Burak Aksu

Effect of extremely low frequency electromagnetic fields on growth rate and morphology of bacteria

Purpose: To determine the effect of extremely low frequency (<300 Hz) electromagnetic fields (ELF-EMF) on the growth rate of Gram-positive and Gram-negative bacteria and to determine any morphological changes that might have been caused by ELF-EMF. Materials and methods: Six bacterial strains, three Gram-negative and three Gram-positive were subjected to 50 Hz, 0.5 mT ELF-EMF for 6 h. To determine growth rate after ELF-EMF application, bacteria exposed to ELF-EMF for 3 h were collected, transferred to fresh medium and cultured without field application for another 4 h. Growth-rate was determined by optical density (OD) measurements made every hour. Morphological changes were determined with Transmission electron microscopy (TEM) for two gram-negative and two gram-positive strains collected after 3 h of field application. Results: A decrease in growth rate with respect to control samples was observed for all strains during ELF-EMF application. The decrease in growth-rate continued when exposed bacteria were cultured without field application. Significant ultrastructural changes were observed in all bacterial strains, which were seen to resemble the alterations caused by cationic peptides. Conclusion: This study shows that ELF-EMF induces a decrease in growth rate and morphological changes for both Gram-negative and Gram-positive bacteria.

Extremely low-frequency electromagnetic fields affect the immune response of monocyte-derived macrophages to pathogens

This study aimed to determine the effect of extremely low-frequency electromagnetic fields (ELF-EMF) on the physiological response of phagocytes to an infectious agent. THP-1 cells (human monocytic leukemia cell line) were cultured and 50 Hz, 1 mT EMF was applied for 4-6 h to cells induced with Staphylococcus aureus or interferon gamma/lipopolysaccharide (IFγ/LPS). Alterations in nitric oxide (NO), inducible nitric oxide synthase (iNOS) levels, heat shock protein 70 levels (hsp70), cGMP levels, caspase-9 activation, and the growth rate of S. aureus were determined. The growth curve of exposed bacteria was lower than the control. Field application increased NO levels. The increase was more prominent for S. aureus-induced cells and appeared earlier than the increase in cells without field application. However, a slight decrease was observed in iNOS levels. Increased cGMP levels in response to field application were closely correlated with increased NO levels. ELF-EMF alone caused increased hsp70 levels in a time-dependent manner. When cells were induced with S. aureus or IFγ/LPS, field application produced higher levels of hsp70. ELF-EMF suppressed caspase-9 activation by a small extent. These data confirm that ELF-EMF affects bacterial growth and the response of the immune system to bacterial challenges, suggesting that ELF-EMF could be exploited for beneficial uses.

Catheter-related Saccharomyces cerevisiae Fungemia Following Saccharomyces boulardii Probiotic Treatment: In a child in intensive care unit and review of the literature

Although Saccharomyces boulardii is usually a non-pathogenic fungus, in rare occasions it can cause invasive infection in children. We present the case of an 8-year-old patient in pediatric surgical intensive care unit who developed S. cerevisiae fungemia following probiotic treatment containing S. boulardii. Caspofungin was not effective in this case and he was treated with amphotericin B. We want to emphasize that physicians should be careful about probiotic usage in critically ill patients.

Effect of extremely low frequency electromagnetic fields on bacterial membrane

Abstract Purpose: The effect of extremely low frequency electromagnetic fields (ELF-EMF) on bacteria has attracted attention due to its potential for beneficial uses. This research aimed to determine the effect of ELF-EMF on bacterial membrane namely the membrane potential, surface potential, hydrophobicity, respiratory activity and growth. Materials and methods: Gram-positive Staphylococcus aureus and Gram-negative Escherichia coli were subjected to ELF-EMF, 50 Hz, 1 mT for 2 h. Membrane potential was determined by fluorescence spectroscopy with or without EDTA (Ethylenediaminetetraacetic acid) with DisC3(5) (3,3-dipropylthiacarbocyanine iodide), zeta potential measurements were performed by electrophoretic mobility, hydrophobicity of the membrane was measured with MATH (Microbial Adhesion to Hydrocarbons) test, respiratory activity was determined with CTC (5-Cyano-2,3-ditolyl tetrazolium chloride), colony forming unit (CFU) and DAPI (4′,6-diamidino-2-phenylindole, dihydrochloride) was used for growth determinations. Results: ELF-EMF caused changes in physicochemical properties of both Gram-positive and Gram-negative bacteria. Hyperpolarization was seen in S. aureus and EDTA-treated E. coli. Surface potential showed a positive shift in S. aureus contrariwise to the negative shift seen in EDTA-untreated E. coli. Respiratory activity increased in both bacteria. A slight decrease in growth was observed. Conclusion: These results show that ELF-EMF affects the crucial physicochemical processes in both Gram-positive and Gram-negative bacteria which need further research.

Value of washed sputum samples in children with lower respiratory tract infections

Lower respiratory tract infections (LRTI) represent one of the serious infections in children with significant morbidity and mortality, especially in developing countries. Accurate identification of respiratory pathogens is the basis of management, and initial appropriate treatment is related to lower mortality rate. Microbiologic diagnosis of LRTI in children is difficult because samples of sputum are difficult to obtain and invasive collection techniques such as bronchoscopy or biopsy are rarely performed. Among respiratory tract samples sputum is a non-invasive clinical specimen, but normal resident bacteria of the oropharynx may contaminate samples and a large number of different species may overgrow, preventing determination of true epidemiologic agent. Previously some authors have suggested that in order to reduce bacterial contamination, sputum can be washed, and, to increase the yield of cultivation, a certain amount of the sample can be inoculated quantitatively. The aim of the present study was therefore to determine the incidence of bacterial pathogens in children with clinically suspected LRTI and to evaluate the value of direct examination and quantitative culture method of sputum samples. Antimicrobial therapy for LRTI is frequently empirical and selection of appropriate drug requires awareness of the patterns and clinical significance of antibiotic resistance. We analyzed the susceptibility patterns of the present bacterial isolates in order to generate antimicrobial susceptibility data to guide physicians in choice of treatment.

Expression of the adeB gene and responsiveness to 1-(1-naphthylmethyl)-piperazine and phenylalanyl-arginyl-β-naphthylamide in clinical isolates of Acinetobacter baumannii

Sir, AdeABC, a resistance–nodulation–cell division (RND)-type efflux pump, is an attractive research target as a cause of multiple antibiotic resistance in Acinetobacter baumannii. The overexpression of the adeB gene encoding the AdeB efflux protein has previously been associated with multidrug resistance in various A. baumannii strains. – 3 These reports led us to perform a study in a group of clinical isolates of A. baumannii, to assess the relative expression of the adeB gene and responsiveness to the potential efflux pump inhibitors (EPIs), 1-(1-naphthylmethyl)-piperazine (NMP) and phenylalanyl-arginyl-b-naphthylamide (PAbN). – 4 Fourteen multidrug-resistant A. baumannii isolates obtained from different patients in an intensive care unit and grouped previously into three clones were included in the study (Table 1). A. baumannii ATCC 19606, A. baumannii SBMox2, A. baumannii U10247 and Escherichia coli ATCC 25922 were used as reference strains. The MICs of ciprofloxacin, gentamicin, erythromycin, chloramphenicol, trimethoprim and tetracycline were determined by the broth microdilution test in the absence and presence of NMP and PAbN, at final concentrations of 100 mg/L. For the detection of adeB transcripts, RT–PCR was performed (RNeasy Kit and Omniscript RT Kit from Qiagen, Hilden, Germany; and FastStart DNA Master SYBR Green I Kit from Roche, Mannheim, Germany) using LightCycler System 2 (Roche) with the primers previously described by Higgins et al. Eight isolates (MU-2, -5, -7, -9, -10, -12, -13 and -14) showed reductions in various antibiotic MICs after the addition of EPIs (Table 1). This demonstrates the ability of NMP and PAbN to partially reverse drug resistance in a significant portion of the isolates. We can conclude that the phenotypic changes in the drug susceptibilities of our EPI-responsive isolates might be associated with the inhibition of functional RND-type drug efflux. – 4 In general, NMP was more active than PAbN. Such differences in the activities of the two compounds have been previously reported and associated with the different mechanisms of action of NMP and PAbN. Significant effects of NMP were observed on ciprofloxacin, erythromycin and trimethoprim MICs (Table 1). Moreover, NMP switched the resistance phenotype to susceptible in two isolates (MU-7 and A. baumannii SBMox2) for trimethoprim and/or tetracycline. This inhibitor may have affinity sites similar to those for these antibiotics inside the efflux pump. The adeB expression of EPI-responsive isolates was as high as that found in A. baumannii SBMox2, which has been previously shown to be an adeB overexpresser (Table 1). This supports our conclusion that the responsiveness of the isolates to EPIs might be due to the inhibition of functional RND-type drug efflux, particularly the AdeB pump. Earlier experiments have also highlighted the causal connection between adeB overexpression and responsiveness to EPIs in various A. baumannii strains. Despite the fact that the aminoglycosides have been shown to be good substrates of the AdeB pump, the gentamicin MICs for most adeB overexpressers did not markedly change in the presence of EPIs. – 3,7,9,10 This could be due to the presence of other resistance mechanisms, such as aminoglycoside modification enzymes and/or some recently recognized efflux pumps, which may have low affinity for the EPIs tested here. Alternatively, the affinity constant for the efflux pump sites could be stronger for gentamicin in comparison with EPIs. In terms of changes in quinolone susceptibility, despite the fact that NMP decreased the ciprofloxacin MICs from 16–32 mg/L to 2–8 mg/L, the adeB overexpressers were still resistant to this antibiotic (Table 1). The persistence of ciprofloxacin resistance in these isolates might be associated with mutations in the gyrA gene. Interestingly, the adeB expression of A. baumannii ATCC 19606 was high and similar to that of A. baumannii SBMox2. Although this strain was initially found to be susceptible to ciprofloxacin and tetracycline, the addition of NMP caused reductions in their MICs (Table 1). This supports earlier findings suggesting the efficiency of the AdeB pump alone was not sufficient to increase various antibiotic MICs to

Investigation Of Macrolide Resistance Mechanisms In Streptococcus Pneumoniae: Results Of Marmara University Hospital Between 2005-2008

ÖZET Amaç: Bu çalışmada enfeksiyon etkeni olarak izole edilen Streptococcus pneumoniae izolatlarında, makrolid direncinin fenotipik karakteristiklerinin ve genetik determinantlarının belirlenmesi amaçlanmıştır. Gereç ve Yöntemler: Eritromisin direnci gösteren 50 S. pneumoniae izolatının 14-, 15üyeli makrolid ve linkozamid duyarlılıkları disk difüzyon ve sıvı mikrodilüsyon yöntemleriyle saptanmıştır. Makrolid direnç fenotiplerini belirlemek üzere eritromisin-klindamisin çift disk testi kullanılmıştır. Makrolid direncinin genetik determinantları, mef(E)/(A), erm(B) ve erm(TR)’nin saptanmasında Polimeraz Zincir Reaksiyonu (PZR) yöntemi kullanılmıştır. Bulgular: İzolatlarımızın % 86’sı Makrolid-LinkozamidStreptogramin B (MLSB) (%76 cMLSB ve %10 iMLSB), % 14’ü M fenotipi göstermiştir. PZR sonuçlarına göre 18 izolatta (% 36) tek başına erm(B); 11 izolatta (% 22) tek başına mef(E)/(A) geni; geri kalan 21 (% 42) izolatta ise her iki gen birlikte saptanmıştır. erm(B) geni taşıyan tüm izolatlar (%78), yüksek düzey makrolid ve linkozamid direnci gösterirken, mef(E)/(A) genini tek başına taşıyan 11 izolatın 7’si M, 4’ü MLSB fenotipi göstermiştir. M fenotipindeki izolatların makrolid MİK’leri ise düşük düzeyde bulunmuştur. Sonuç: Çalışmamızda mef(A)/(E) geninin tek başına ya da erm(B) ile birlikte izolatların %64’ü tarafından taşındığının gösterilmesi ayrıca M fenotipine sahip izolatların oranının da oldukça yüksek (%14) bulunması, aktif makrolid pompasının, ribozomal hedef mutasyonuyla birlikte hastanemiz pnömokok izolatlarının makrolid direncinde çok önemli rolü olduğunu ortaya koymaktadır. Anahtar Kelimeler: Streptoccocus pneumoniae, makrolid direnci, mef(A)/(E), erm(B) ABSTRACT Objective: In this study, we aimed to determine the phenotypic characteristics and genetic determinants of macrolide resistance in the clinical isolates of Streptococcus pneumoniae. Materials and Methods: In 50 erythromycin resistant S. pneumoniae isolates, 14-, 15membered macrolides and lincosamide susceptibilities were determined by both disk diffusion and broth microdilution methods. The erythromycin-clindamycin double disk method was applied for the detection of macrolide resistance phenotypes. Genetic determinants of macrolide resistance, erm(B), erm(TR) and mef(A)/(E) were investigated by Polymerase Chain Reaction (PCR). Results: The percentages of the isolates presenting Macrolide-Lincosamide-Streptogramin B (MLSB) and M phenotypes were 86% (cMLSB phenotype: 76%; iMLSB phenotype: 10%) and 14%, respectively. According to the PCR results, 18 isolates (36%) had erm(B) gene, 11 isolates (22%) had mef(E)/(A) gene and the remaining 21 (%42) isolates had both erm(B) and mef(A)/(E) genes. All erm(B) positive isolates (78%) presented high level macrolide and lincosamide resistance. Seven out of 11 isolates carrying the mef(A)/(E) gene alone presented M phenotype with low level macrolide MICs. Conclusion: The demonstration of the mef(A)/(E) gene either alone or together with erm(B) in a high proportion of the isolates (64%) in addition to the high rate of M phenotype (14%) signifies that active macrolide efflux together with the ribosomal target mutation has a significant role in the macrolide resistance of our pneumococcal isolates.

Group B Streptococci Induce Interleukin 8 Production in Human Cervical Epithelial Cell Cultures: The Role of Capsule Polysaccharide

Objective: Group B streptococci (GBS) are the major cause of pneumonia, sepsis, and meningitis in neonates and adults. Epithelial invasion and early cytokine response of female genital tract considered to be important in the pathogenesis of GBS infection. In this study, we studied the IL-8 induction in cervical epithelial cells in response to stimulus with encapsulated (COH1) and unencapsulated (COH1-13) strains of group B streptococci. Methods: Human cervical epithelial cancer cell (HeLa) cultures were stimulated with different concentrations (106 CFU/ml and 108 CFU/ml) of two GBS strains. E.coli LPS was used as positive control and at specified time points (4, 8 and 24 hour) cell culture supernatant samples were collected. IL-8 level in samples was quantified by using ELISA assay. Results: Both GBS strains caused an equal IL-8 response in HeLa cells in a time-dependent manner. In addition, cytokine levels triggered by different bacterial concentrations were similar and comparable with LPS. Conclusion: Our study showed that GBS induce proinflammatory IL-8 levels in cervix epithelial cells. This induction seems to be independent from capsule polysaccharides and suggesting that other bacterial components are involved in IL-8 stimulation.

Performance of “RESIST-3 O.K.N. K-SeT” immunochromatographic assay for the detection of OXA-48 like, KPC, and NDM carbapenemases in Klebsiella pneumoniae in Turkey

In this study, the performance of the “RESIST-3 O.K.N. K-SeT” (Coris BioConcept, Gembloux, Belgium) immunochromatographic assay was evaluated in 132 Klebsiella pneumoniae comprising 102 carbapenem resistant and 30 carbapenem susceptible isolates. Genotypically known isolates of Gram negative bacteria (n = 22) including various species were also tested by the assay as controls. The isolates tested by the immunochromatographic assay and also were run PCR for blaKPC, blaIMP, blaVIM, blaNDM, and blaOXA-48. The rates of blaNDM, blaOXA-48, and blaKPC in carbapenem resistant isolates were found at 52.9%, 39.2%, and 2.0%, respectively. Both blaNDM and blaOXA-48 were found in six (5.9%) isolates. The results of the assay showed 100% concordance with those obtained by PCR in 132 K. pneumoniae. The agreement between the two methods was found to be identical at the isolate level. The assay also correctly detected all genotypically known isolates of Escherichia coli, Serratia marcescens, Citrobacter freundii, Enterobacter cloacae, K. pneumoniae carrying blaKPC, blaNDM, and/or blaOXA-48. On the other hand, the assay did not exhibit any cross-reaction in control isolates harboring blaIMP and blaVIM. We conclude that the RESIST-3 O.K.N. K-SeT is a reliable, rapid, and user friendly test and we recommend it for routine diagnostic laboratories.

Helicobacter pylori prevalence and relation with obesity

I n d I a n J o u r n a l o f P a t h o l o g y a n d M I c r o b I o l o g y ¦ V o l u M e 6 0 ¦ I s s u e 3 ¦ J u l y s e P t e M b e r 2 0 1 7 451 2012;10:1029‐36. 13. Terwijn M, Kelder A, Snel AN, Rutten AP, Scholten WJ, Oussoren YJ, et al. Minimal residual disease detection defined as the malignant fraction of the total primitive stem cell compartment offers additional prognostic information in acute myeloid leukaemia. Int J Lab Hematol 2012;34:432‐41.