The Microbiological Profile of Bicanalicular Silicone Tubes Placed During External Dacryocystorhinostomy

Objectives: To identify microbiological growth on bicanalicular silicone tubes (BST) placed during dacryocystorhinostomy (DCR) surgery and to analyze the association between culture results and surgical outcomes and BST removal time. Materials and Methods: A total of 80 lacrimal drainage systems of 68 patients who had external DCR with bicanalicular silicone intubation were included the study. Twenty-five tubes (31.3%) were removed up to 8 weeks, 28 tubes (35.0%) were removed between 9 and 11 weeks, and the remaining 27 tubes (33.7%) were removed 12 weeks or more after surgery. The tubes were transferred to Stuart medium and sent for microbiologic examination. The disc diffusion method was used to determine antibiotic resistance. Results: Culture positivity was observed for 96.2% of the tubes. Among a total of 109 isolates, 63 were gram-positive bacteria (57.8%), 37 were gram-negative bacteria (34%), and 9 were fungi (8.2%). The most commonly isolated gram-positive and gram-negative bacteria were Staphylococcus aureus (66.6%) and Enterobacter spp. (29.7%), respectively. Penicillin, clindamycin, erythromycin, and tetracycline resistances were higher among gram-positive pathogens. Cephalothin, amoxicillin-clavulanic acid, and ampicillin resistances were higher among gram-negative pathogens. There was no significant difference in terms of the microbiological profile between the three groups of removed tubes. Haemophilus influenzae was isolated at a significantly higher rate in patients with surgical failure (p=0.04). Conclusion: Although a variety of agents were isolated from removed BST, gram-positive organisms were more frequent than gram-negatives and fungi. S. aureus and Enterobacter were the most common gram-positive and gram-negative isolates. Later BST removal was associated with the isolation of significantly more bacterial strains per tube. There was no correlation between multiple infections and surgical failure. H. influenzae was more common in failed DCR cases.

Molecular Identification and Antimicrobial Activity of Vaginal Lactobacillus sp.

BACKGROUND: The genus Lactobacillus has recently been the focal point of researchers due to their ability to produce secondary metabolites with antimicrobial effects. OBJECTIVE: The aim of this study was to identify vaginal Lactobacillus sp. by analyzing 16S rRNA gene sequence, to investigate into antimicrobial activities of secondary metabolites produced by these isolates and to determine the quantities of lactic acid, acetic acid and hydrogen peroxide in secondary metabolites using High-Performance Liquid Chromatography. METHODS: Antimicrobial activities of secondary metabolites were investigated against test microorganisms using Agar Well Diffusion and Agar Spot Methods. RESULTS: According to the results, 18 L. crispatus, 17 L. gasseri, 5 L. jensenii, 4 L. vaginalis, 3 L. fermentum, 2 L. coleohominis, 1 L. saerimneri, 1 L. reuteri, 1 L. johnsonii and 1 L. helveticus were identified. Isolates were frequently found to be effective against Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853, Klebsiella pneumoniae RSKK 578 and Bacillus subtilis ATCC 6633. None of the isolates showed activity against Staphylococcus aureus ATCC 43300, S. epidermidis ATCC 12228, S. epidermidis ATCC 35984 and Candida albicans ATCC 10231. Secondary metabolites of all tested isolates contain hydrogen peroxide between 7.306 and 0.33 µg/µL range. CONCLUSION: It was found that the secondary metabolites of some isolates contained both acetic and lactic acid, while some of them contained either acetic or lactic acid.

Determining the persistence of maternally acquired antibodies to hepatitis A and varicella zoster during the first 2 years of life in Turkey.

UNLABELLED: Vaccination should be timed to take into account the potential interference of maternal antibodies. The purpose of this study was to determine the persistence of maternally acquired antibodies to hepatitis A and varicella zoster in a group of healthy infants between 6 and 24 months of age. These infants were divided into four groups according to the age at the time of follow-up visits. The study group consisted of infants who were brought to the 6-month follow-up visit (group 1, n=100), 12-month follow-up visit (group 2, n=99), 18-month follow-up visit (group 3, n=59), and 24-month follow-up visit (group 4, n=59). Hepatitis A, varicella IgG, and IgM antibodies were analyzed qualitatively. Hepatitis A IgG seropositivity was determined as 71 % in group 1, 41.4 % in group 2, 0 % in group 3, and 8.5 % in group 4 (p<0.001). Varicella IgG seropositivity was found to be 5 % in group 1, 4 % in group 2, 4 % in group 3, and 1 % in group 4 (p>0.05). CONCLUSION: We found that maternal hepatitis A antibodies in children disappear between 12 and 18 months, whereas maternal varicella antibodies substantially diminish following the sixth month. Therefore, the vaccination timing should be based on factors such as the interference of maternal antibodies, disease susceptibility period, and immune maturation.

[Investigation of seven different RNA viruses associated with gastroenteritis in children under five years old]. / Bes yas alti çocuklarda gastroenterite neden olan yedi farkli RNA virusunun arastirilmasi

Viruses are the most frequently detected etiologic agents of gastroenteritis seen in small children. In addition to classical gastroenteritis viruses namely rotavirus, norovirus, adenovirus type 40/41, astrovirus and sapovirus, some novel picornaviruses (Aichi virus, parechovirus, enterovirus) that have been identified in parallel to the developments in molecular diagnostic methods, thought to be associated with diarrhea in humans. However, the data are not enough to prove their actual roles in the pathogenesis of gastroenteritis. The aim of this study was to investigate the presence of rotavirus, norovirus, sapovirus, astrovirus, Aichi virus, parechovirus and enterovirus in the stool samples of children with diarrhoea by reverse-transcriptase polymerase chain reaction (RT-PCR). A total of 50 samples from children admitted to our hospital with diarrhoea between June-December 2012 were included in the study. All the patients were under 5 years of age. Routine bacteriological and parasitological examinations of the patients' stool samples were negative. Total RNAs were extracted from each of the samples and cDNAs were obtained by reverse transcription. All cDNAs were investigated first with the internal control (IC) using PCR. Thirty-one of the 50 cDNAs (62%) were found IC positive. Those 31 samples were further investigated in terms of rotavirus group A and C, norovirus (NoV) genogroup GI and GII, sapovirus, astrovirus, Aichi virus, parechovirus and enterovirus by PCR using specific primer pairs. The predicted sized PCR products obtained were cloned into the pBSK cloning vector and were sequenced. Sequences obtained were subjected to a BLAST search with registered sequences in the GenBank database for the confirmation of the PCR product. Out of 31 RNA positive stool specimens, 12 (38.7%) were found positive for five types of the target viruses. NoV GII (6/31, 19.3%) were detected as the most prevalent virus, followed by NoV GI (2/31, 6.5%), rotavirus group A (2/31, 6.5%), astrovirus (1/31, 3.2%) and sapovirus (1/31, 3.2%). The results of this study revealed that the most frequently detected agents were noroviruses (8/50, 16%) followed by rotavirus (2/50, 4%), astrovirus (1/50, 2%) and sapovirus (1/50, 2%). It was concluded that RT-PCR performed with the primers used in this study could be applied effectively for the molecular epidemiological analysis of RNA viruses leading to gastroenteritis. Larger scale studies conducted in different areas for longer time periods and with a larger population size will provide data for the epidemiological properties of agents of viral gastroenteritis.

HHV-6 is ubiquitously found using Western blot in tonsils and adenoid tissues of healthy people.

Few studies have examined the prevalence and cellular proclivity of latent human herpesvirus 6 (HHV-6) in healthy populations. Difficulties in detection of HHV-6 genome in different tissues using polymerase chain reaction (PCR) and immunohistochemistry (IHC) techniques have been reported by various researchers. We examined tonsils and adenoid tissues of 54 patients who had undergone tonsillectomy or adenoidectomy without any evidence of acute infection for the presence of latent HHV-6 infection. While we were investigating the prevalence of HHV-6, we tested the efficiency of PCR, IHC and Western Blot (WB) for detection of HHV-6 in tonsil tissues. We found that 100% of tonsil tissues were positive for HHV-6 with WB, 40% of tonsils were positive with PCR and no tonsil was positive with IHC. This result correlates well with most studies claiming HHV-6 is a ubiquitous organism in various populations and tissues. Western blot may be a good choice for detecting HHV-6 in tissues. Expression of the HHV-6 gp60/110 envelope protein disclosed by WB may indicate that HHV-6 does not have true latency. To our knowledge, this is the first report to use WB to test for HHV-6 in tissues.

Expression analysis of efflux pump genes among drug-susceptible and multidrug-resistant Mycobacterium tuberculosis clinical isolates and reference strains.

Finding a gene or genes that are involved with multidrug resistance will be useful for finding a new target for the treatment of drug resistant tuberculosis. In this study, we aimed to compare the differences of the expression of 15 putative multidrug efflux pump genes in clinically isolated drug sensitive and multidrug resistant (MDR) Mycobacterium tuberculosis isolates, and reference strains. We found that these genes in the drug-sensitive and MDR M. tuberculosis isolates have similar rates of expressions. However, we found the expression levels of the all the genes are significantly higher in the clinical strains compared to the expression level of genes in the reference strains. In addition to this, it is found that standard strain has lower MIC value for the drugs including streptomycin and rifampin compared to the clinical isolate. We presume that the increase of the gene expression in the clinical strains is due to the exposure of antituberculosis drugs during treatment of patients, which cause constitutive expression of efflux systems, which might increase MIC levels of the major anti-tuberculosis drugs.

[Identification of a novel lytic bacteriophage obtained from clinical MRSA isolates and evaluation of its antibacterial activity]. / Klinik MRSA Izolatlarindan Elde Edilen Yeni Bir Litik Bakteriyofajin Tanimlanmasi ve Antibakteriyel Etkisinin Degerlendirilmesi.

Multidrug-resistant bacteria particularly MRSA is well known as a worldwide problem. Since the rate of development of novel antimicrobial agents has been slowed down during the last years, there have been a need for the exploration of alternative solutions for the treatment of resistant bacterial infections. Treatment of infections by bacteriophages (phages) that specifically kill the infecting pathogen, i.e. by the process known as phage therapy, is considered as a possible approach to treat multidrug resistant bacteria. Phage treatment has also been considered to treat Staphylococcus aureus infections. This study was aimed to evaluate the antibacterial and cytotoxic activities of a new lytic phage obtained from clinical MRSA strains. This lytic phage named as f LizAnk was obtained during the phage infectivity studies performed with 13 lysogenic phages against MRSA strains. The antibacterial activity of the f LizAnk phage was determined in vitro in BHI (Brain Heart Infusion) and LB (Leuria Bertani) broths and the in vivo antibacterial activity against MRSA strains and possible cytotoxic effect against mammalian cells were tested on fibroblastic cell cultures (3T3). This study was conducted using 20 MRSA strains isolated from hospitalized patients. Identification of the isolates was performed by conventional methods and methicillin resistance was detected with oxacillin disk diffusion test and mecA gene detection by PCR. The method described by Kaneko et al. [Biosci Biotechnol Biochem 1997; 61(11): 1960-2] was used with some modifications, for induction and isolation of the phages. In vitro studies indicated that this phage killed the six different MRSA strains (in 107 cfu/ml concentrations) in 8 hours, and this powerful lytic effect was similar in both of the liquid media. In vivo studies were performed by using cell cultures prepared in microplates, and the wells have been inoculated with only phage, phage + MRSA mixture, and only MRSA. The cells were then evaluated microscopically as well as by MTT assay which detected alive cells colorimetrically, at 2nd and 24th hours. In our study, the f LizAnk phage did not cause any toxic effect on fibroblast cell cultures, in addition it was observed that the antibacterial effect of the phage against MRSA has proceeded in the cell culture. In conclusion, since the fLizAnk phage described in this study exhibited strong antibacterial activity against MRSA strains and no cytotoxic effect was detected against mammalian cells, it might be safely used alone or in a phage cocktail to treat skin infection caused by MRSA.

[Investigation of Salmonella serotype Enteritidis isolates by plasmid profile analysis and pulsed field gel electrophoresis]. / Salmonella Serotip Enteritidis Izolatlarinin Plazmid Profil Analizi ve "Pulsed Field" Jel Elektroforezi ile Incelenmesi.

In this study a total of 122 Salmonella serotype Enteritidis stock strains selected from the culture collection of Enterobacteriaceae Laboratory of Ankara University Faculty of Medicine, Department of Medical Microbiology, were investigated by plasmid profile analysis with the method defined by Kado and Liu and pulsed field gel electrophoresis (PFGE) according to World Health Organization protocols using SpeI and XbaI macrorestriction enzymes, for better understanding of the molecular epidemiology of S. Enteritidis. The study strains were selected from a collection of previously isolated epidemic (n= 13) and sporadic (n= 109) strains (103 stool, 16 blood and one each bile, urine and cerebrospinal fluid) obtained from 10 different cities after the year 2000. PFGE patterns were analyzed with Gene Directory software (Syngene, UK) and a similarity index was determined by using Dice coefficient and the unweighted pair group method with mathematical averaging (UPGMA). Plasmid-carrying 110 (90%) strains that harbored 1-4 plasmids with sizes ranging from 2.0 to 100 kb were separated into patterns more than 14 (p1-p14). A total of 85 (69.7%) isolates harbored the 57 kb plasmid solely or in combination with other plasmids. By PFGE, 11 distinct patterns were shown with each enzyme SpeI and XbaI. S. Enteritidis strains after digestion with macrorestriction enzyme SpeI generated 11 different PFGE patterns (A to K), whereas XbaI generated also 11 different PFGE patterns (a to k). PFGE pattern A consisted of 93 strains (76.2%) after digestion with macrorestriction enzyme SpeI, while PFGE pattern a consisted 53 (43.4%) and PFGE pattern b 42 strains (34.4%) after digestion with macrorestriction enzyme XbaI. Using two macrorestriction enzymes two PFGE cluster profiles Aa (50 strains, 40.9%) and Ab (42 strains, 34.4%) were found to be predominating among 17 different PFGE clusters. Our results confirmed the clonal nature of S. Enteritidis strains in Turkey. The use of two enzymes in PFGE analysis appeared to increase the discriminatory power of PFGE, leading to greater diversity among strains. PFGE analysis performed by SpeI and XbaI enzymes combined with plasmid profiling could be established as a useful tool for detection of genetic relationship between isolates.

A new, simple, rapid test for detection of DNase activity of microorganisms: DNase Tube test.

The DNase test is a simple, economical method that has traditionally been used as a supplemental test to identify pathogenic Staphylococcus. This test also aids in the differentiation of closely-related genera within the Klebsiella-Enterobacter-Serratia division of Enterobacteriaceae and several other pathogens, including screening of C. diphtheriae. Currently DNase activity of microorganisms was tested using DNase agar plate methods. These tests have some drawbacks including the necessity of the extensive time to see the results of DNase activity of bacteria. In here, we developed a new method which is simple, rapid, inexpensive and applicable to examine DNase activity of any bacteria. In this method, simply, bacteria is added to the broth medium containing DNA and followed for the DNA degradation caused by the DNase of bacteria by running in agarose gel. This method we called DNase Tube test showed DNA degradation as fast as in half an hour depending on the DNase activity of the bacteria.

[A novel Salmonella Typhimurium plasmid, pAnkS: an example for plasmid evolution in antibiotic resistance]. / Yeni bir Salmonella Typhimurium plazmidi, pAnkS: antibiyotik direnç gelisiminde plazmid evriminin bir örnegi.

In this study, a plasmid, carrying ampicillin resistance (ampR) gene, isolated from a clinical isolate of Salmonella enterica serotype Typhimurium presenting ACSSuT (ampicilin, chloramphenicol, streptomycin, sulphonamide, tetracycline) resistance phenotype, was defined. The length of complete sequence of this plasmid was 8271 base pairs (bp), and it was named as pAnkS owing to its isolation place (plasmid-Ankara- Salmonella). The plasmid was analyzed for potential reading frames and structural features indicative of transposons and transposon relics. The Xmnl enzyme restriction fragments of pAnkS were cloned into E. coli plasmid vectors (pBSK), sequenced and analyzed with the BLAST programs. Plasmid pAnkS has contained a previously defined enterohemorrhagic E. coli (EHEC) plasmid p4821 as a core region and also contained a complete Tn3-like transposon of 4950 bp consisting of the left terminal repeat, Tn3-related tnpR and tnpA genes for transposition functions, ampicillin resistance gene bla(TEM), and the right terminal repeats, pAnkS showed strong homology with another Salmonella plasmid, pNTP16, for sequences that belong to p4821 and partial Tn3 segments. It was found that pNTP16 also carries kanamycin resistance gene (kanR) in addition to ampR gene. Plasmid pAnkS is one of the few completely sequenced plasmids from Salmonella Typhimurium and is in the middle of the pathway of evolution of plasmid from p4821 to pNTP16. The identification of pAnkS might help better understanding of plasmid evolution.

Detection of herpes simplex virus type 1 in addition to Epstein-Bar virus in tonsils using a new multiplex polymerase chain reaction assay.

HSV-1, HSV-2, CMV, EBV, which are the members of the herpes virus family colonize and establish latent infection in human. Although EBV is a well known virus most involved in recurrent bouts of acute tonsillitis, the role and possibility of HSV-1, HSV-2, and CMV for establishing infection in tonsils are not clear. The purpose of this study is to verify whether the tonsils might harbor the HSV-1, HSV-2, and CMV, in addition to EBV, in chronically hyperplastic nasopharyngeal lymphoid tissue. To accomplish the purpose, we developed a new Multiplex Polymerase Chain Reaction (M-PCR) assay using a single consensus forward primer and virus specific reverse primers for DNA polymerase gene of HSV-1, and 2, EBV, and CMV, and investigated its efficiency for detecting HSV1, HSV2, CMV, and EBV. The sample of 52 patients underwent tonsillectomy or adenectomy because of chronic lymphoid hyperplasia without any evidence of acute infections and were investigated for presence of HSV-1, HSV-2, CMV, and EBV. Of the 54 samples, 11 (20.4%) of them were positive for EBV, 4 of them (7.4%) were positive for HSV-1, and none of the samples were positive for HSV-2 and CMV. To the best of our knowledge, this is the first report that tonsils may be the reservoir for HSV-1 in addition to EBV, and HSV-1 may have a role in recurrent tonsillitis and systemic diseases. The MC-PCR assay presented in this study can provide a rapid, sensitive, and economical method for detection of HSV-1, HSV-2, EBV, and CMV in a single PCR tube.

Antimicrobial effect, in vitro, of gutta-percha points containing root canal medications against yeasts and Enterococcus faecalis.

OBJECTIVE: The objective of this study was to determine the in vitro susceptibility of Enterococcus faecalis and the most prevalent Candida species as therapy-resistant microorganisms to gutta-percha points containing root canal medications. STUDY DESIGN: Gutta-percha points containing calcium hydroxide (Calcium Hydroxide Plus Points), chlorhexidine diacetate (Activ Points), or calcium hydroxide-chlorhexidine combinations (CHX/Ca Combi Points) were tested for their ability to inhibit growth of pure cultures of Candida albicans, Candida glabrata, Candida parapsilosis, Candida krusei, Candida tropicalis, Saccharomyces cerevisiae, and Enterococcus faecalis. Approximately 2 x 10(7) microorganisms per assay were suspended in diluted human serum and co-incubated with the gutta-percha points placed in Eppendorf tubes in an incubator for up to 2 weeks. A tube was removed at 1, 2, 3, 4, 7, and 14 days, and then opened and microorganism suspensions were serially diluted in a sterile 0.9% NaCl solution. Aliquots of the dilution steps were streaked onto solid medium. After incubating the plates in an incubator at 37 +/- 1 degrees C for 48 hours, CFU numbers per milliliter of suspension were calculated. RESULTS: Calcium Hydroxide Plus Points or Activ Points did not exhibit sufficient antimicrobial or anticandidal activity for Enterococcus faecalis, Candida albicans, C. glabrata, C. parapsilosis, C. krusei, or C. tropicalis within 14 days. Only Saccharomyces cerevisiae was susceptible to the calcium hydroxide or chlorhexidine diacetate containing gutta percha points. CHX/Ca Combi Points killed C. albicans, C. glabrata, C. krusei, C. tropicalis, and S. cerevisiae completely. However, E. faecalis and C. parapsilosis were resistant to CHX/Ca Combi points within 14 days. CONCLUSION: The results show the gutta percha points containing a mixture of calcium hydroxide and chlorhexidine diacetate have efficacies superior to calcium hydroxide or chlorhexidine diacetate alone against some microorganisms except E. faecalis and C. parapsilosis.

Effects of probiotics on radiation-induced intestinal injury in rats.

OBJECTIVE: Radiotherapy is an important aspect of multimodal cancer therapy, but radiation-induced acute intestinal injury is a common and serious problem. Disruption of morphologic mucosal integrity and normal bacterial microflora after abdominal radiation leads to malabsorption and bacterial translocation. METHODS: Lactobacillus bulgaricus strain isolated from yogurt was given as a probiotic to rats subjected to radiotherapy. On postradiation day 8 rats were killed. Mesenteric lymph nodes, liver, and spleen were excised for microbiologic examinations. Segments of jejunum, ileum, and colon were evaluated for the presence of inflammation, vascularity, and mucus cells. RESULTS: The results of this study suggest that probiotics may have a protective effect on intestinal mucosa. CONCLUSION: Probiotics added as substrates can be given by an oral or enteral route to patients who undergo radiotherapy to prevent radiation-induced enteritis and related malnutrition.

[Investigation of the role of ribonuclease L gene in the response of tonsil tissues against viral infections]. / Tonsil dokusunun viral enfeksiyonlara karsi yanitinda ribonükleaz L geninin rolünün arastirilmasi.

Mutations or variants that impair function of ribonuclease L (RNase-L), particularly R462Q, have been proposed as susceptibility factors for the innate antiviral response. The aim of this study was to investigate and compare the expression levels of RNase-L and mutation of R462Q in the tonsils of tonsillectomy patients who were infected and not infected with herpes simplex virus type 1 (HSV-1), Epstein-Barr virus (EBV) and human herpes virus 6 (HHV-6). Six tonsils were included in the study. One tonsil was infected with all of these three viruses, two were infected with at least one of these viruses, and three were not infected with these viruses. The presence of viral DNAs in the tonsil tissues had been searched by polymerase chain reaction (PCR) in our previous study. Reverse transcriptase PCR method was used for RNase-L expression analyses, and single strand conformation polymorphism (SSCP) and direct sequencing methods were used for the mutation analyses. PCR products containing R462Q mutation site in the genomic DNA were used for SSCP analysis. In addition to SSCP analyses, partial sequencing of the cDNA PCR product containing R462Q mutation site were performed. As a result, no difference between the virus-infected and non-infected tonsils for the expressions of RNase-L were detected, and there were no mutations detected by SSCP and sequencing analyses. It was concluded that other factors than RNase-L protein, might be involved in the innate defense mechanisms of tonsil cells against viruses.