological study was to investigate the prevalence of azole resistance in clinical and environmental A. fumigatus isolates in Turkey. METHODS: Twenty-one centres participated in this study from 1 May 2018 to 1 October 2019. One participant from each centre was asked to collect environmental and clinical A. fumigatus isolates. Azole resistance was screened for using EUCAST agar screening methodology (EUCAST E.DEF 10.1) and was confirmed by the EUCAST E.DEF 9.3 reference microdilution method. Isolates with a phenotypic resistance pattern were sequenced for the cyp51A gene and microsatellite genotyping was used to determine the genetic relationships between the resistant strains. RESULTS: In total, resistance was found in 1.3% of the strains that were isolated from environmental samples and 3.3% of the strains that were isolated from clinical samples. Mutations in the cyp51A gene were detected in 9 (47.4%) of the 19 azole-resistant isolates, all of which were found to be TR34/L98H mutations. Microsatellite genotyping clearly differentiated the strains with the TR34/L98H mutation in the cyp51A gene from the strains with no mutation in this gene. CONCLUSIONS: The rate of observed azole resistance of A. fumigatus isolates was low in this study, but the fact that more than half of the examined strains had the wild-type cyp51A gene supports the idea that other mechanisms of resistance are gradually increasing.

Epidemiology and Risk Factors of Candidemia Among Hospitalized Patients in a Turkish Tertiary Care Hospital.

BACKGROUND: Candidemia, which constitutes 50 - 70% of invasive Candida infections, is an important clinical condition with high mortality and difficulty in diagnosis and treatment. Our objective was to determine the epidemiology, risk factors of candidemia, the distribution, and antifungal susceptibilities of Candida spp. responsible for candidemia among hospitalized patients in Gazi University Medical Faculty Hospital. METHODS: This was a laboratory-based, prospective observational study conducted between 2009 and 2010. The definition of nosocomial candidemia was based on CDC criteria. All relevant demographic and clinical data were collected from patient files. Candida spp. were identified by API ID32C system. Antifungal susceptibility testing was performed using broth microdilution method according to CLSI. RESULTS: Seventy-one candidemia episodes were identified with the incidence of 0.94 cases/1,000 hospital admissions. C. albicans was isolated in 47.9% of the admissions and in 52.1% of non-albicans Candida admissions. Among the latter, C. parapsilosis and C. tropicalis were the most frequent species. The most common risk factors were use of antibiotics (94.4%), hospitalization in the last 1 month (93%), history of hospitalization in ICU (74.6%), and CVC use (70.4%). Abdominal surgery, urethral catheter insertion, and use of piperacillin/tazobactam was found to increase the risk of C. albicans. A history of hospitalization within the last 3 months increased the risk of developing candidemia with non-albicans Candida spp. In total, fluconazole resistance was 20% (24.2% for C. albicans and 16.2% for non-albicans Candida strains) and voriconazole resistance was 5.7% (12.1% for C. albicans and 0% for non- albicans Candida). CONCLUSIONS: This study provided a relevant source of information for the prediction of high-risk patients and the implementation of prevention strategies for nosocomial candidemia.

[A case of Peritonitis Caused by Wickerhamomyces anomalus (Candida pelliculosa) Related to Peritoneal Dialysis]. / Wickerhamomyces anomalus (Candida pelliculosa)'un Etken Oldugu Periton Diyalizi Iliskili Bir Peritonit Olgusu.

Fungal peritonitis is less commonly seen than bacterial peritonitis in patients undergoing peritoneal dialysis (PD), but it is a serious complication with high morbidity and mortality. It often results in catheter loss and modifying therapy from PD to hemodialysis. The causative organisms are often Candida species. In this report, a PD-associated peritonitis caused by Wickerhamomyces anomalus (Candida pelliculosa), a rare fungal infection agent with increasing clinical importance by causing different clinical pictures was presented. An outpatient peritoneal fluid culture was sent from a 48-yearold male patient, who had been undergoing continuous peritoneal dialysis (CAPD) for 9 years, due to abdominal pain and blur in peritoneal fluid during dialysis. The patient admitted to the emergency department four days later due to the persistence of his complaints. A sample of peritoneal fluid was taken in the emergency department and sent to the laboratory for microbiological analysis. In the direct microscopical examination of the peritoneal fluid; cell number was determined as 210/mm3, and no microorganisms were seen in the Gram and methylene blue staining. The patient was admitted to the nephrology service with a pre-diagnosis of PD-associated peritonitis. Enterobacter aerogenes was grown in the peritoneal fluid culture which was sent from the dialysis outpatient clinic four days ago. The peritoneal fluid sample sent from the emergency department was inoculated on 5% sheep blood , EMB and chocolate agars and no growth was detected. As the patient's complaints and peritoneal fluid leukocyte count continued to increase, peritoneal fluid cultures were repeated and recurrent growth of yeast was detected in cultures. The yeast was identified as Candida pelliculosa by matrix assisted laser desorption ionization time-of-flight mass spectrofotometry (MALDI-TOF) VITEK®MS (bioMerieux, France). The species identification was confirmed by sequencing the target ITS gene regions on the rRNA and the isolate was identified as 100% Wickerhamomyces anomalus (sexual reproduction form of Candida pelliculosa, teleomorph). The reference microdilution method was performed according to the recommendations of the Clinical and Laboratory Standards Institute (CLSI) in order to test the antifungal susceptibility. After 24 hour incubation, the minimal inhibitory concentrations (MIC) were determined as 0.03 µg/ml for amphotericin B, 0.125 µg/ml for caspofungin 0.125 µg/ml for voriconazole, 0.03 µg/ ml for itraconazole and 4 µg/ml for fluconazole. Fluconazole and anidulafungin were started for the treatment of fungal peritonitis. The patient's peritoneal dialysis catheter was removed and hemodialysis was applied to the patient. Clinical and laboratory symptoms regressed with antifungal therapy and the patient's anidulafungin treatment was discontinued for 14 days after the catheter removal. In conclusion, in patients undergoing CAPD, as in our case, fungal pathogens should also be considered although it is rare, when there is no laboratory and clinical improvement, and the response to treatment is not complete in PD-associated peritonitis to prevent delays in diagnosis and treatment.

Retrospective analysis of the association of the expression and single nucleotide polymorphisms (SNPs) of the TLR4, PTX3 and Dectin-1 (CLEC/A) genes with development of invasive aspergillosis among haematopoietic stem cell transplant recipients with oncohaematological disorders.

OBJECTIVES: Several studies described single nucleotide polymorphisms (SNPs) on pattern recognition receptor (PRR) such as toll-like receptors (TLRs), dendritic cell-associated C-type lectin-1 (Dectin-1/CLEC7A) genes of patients with invasive fungal infections (IFIs) caused by Candida and Aspergillus. We screened TLR4, Dectin-1 and PTX3 polymorphisms in a Turkish population with invasive aspergillosis (IA) underlying haematological malignancies. METHODS: In this case-control study, a cohort of 59 patients with haematological malignancies were included. There were 26 IA patients assigned by the EORTC-MSG criteria and 33 patients with no evidence of fungal disease. DNA and RNA were isolated from frozen bone marrow and serum samples. RNA levels and polymorphisms of TLR4 (rs4986790, rs4986791), Dectin-1 (rs16910526, rs7309123) and PTX3 (rs2305619, rs3816527) were determined. The odds ratios (ORs) and corresponding 95% confidence intervals (CIs) were calculated by unconditional logistic regression analysis. RESULTS AND CONCLUSIONS: TLR4, PTX3 and Dectin-1 genes were downregulated in aspergillosis cohort under similar haematological conditions. TLR4 expression was 0.0626 ± 0.032 in controls when compared to IA patients as 0.0077 ± 0.014, and the difference was significant (P = .026). There was a difference in also the PTX3 gene among IA (0.0043 ± 0.004) and control (0.5265 ± 0.0043) groups (P = .035). The Dectin-1 (CLEC/A) expression was downregulated in IA group (0.1887 ± 0.072 & 0.0655 ± 0.010) but not statistically significant (P > .05). Conditional logistic regression analyses indicated that the GT genotype of rs16910526 polymorphism in Dectin-1 gene was associated with lower risk of IA (odds ratio = 3.635, 95% confidence interval = 0.690-3.138, P = .04).

[Candida hellenica Candidemia Occurred After Esophagus Surgery]. / Özofagus Cerrahisi Sonrasi Gelisen Candida hellenica Kandidemisi.

In this study, a case of candidemia caused by Candida hellenica as the first report in our country was presented. Fluconazole and liposomal amphotericin B treatment was initiated in a 20-year-old male patient in October 2018 due to the diagnosis of candidemia following esophageal surgery. The patient had a history of multiple esophageal operations. The patient was discharged during the last 24 hours due to the lack of fever, improvement in general condition and lack of growth in blood cultures. Germination tube test of the Candida isolate grown in blood culture was negative and the colony morphology in corn meal tween 80 agar was not defining. It was identified as C.hellenica according to the profile obtained from the ID32C® (bioMérieux, France) method based on carbohydrate assimilation. The target ITS regions of the rRNA genes were amplified by polymerase chain reaction and sequenced using suitable primers for the confirmation of the identification on species level. The DNA sequences obtained were searched by using the "National Center for Biotechnology Information (BLAST)" (http://www.ncbi.nlm.nih.gov/ BLAST/) database and the isolate was identified as C.hellenica with a 99% homology with GenBank sequences. MALDI-TOF (Vitek MS, bioMerieux) could not identify the yeast isolate. The reference microdilution method was performed according to the recommendations of the Clinical and Laboratory Standards Institute in order to test the antifungal susceptibility. The minimal inhibitory concentrations for the isolate, determined after 24-hour incubation were 0.25 µg/ml for amphotericin B, 8 µg/ml for fluconazole, 0.25 µg/ml for voriconazole, and 0.25 µg/ml for itraconazole. As our case had a previous history of gastrointestinal tract surgery it was thought that gastrointestinal tract was the endogenous source of candidemia by leading to mucosal disruption and this mucosal disruption might facilitate the translocation of Candida. The carbohydrate assimilation test ID32C®, was able identify the causative agent of candidaemia at the species level in this case. However, uncommon or previously unrecognized organisms may be misidentified by commercial systems. While the phenotypic definition is sufficient in routine laboratories, it is mandatory to confirm the microorganism species definition by DNA sequence analysis, as done in this case. We have presented a correctly identifed and successfully treated candidemia case. Although the candidemia was not mortal in our patient, the mortality rate of candidemia which is 50%, should be remembered. A total of two C.hellenica infections have been reported in the literature, including one candidaemia and one respiratory tract colonization. Our successfully treated case was presented to draw attention to this rare infectious agent.

[Characterization of Klebsiella pneumoniae strains exposed to subinhibitory concentrations of chlorhexidine]. / Düsük düzeyde Klorhekzidin ile karsilastirilmis Klebsiella pneumoniae izolatlarinin karakterizasyonu.

Chlorhexidine, a topical antiseptic, acts as a cationic biguanide altering the osmotic transport of the bacterial cell wall that has been used throughout the world to prevent healthcare-associated infections for decades. The routine application of chlorhexidine can result in decreased susceptibility of bacteria over time. The aim of this study was to develop Klebsiella pneumoniae strains after exposure to chlorhexidine and characterize these adapted strains in terms of their virulence ability both by in vivo and in vitro methods. Two clinical strains of K.pneumoniae were included in the study. One strain was completely susceptible and the other was resistant to certain antibiotics. Susceptible strain was subjected in the exposure assay as parent/wild strain. Exposure was performed by increasing chlorhexidine concentrations in agar plates. Chlorhexidine concentrations were gradually decreased reaching a final concentration of 0.12 mg/L after five weeks. Chlorhexidine-adapted viable colonies were selected and isolated. Minimal inhibitor concentrations of chlorhexidine, sodium hypochloride, benzalkonium chloride and triclosan for K.pneumoniae strains were determined using broth microdilution method. Reverse transcription-polymerase chain reaction analysis were performed for efflux pumps named cepA, kdeA and acrKp expressions. Fluorimetric efflux assay by using Rhodamine 6G was performed. Galleria mellonella killing assay and in vitro virulence determinants such as esculin hydrolysis, biofilm production, lecithinase, DNase activity, hemolytic activity, lipase production, mucoviscocity, casein hydrolysis and complement-mediated serum killing were evaluated. K.pneumoniae strains exposed to chlorhexidine did not show any antibiotic resistance. MICs for chlorhexidine, sodium hypochloride, and benzalkonium chloride were increased in the adapted strain. Efflux pumps of cepA and kdeA were over-expressed in the chlorhexidine adapted strain. Rhodamine 6G assay showed an increased efflux in the adapted strain. G.mellonella killing assay showed median virulence score. All strains, were esculin positive, while biofilm production, lecithinase, DNase, hemolytic activity, lipase production, mucoviscocity, casein hydrolysis were all negative. The susceptible parent/wild strain was susceptible to the complement-mediated serum killing, while the chlorhexidine adapted strain showed intermediate susceptibility. Chlorhexidine adapted strains of K.pneumoniae showed increased efflux pump expression, enhanced G.mellonella killing and raised resistance to serum killing. No difference was determined for other determinants. Minimal correlation was found between chlorhexidine resistance and virulence in K.pneumoniae.

Corneal Cross-Linking Has No Effect on Matrix Metalloproteinase 9 and 13 Levels During Fungal Keratitis on the Early Stage.

The aim of our study was to investigate matrix metalloproteinases, MMP-9 and MMP-13 levels, in the rabbit model of Fusarium and Candida keratitis treated by corneal cross-linking (PACK-CXL). Rabbit corneas were inoculated with fungal inoculum for keratitis. Each group divided into four subgroups, including un-treated group, PACK-CXL group, voriconazole group and PACK-CXL plus voriconazole group. PACK-CXL was applied with 0.25% riboflavin in accelerated Dresden protocol, and 0.1% voriconazole drops were administered. All corneal buttons excised at tenth day after ophthalmological examination. Fungal cell counts and Scheiber scores were determined in all groups. Corneal tissue MMP mRNA levels were evaluated quantitative reverse transcriptase PCR. The difference in MMP-9 and MMP-13 levels at all groups was not statistically significant (p > 0.05). PACK-CXL with 0.25% riboflavin either alone or combined with antifungal drops was unable to provide decline in inflammatory findings in both macroscopic and microscopic levels similar to medical antifungal treatment.

[Investigation of antifungal activities of some disinfectants on Candida albicans]. / Bazi dezenfektanlarin Candida albicans üzerine antifungal aktivitelerinin incelenmesi.

Disinfectants may have fungicidal or fungistatic effects against fungal cells. The mechanism of action of disinfectants on fungal cells believed to be similar to the antibacterial activity. The aim of this study was to demonstrate the efficacy of some disinfectants against Candida albicans and to investigate the relationship between virulence and disinfectant resistance. In this study, the susceptibility of 417 clinical C.albicans and reference isolates against disinfectants were determined. The minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) values of disinfectants were obtained by using broth microdilution (BMD) assay. Epidemiological cut-off values (ECVs) were determined by using the MIC and MFC values. Crystal violet assay was carried out to investigate membrane permeability in disinfectant resistant and susceptible isolates. Rhodamine 6G (R6G) flourescence stain was used to show the increase in the number of efflux pumps among selected isolates. The relationship between virulence and disinfectant resistance was determined by in vitro and in vivo investigations. Virulence factors secretory acid proteinase (SAP), phospholipase, esterase, hemolytic activity and slime factor production were examined in vitro. In vivo virulence assay was performed by infecting Galleria mellonella larvae. The relationship between virulence factors and disinfectant resistance was evaluated according to the mortality rates of G.mellonella larvae. The range of MIC values for benzalkonium chloride (BZC) and chlorhexidine digluconate (CHX), triclosan (TRC) and sodium hypochlorite (SHC) were 0.25-8 mg/L, 0.06-4 mg/L and 256-16.384 mg/L, respectively. ECV values for BZC, CHX, TRC and SHC were determined as 4, 2, 1 and 4096 mg/L, respectively. The rate of crystal violet uptake was found between 26.5-57.6% for disinfectant susceptible isolates, and between 33-79.2% for resistant isolates. It is concluded that the disinfectant resistance was related with efflux pumps. Due to the lack of number of isolates that were used in this assay, the relationship between disinfectant resistance and virulence factors could not be assessed. There was no difference in the mortality of larvae infections caused by disinfectant resistant and susceptible isolates. As a result, in this study, resistant isolates against BZC, CHX, SHC and TRC were found among 417 isolates. Input and output of disinfectants were found to be associated with the cell membrane efflux pumps of C.albicans.

[Detection of virulence factors and antifungal susceptibilities of Fusarium strains isolated from keratitis cases]. / Keratit olgularindan elde edilen Fusarium izolatlarinin virülans faktörlerinin ve antifungal duyarliliklarinin belirlenmesi.

Fusarium species have gained importance as a cause of keratitis. The pathogenicity and virulence factors of genus Fusarium remain largely unknown. Several putative virulence factors have been reported for fungal pathogens, including biofilm formation, production of proteinases and other hydrolytic enzymes. It has been emphasized that Fusarium species are generally resistant to antifungals but the resistance may vary depending on the species and even according to the isolate. For this reason, pathogenic features and antifungal susceptibility of the clinical isolates gained importance for the management of keratitis cases. The aim of this study was to identify clinical Fusarium isolates, to evaluate their virulence factors and to show antifungal susceptibility patterns. The identification of Fusarium was made on genus level isolated from 25 keratitis cases. Among them, 13 of the isolates were identified by ITS sequencing on species complex level. The production of hemolytic activity, caseinase, esterase, proteinase and phospholipase activity were investigated in 13 of the isolates. Biofilm production was searched among all 25 isolates. Galleria mellonella larvae was used as in vivo infection model. Antifungal susceptibility for amphotericin B, itraconazole, voriconazole and posaconazole was performed according to the Clinical and Laboratory Standards Institute (CLSI) M38-A2 microdilution assay guidelines. As the subcommittee on antifungal susceptibility tests did not determine the clinical resistance breakpoints (CBP) specific to Fusarium species complex, the epidemiological cut off values (ECV) were used for the interpretation of the minimum inhibitory concentration (MIC) values of the antifungal drugs. Isolates were identified as six F.oxysporum, six F.solani species complex and one F.brachygibbosum. One F.solani, one F.oxysporum were positive for hemolytic activity; all isolates were caseinase positive; three F.oxysporum and two F.solani isolate were esterase positive; one F.solani isolate was proteinase positive; five F.oxysporum and two F.solani isolates were phospholipase positive; biofilm activity was positive in 52% of the 25 isolates. The larvae survived for seven days after Fusarium inoculation in the G.mellonella larvae model. MIC range was 0.5-8 µg/ml for amphotericin B, 2-32 µg/ml for itraconazole, 0.5-8 µg/ml for voriconazole, 0.5-16 µg/ml for posaconazole and according to the ECV values F.solani and F.oxysporum isolates were determined as wild type for four antifungal agents. As a result, it was shown that Fusarium isolates have some virulence factors, there was a concordance between in vitro virulence properties and in vivo virulence characteristics and some of the isolates were classified as antifungal susceptible wild type isolates.

The in vitro effect of antimicrobial photodynamic therapy on Candida and Staphylococcus biofilms

Background/aim: This study was designed to evaluate the effect of antimicrobial photodynamic treatment (APDT) in a biofilm model using combinations of various dyes (rose bengal, riboflavin, and methylene blue) as photosensitizers and light sources (LED and UVA) against staphylococcal and candidal biofilms. Materials and methods: Sterile microtiter plates were used for the development and quantification of the biofilms. APDT was carried out using combinations of the light sources and dyes. The percentage of the growth inhibition was then calculated using a spectrophotometer. The broth media in the wells were aspirated, wells were stained with crystal violet, and optical density values were measured spectrophotometrically. SEM analysis of the impact of APDT on bacterial and fungal biofilms was also performed. Results: The experiments showed that the most efficacious combination was red LED + methylene blue against both staphylococcal and candidal biofilms. A marked inhibition (45.4%) was detected on both C. albicans and C. parapsilosis biofilms. Red LED + methylene blue was also effective on S. aureus and S. epidermidis biofilms. SEM images suggested that the number of adherent cells and biofilm mass were markedly reduced after APDT treatment. Conclusion: Although the results of this study indicated the in vitro efficacy of APDT, it might also be a promising technique for the control of biofilm growth within intravenous catheters.

[Emerging pathogen: Candida kefyr (Kluvyeromyces marxianus)]. / Önem kazanan patojen: Candida kefyr (Kluvyeromyces marxianus).

In the central microbiology laboratory of Gazi University Hospital Candida kefyr was isolated from different clinical samples as 5.3% in 2016 and in 2017 this rate increased to 9.3% which was nearly two-fold and this has drawn our attention. The aim of this study was to evaluate the special characteristics, antifungal susceptibility and virulence properties of C.keyfr species. Germ tube, corn meal-tween 80 agar morphology and carbohydrate assimilation profiles on ID32C yeast identification system were used for the diagnosis of Candida species. In this study, DNA sequencing was performed using ITS1 and ITS4 primers amplifying fungal gene between 5.8S and 18S regions of rRNA. Antifungal susceptibility was performed using M27A microdilution method recommended by Clinical and Laboratory Standards Institute (CLSI). Minimum inhibitory concentration (MIC) values for amphotericin B, fluconazole, voriconazole and itraconazole were determined. MIC distribution, MIC50 and MIC90 values and geometric mean (GM) were detected. The existence of virulence factors caseinase, secreted aspartyl proteinase, esterase and phospholipase were investigated in vitro. A total of 865 Candida species were isolated from different clinical samples in the central microbiology laboratory of Gazi University Hospital in 2016. Among them, 46 (5.3%) were C.kefyr. In the first four months of 2017, 30 (9.3%) C.kefyr were identified among 320 Candida isolates. Ten isolates which have shown atypical morphology on corn meal agar were selected. Among these 10 isolates, nine of them were identified as C.kefyr by using ID32C system and DNA sequencing method. Amphotericin B MIC value was 2 µg/ml for one isolate, and fluconazole MIC value was 8 µg/ml for another isolate among 46 isolates. Among the 30 isolates of the year 2017, one of them presented MIC value for fluconazole as 8 µg/ml. No marked antifungal resistance was detected in our isolate group. Caseinase was positive in one C.kefyr isolate, and phospholipase were positive in eight of nine isolates. As a result, the reason of increase in the incidence of this Candida species, which does not show significant resistance and presents mostly phospholipase activity as a virulence factor, should be investigated in more detail.

Circulating Th17 cytokine levels are altered in Hashimoto's thyroiditis.

The disrupted autoimmune response in Hashimoto's thyroiditis (HT) has long been considered to be dominantly T helper type 1 (Th1) mediated. Recent advances in the field of immunology have introduced a new class of effector T cells, named 'Th17', which plays important roles in autoimmune disorders once thought to be merely Th1 mediated. We aimed to examine the levels of major Th17 cytokines in patients with HT in this study. We studied serum interleukin 17 (IL-17) and interleukin 23 (IL-23) levels in 46 newly diagnosed, untreated patients with HT (40 women and 6 men, aged 40.0 ± 11.8 years) divided into euthyroid (n=22) and hypothyroid (n=24) groups and compared them with age and sex matched 26 healthy euthyroid controls without HT (21 women and 5 men; aged 36.0 ± 12.9 years). Serum IL-17 and IL-23 levels were significantly different among euthyroid and hypothyroid HT patients and controls, with highest levels obtained in the euthyroid HT group (p=0.041 for IL-17 and p<0.001 for IL-23). TSH was negatively and FT4 was positively correlated with IL-17 (p=0.016 for TSH and p=0.004 for FT4) and IL-23 (p<0.001 for TSH and p=0.003 for FT4) levels. There were no correlations between thyroid volumes calculated on thyroid ultrasonography and IL-17 (p=0.630) or IL-23 (p=0.321) levels. In conclusion, the levels of IL-17, one of the major effector cytokines of the Th17 system, and IL-23, which had been implicated in the generation, survival and expansion of Th17 cells, are altered in HT. How thyroid hormone status and the course of disease affect Th17 system in chronic autoimmune thyroiditis needs to be determined with further studies.

Microbiological and Histopathological Effects of Nasal Packing Containing Probiotics on Nasal Mucosa.

BACKGROUND: Nasal packing is frequently used after surgical interventions to prevent bleeding and synecchia formation and for the treatment of diseases such as epistaxis. One of the most morbid complications of nasal packing applications is the toxic shock syndrome (TSS). Owing to the microbiological structure of nasal mucosa, antibiotics are administered to all patients who are applied nasal packages for prevention of TSS. AIM: The aim of this study is the evaluation of microbiological and histopathological changes taking place in nasal mucosa with nasal packing containing probiotics. METHODS: Three groups were formed with 6 rats in each group. The nasal packings with the same characteristics were applied to nasal cavities of rats in all 3 groups. In group 1, only nasal packs were used. Probiotics or parenteral antibiotics were not used. In group 2, parenteral antibiotics were used along with nasal packs. In group 3, nasal packs with probiotics containing Lactobacillus strains were applied. No parenteral antibiotics were used. After 3 days packages were removed and nasal cavity was irrigated with saline. Both packages and irrigation materials were analyzed for microbiological content. After scarification, nasal and paranasal structures were examined for histopathological changes. RESULTS: In group 3 statistically the total bacteria load was significantly lower in comparison to the other groups. However, in the histopathological evaluation of the mucosa of rats in group 3, bleeding and inflammation findings were significantly higher statistically. CONCLUSIONS: It has been determined that the total microbiological load significantly decreases with the application of packing containing probiotics. So, the use of probiotics along with nasal packings is promising to prevent unnecessary use of medications.

[A rare and mortal infection agent in patients with hematological malignancies: Saprochaete capitata]. / Hematolojik maligniteli hastalarda nadir ve mortal bir enfeksiyon etkeni: Saprochaete capitata.

Saprochaete capitata (formerly known as Geotrichum capitatum and Blastoschizomyces capitatus) is a rare invasive fungal agent that may lead to mortal clinical course in patients with hematological malignancies. This agent can be colonized in skin, lungs and intestines, and it can cause major opportunistic infections. Invasive systemic infections due to S.capitata have been reported in immunosuppressed patients. In this report, two patients with invasive S.capitata infections detected during the course of persistent neutropenic fever in acute leukemia, were presented. In both cases empirical caspofungin was added to the treatment, as no response was obtained by board-spectrum antibacterial therapy in neutropenic fever. In the first patient, there were no significant findings except the chronic inflammation observed in the biopsies which was performed for the symptoms of lymphadenitis, myositis, and hepatosplenic candidiasis. While persistent fever was on going, S.capitata was isolated from the blood and catheter cultures. There was no response after catheter removing and the introduction of amphotericin B and voriconazole therapy, therefore allogeneic stem cell transplantation plan for the second time for bone marrow aplasia was taken an earlier time. However, the patient died due to progressive pericardial and pleural effusion and multiorgan failure, although an afebrile process after stem cell transplantation could be obtained. Similarly the second patient had persistent fever despite empirical caspofungin treatment. The additional symptoms of diarrhea, abdominal pain and subileus have indicated an intraabdominal infection. During the follow up, S.capitata was isolated from the blood and catheter cultures. Catheter was removed and amphotericin B was initiated. No response was obtained, and voriconazole was added to treatment. Despite of an afebrile and culture-negative period, the patient died as a result of Acinetobacter sepsis and multiorgan failure. Minimal inhibitory concentration values for both of the Saprochete strains were found as 0.25 µg/ml for amfoterisin B, 1 µg/ml for flukonazol, 0.125 µg/ml for vorikonazol and 0.25 µg/ml for itrakonazol. Virulence model was created by injecting the isolates to the Galleria mellonella larvae, and the life cycle of the larvae were determined. The observation revealed that the infected larvae began to die on the second day and there was no live larvae remained on the eleventh day. In conclusion, S.capitata should be considered as an infection agent with high mortality risk in the neutropenic patients with hematologic malignancies, especially in the presence of persistent fever during the use of caspofungin.

Successful treatment of Paecilomyces variotii peritonitis in a liver transplant patient.

Paecilomyces variotii has previously been reported as a causative pathogen for peritonitis in patients on continuous ambulatory peritoneal dialysis and shown to be usually sensitive to amphotericin B and resistant to voriconazole. We report the first case, to our knowledge, of P. variotii peritonitis in a liver transplant patient, which was unresponsive to initial liposomal amphotericin B (L-AmB) treatment and resolved dramatically after the addition of voriconazole. The present case provides evidence for the clinical and microbiological effectiveness of voriconazole combined with L-AmB in treating P. variotii peritonitis refractory to initial L-AmB treatment.

[Evaluation of risk factors for invasive pulmonary aspergillosis and detection of diagnostic values of galactomannan and PCR methods in bronchoalveolar lavage samples from non-neutropenic intensive care unit patients]. / Nötropenik olmayan yogun bakim hastalarinda invazif pulmoner aspergilloz için risk faktörlerinin degerlendirilmesi ve bronkoalveoler lavaj örneklerinde galaktomannan ve PCR testlerinin tanisal degerinin belirlenmesi.

Non-neutropenic intensive care unit (ICU) patients are at particular risk for invasive pulmonary aspergillosis. In these cases, radiological and microbiological methods (direct microscopy, culture), which can be used for diagnosis, have quite low sensitivity and specificity. The aims of this study were to evaluate the risk factors for invasive pulmonary aspergillosis (IPA) in non-neutropenic ICU patients and to determine the diagnostic values of galactomannan (GM) antigen and Aspergillus nucleic acid detection methods. A total of 44 patients (13 female, 31 male; age range: 36-96 years) who had been followed at pulmonary ICU with invasive mechanical ventilation and undergone bronchoscopy between January to December 2013, were included in the study. Consecutive bronchoalveolar lavage (BAL) and serum samples were obtained from all of the patients. BAL samples were tested for the presence of Aspergillus DNA by polymerase chain reaction (PCR) and both serum and BAL samples were tested for GM antigen by EIA method (Platelia Aspergillus, BioRad, France). EORTC/MSG criteria were used for the case definition of IPA. Patients were classified as high-probable IPA, possible IPA and non-IPA. ROC (receiver operating characteristics) analysis was used to determine the diagnostic values of BAL Aspergillus PCR and BAL GM in the diagnosis of IPA. Five patients were defined as high-probable IPA and six were defined as possible IPA; thus the incidence rate of IPA was estimated as 11.4% (5/44) among non-neutropenic intensive care unit patients. In high-probable IPA patients, BAL GM levels were significantly higher than non-IPA patients (p< 0.05). The prolonged duration in ICU, presence of septic shock and the use of high cumulative doses (> 460 mg) of steroid were found to be risk factors for IPA development. The cut-off value for GM in BAL samples was determined as 0.7, with a sensitivity rate of 100% (95% confidence interval: 47.9-100) and a specificity rate of 87.9% (95% confidence interval: 71.7-96.5), so optimal GM level in BAL was considered as ≥ 0.7 for the diagnosis of IPA. The specificity rates of serum GM and BAL Aspergillus PCR methods were high (97.1% and 93.9%, respectively), however their sensitivity rates were found quite low (33.3% and 40%, respectively), in the diagnosis of IPA. In conclusion, development of IPA should be assessed in non-neutropenic patients when the stay in ICU extends and high dose cumulative steroids are used. GM antigen detection in BAL can be used effectively for diagnosis of IPA in these patients compared to other diagnostic methods.