Use of fluorescence resonance energy transfer for rapid detection of isoniazid resistance in Mycobacterium tuberculosis clinical isolates.

SETTING: Rapid detection of drug resistance in Mycobacterium tuberculosis is important to select effective treatment and prevent transmission of resistant isolates. OBJECTIVE: To evaluate the use of fluorescence resonance energy transfer (FRET) for rapid detection of isoniazid (INH) resistance in M. tuberculosis clinical isolates. DESIGN: One hundred INH-resistant and 50 INH-susceptible isolates of M. tuberculosis were included in the study. The drug susceptibility of all isolates was determined by the standard agar proportion method, and all isolates were then tested by FRET. Three genes associated with INH resistance, katG, inhA and ahpC, were analysed. All isolates were amplified with three pairs of primers. Three pairs of fluorescently labelled DNA probes specific to codon 315 of katG, nucleotide 209 in the regulatory region of inhA and a frequent mutation site in the intergenic region of oxyR-ahpC, were used for mutation detection. RESULTS: The results obtained using FRET were compared with those from the proportion method. The sensitivity and specificity of FRET were respectively 76% and 100%. The frequencies of mutations were 48% in katG, 17% in inhA, 8% in ahpC, 2% in inhA-ahpC and 1% in inhA-katG. CONCLUSION: FRET is a rapid, specific method that can be useful to detect INH resistance in M. tuberculosis clinical isolates.

Epidemiology of chronic Pseudomonas aeruginosa infections in cystic fibrosis.

Chronic lung infection with Pseudomonas aeruginosa is primarily responsible for pulmonary deterioration of cystic fibrosis patients. The purpose of this study was to type the P. aeruginosa isolates collected sequentially from cystic fibrosis patients, chronically colonized with P. aeruginosa, by random amplified polymorphic DNA fingerprinting-PCR (RAPD-PCR). Sequential P. aeruginosa isolates (n: 130) that had been collected from 20 CF patients over at least 9 years were investigated. The isolates were analyzed by RAPD-PCR using two arbitrary primers. Antimicrobial susceptibility testing of all isolates was performed by the disc diffusion method. RAPD-PCR typing demonstrated that strains dissimilar in colony morphotype and of different antibiotic susceptibility patterns could be of the same genotype. Some CF patients were colonized with a rather constant P. aeruginosa flora, with strains of different phenotypes but of one genotype. However, some patients may be colonized with more than one genotype. The results also demonstrated that there might be a risk of cross-colonization between CF patients followed-up at the same center.

Neutrophil chemotaxis in acutely infected and clinically stable cystic fibrosis patients.

PURPOSE: The aim of the present study was to evaluate the role of neutrophil chemotaxis in cystic fibrosis (CF) and to also determine whether an acute bacterial infection and the nutritional status of a child can affect neutrophil chemotaxis. METHODS: Twelve acutely infected and 12 clinically stable CF patients and 10 healthy age-matched controls were studied. Neutrophil chemotaxis and random migration were investigated in vitro in the peripheral blood of subjects by the Boyden chamber method and the results were expressed as chemotactic index (CI). The nutritional status of the cases was evaluated as body mass index (BMI). RESULTS: The CI values in the acutely infected group were found to be significantly lower than the clinically stable and healthy control groups (P < 0.05 and P < 0.005, respectively). There was no significant difference between the clinically stable CF group and the healthy control group (P > 0.1). No significant correlation was detected between the CI and BMI of the two groups of CF patients (P > 0.05). CONCLUSIONS: The present study confirms that neutrophil chemotaxis and random migration are normal in clinically stable CF patients. The decreased CI in the acutely infected patients indicates the possible role of infection itself on neutrophil chemotaxis.

Cryptosporidiosis in immunocompromised patients in a Turkish university hospital.

The prevalence of Cryptosporidiosis in 18 immunosuppressed diarrheic patients was evaluated by examination of fecal samples by direct staining (Modified Kinyoun and Giemsa), direct and indirect immunofluorescence methods. Forty patients (10 nondiarrheic immunosuppressed, 10 nondiarrheic immunocompetent, and 20 diarrheic immunocompetent) were included in the study as the control group. 11 of 18 samples were positive for cryptosporidial oocysts by at least one of the methods. Oocysts were detected in all (n = 7) of the AIDS patients. This high frequency was attributed to a probable nosocomial infection. None of the samples from control subjects were found positive for Cryptosporidium. Our results indicate that Cryptosporidial oocysts should be detected particularly in immunosuppressed patients with diarrhea. Modified Kinyoun staining method is practical and reliable for this purpose. Immunofluorescence staining methods can be applied for confirmation of the results.

Detection of Listeria monocytogenes in milk by the polymerase chain reaction.

A on the polymerase chain reaction (PCR) method based was developed for detection of Listeria monocytogenes in milk samples after enrichment culture. It consists of culturing samples in Listeria enrichment broth, followed by DNA extraction and detection of the organism using PCR. Dilutions of L. monocytogenes in milk were subjected to PCR amplification after enrichment culture. When determining the sensitivity of the method, it was found to be possible to detect 37 CFU (colony forming unit gl/ml) of the bacterium in milk. The method was assessed as a sensitive, specific, times-saving and practical way of detecting L. monocytogenes in milk samples.

Visceral and cutaneous leishmaniasis (report of 2 cases).

Leishmaniasis is an endemic, sporadic infection in many parts of the world. Turkey is geographically unique in linking Asia and Europe. Of special interest is leishmaniasis, as various forms of this disease have long been reported in the surrounding regions. Visceral and cutaneous leishmaniasis are endemic in the western and southeastern parts of Turkey, respectively. Here, we report a cutaneous and a visceral leishmaniasis case, to draw attention to the increase in the incidence of leishmaniasis in Turkey. In the patient with cutaneous leishmaniasis, the ulcerative lesion on the cheek had persisted for two months before admittance to the hospital. Direct smears prepared from this lesion were negative for leishmania amastigotes whereas the promastigote forms were maintained in NNN (Novy-MacNeal-Nicolle) medium. The second patient was hospitalized with a prediagnosis of haematological malignancy, but the smears prepared from the bone marrow aspirates revealed leishmania amastigotes and promastigotes were seen on the smears from NNN cultures. These two reports mark the importance of inoculation of the specimens to NNN medium for the recovery of the promastigote forms. Cutaneous and visceral leishmaniasis have become endemic in considerable number of foci in Turkey, possibly due to the cessation of vector control programmes and increase in the agricultural and irrigation areas. These two reports also point out the increased prevalence of leishmaniasis in Turkey after 1980's.

Cryptosporidium parvum prevalence in a group of Turkish children.

Stool samples from two hundred children with diarrhea and from 50 healthy children were examined, by modified Kinyoun's acid-fast staining (MAF), Giemsa staining and direct (DFA) and indirect immunofluorescence antibody (IFA) methods, in order to determine cryptosporidiosis prevalence under the age of 12 and to detect the most efficient identifying method for use in our country. Cryptosporidium oocysts were detected in seven (3.5%) of the cases. None of the samples from the control subjects was found to be positive for Cryptosporidium. Our results indicate that Cryptosporidial oocysts should be detected in children with diarrhea. Modified Kinyoun staining method is practical and reliable for this purpose. Immunofluorescence staining methods can be applied for conformation of the results, if available.

Rate of carriage, serotype distribution and penicillin resistance of Streptococcus pneumoniae in healthy children.

This study was aimed to define the carriage rates for Streptococcus pneumoniae in a given population in Ankara and also to determine the serotypes and penicillin resistance of these strains. Oropharyngeal swabs were taken from a total of 661 children aged between 0-11 years and living in a province of Ankara between January 1995-January 1997. Serotyping was performed by detection of the Quellung reaction. The isolates were screened for penicillin susceptibility by the agar dilution method according to the guidelines of NCCLS. The total rate of pneumococcal carriage in the study population was 23.90% and the isolation rate was found to be statistically associated with age, being higher in small children. Among the 158 S. pneumoniae isolates, the most prevalent serotypes (in order of frequency) were 6, 19, 9, 23, 3 and 14. Penicillin susceptibility was examined in 120 of the isolates. 55 of them (45.83%) were susceptible, 53 (44.17%) were intermediately and 12 (10.0%) were highly resistant to penicillin. Evaluation of the results showed that serotypes 6, 14 and 23 were those most often associated with penicillin resistance. The significant rate of isolation of penicillin-resistant pneumococci in healthy carriers points to the importance of active immunization in risk groups and also the importance of the rational use of antibiotics to limit the spread of resistant strains.

Detection of Listeria in raw and pasteurized milk.

One hundred raw milk samples from different regions of Anatolia and 20 pasteurized milk samples from three different manufacturers in Ankara were analyzed for the presence of Listeria spp. L. monocytogenes was found in 1% of the raw milk samples and in 5% (1/20) of the pasteurized milk samples. L. innocua and L. seeligeri were found in 8 and 2% of the raw milk samples, respectively. No other species of Listeria was found. The overall incidence of Listeria spp. was 10% in the raw milk samples and 5% in the pasteurized milk samples.

Trends in antimicrobial resistance of Streptococcus pneumoniae in children in a Turkish hospital.

The antimicrobial susceptibilities of 143 isolates of Streptococcus pneumoniae recovered from the sputa of children with lower respiratory tract infections in a Turkish university hospital were determined. Five isolates (3.5%) were resistant and 57 isolates (39.9%) intermediately resistant to penicillin. The most common serotype among these resistant isolates was serotype 23, followed by serotypes 19 and 14. The overall resistance rates were 31% for tetracycline, 11% for erythromycin and cefaclor, 4% for chloramphenicol, 2% for cefotaxime and 0% for vancomycin. The data highlight the need for surveillance of resistance and serotype distribution of S. pneumoniae in our geographical area.

Isolation, in vitro antimicrobial susceptibility and penicillin tolerance of Arcanobacterium haemolyticum in a Turkish university hospital.

Arcanobacterium haemolyticum (Ah) was isolated from 5 (0.3%) out of 1531 throat cultures of patients with presumed pharyngotonsillitis. The age of the patients who had a positive culture for Ah varied between 6 and 22. The isolation rate of beta-haemolytic streptococci (BHS) was 7.4%, 72.6% of which belonged to Group A, followed by groups G, C and B. None of the throat samples yielded simultaneous growth of Ah and BHS. Antimicrobial susceptibility of Ah isolates to phenoxymethylpenicillin, cephalexin, cefotaxime, vancomycin, erythromycin, azithromycin, doxycycline, ciprofloxacin, and trimethoprim-sulfamethoxazole was tested by the agar dilution method. The isolates were found to be susceptible to all antimicrobials tested except trimethoprim-sulfamethoxazole. Penicillin tolerance could be detected in none of the Ah strains, including the reference strain Ah ATCC 9345. We conclude that Ah should be kept in mind as a potential pathogen causing pharyngitis in adolescents and young adults.

[Determination of the immune potency of lymphocyte-promoting factor (LPF) and filamentous hemagglutinin (FHA) molecules by intranasal challenge in mice]. / Lenfositozis promoting faktör (LPF) ve filamentöz hemaglütinin (FHA) moleküllerinin bagisiklama gücünün farelerde buruniçi challenge yoluyla belirlenmesi.

All mice immunized with toxoid LPF and FHA molecules were found to be protected against infections by intranasal instillation of Bordetella pertussis. While a significant weight loss was observed in control mice within ten days following challenge, immunized mice were found to gain weight. Mice immunized with 20 micrograms doses of toxoid LPF were found to be also protected against intracerebral infection. FHA did not protect mice against such an infection.

[Brain abscess bacteriology and antibiotic sensitivity]. / Beyin abselerinin bakteriyolojisi ve antibiyotiklere duyarlilik durumlari.

In this study pus samples obtained during operation from 28 patients with brain abscess have been examined microbiologically. In three of the 28 patients the cultures were negative (10%) obligate anaerobic bacteria from 20 (80%) patients, microaerophilic bacteria from 2 (4%) patients, aerobic bacteria from 11 (44%) patients have been isolated. Among the 15 obligate bacteria and 2 microaerophilic bacteria, the highest resistance has been shown against tetracycline, among the 11 aerobic bacteria the highest resistance has been shown against Gentamicin and Tetracycline.

[HBeAg and anti-HBe levels in HBsAg positive blood donors]. / HBs antijeni pozitif donörlerde HBe antijen ve antikor düzeyi ile ilgili bir arastirma.

Serum samples from 110 blood donors who were carriers of hepatitis B surface antigen (HBsAg) were examined for the presence of hepatitis Be antigen (HBeAg) and its antibody. HBeAg was detected in 21 (19.09%) and anti-HBe in 87 (79.09%) blood donors. No significant association were found with blood groups and HBeAg or anti-HBe. The analysis of serum samples for HBeAg and anti-HBe at the blood transfusion centres, will be useful for determining if HBsAg carriers are infectious.

[Synthetic peptide antigens for medical cell biology]. / Sentetikik peptidlerin antijen olarak kullanilmasi.

The research in cell biology and medical sciences have been advanced through the use of antibodies specifically reactive with cellular proteins. In addition to conventional polyclonal and monoclonal antibodies, a third type of antibodies has come wide spread use within the past five years. Synthetic peptide antigens have been shown to elicit antibodies that can react with full length protein containing that peptide. Such antibodies are directed against a specific region of the protein chosen in advance by the investigator and so have a predetermined specificity. In basic research, these antibodies are usefull in identifying the protein product to particular cells or subcellular organelles and purifying the protein by immunoaffinity chromatography techniques. In medicine, such antibodies may provide reagents for passive vaccination, antitoxin therapy and targetted immunotherapy of neoplasia.

[Actinomyces and other bacteria isolated from cervical cultures of women using IUDs]. / RIA kullanan kadinlarin serviks kültürlerinden enfeksiyon etkeni olarak izole edilen actinomyces türleri ve diger bakteriler.

In this study we performed endocervical cultures from 517 patients with infection. In 75 of these 517 cultures (14.5%) Actinomyces has been grown. The incidence of Actinomyces has increased in correlation with IUD period, but the type of IUD has not affected the incidence of Actinomyces. Actinomycosis of uterus is usually superficial. It rarely spreads and causes pelvic actinomycosis. Even it can change to systemic infection. For this reason, in women who have used IUDs for long period Actinomyces infection should be kept in mind. Clinicians should be in corporation with the laboratories, this is important for identification of the microorganism and for the right therapy.

[Anaerobic bacteria in various infections and their antibiotic susceptibility patterns]. / Anaerobik bakterilerin çesitli infeksiyonlardaki rolü ve antibiyotiklere duyarlilik durumlari.

In this study we have examined antibiotic susceptibility of anaerobic bacteria isolated from various samples of 75 patients. Anaerobic bacteria from 22 (29.3%) patients, and only aerobic bacteria from 29 (38.7%) patients have been isolated; there has been no growth in cultures from 24 (32%) patients. Of 75 isolates from 51 patients 38 (50.6%) are anaerobic and 37 (49.4%) are aerobic bacteria. Following anaerobic bacteria have been isolated: Peptostreptococcus (13), Peptococcus (5), microaerophilic streptococci (2), Veillonella (1), Bacteroides (9), Fusobacterium (4) and non spore forming anaerobic gram positive bacilli (4). Among the 38 anaerobic bacteria the highest resistance has been shown against tetracycline (65.8%). Resistance rates for cephalothin, erythromycin and penicillin G are 28.9%, 15.8% and 7.8%. In our investigation we have shown no resistance against carbenicillin, clindamycin and chloramphenicol.

[In vitro susceptibility of gram-negative bacteria isolated from urinary tract infections to acylureidopenicillins]. / Idrar yolu enfeksiyonlarindan izole edilen gram negatif bakterilerin acylureido penicillin'lere olan duyarliliginin in vitro saptanmasi.

Ureidopenicillins exert antibacterial activity on many Gram negative bacteria and also some Gram positive bacteria Two acylureido penicillins called azlocillin and mezlocillin have been introduced recently into medical practice in many countries as broad spectrum antibiotics. In this communication we present the sensitivity of Gram negative bacteria isolated from urinary tract infection of pediatric age patients to azlocillin, mezlocillin, another semisynthetic penicillin called pipercillin and aminoglycoside antibiotic gentamicin by MIC and disk-diffusion methods. The results are discussed in the view of the use of new semisythetic penicillins in clinical practice.

[Antibiotic susceptibility of E. coli strains isolated from urinary tract infections and the role of metabolically deficient strains in these infections]. / Idrar yolu enfeksiyonlarindan izole edilen E. coli'lerin çesitli antibiyotiklere duyarliliklari ve bu enfeksiyonlarda metabolik defektli suslarin rolü.

In this study; E. coli strains isolated from 100 patients with urinary tract infections were investigated for their antibiotic susceptibility and metabolic deficiencies. The strains were found to be highly susceptible to gentamicin (%97), Bactrim (R) (%77) and resistant to the other antibiotics in changing but important degrees. Study of metabolic deficiencies revealed that 9 of the strains were in accord with the deficiency criteria and only one of them was thymin-dependent.

[Importance of vaccination and bacteriological studies in diphtheria epidemics]. / Difteri epidemilerinde asilama ve bakteriyolojik incelemelerin önemi.

The Ouchterlony - Elek gel diffusion test was performed in twenty-nine patients having a positive culture result for diphtheria bacilli on Loeffler and tellurite agars in January and February, 1984 and toxin production was detected in 20 of them. Except one adult patient, the cases ranged in age 2 and 12 years. In 14 patients who have received at least one immunization with diphtheria vaccine had no complication, besides three patients without immunization in whom two received antibiotics plus antitoxin therapy had carditis and soft palate paralysis. Five patients in the non-immunized group and 6 patients in the group received at least one immunization had diphtheritic membrane. In spite of three patients with cardiac and neural complications, with early diagnosis and beginning therapy with antibiotics and antitoxin before the result of in vitro toxigenicity test, O% mortality rate was obtained.