Obstacles to progress in family planning.

Two dip-tests, Testuria and Uricult, were studied as screening tests for bacteriuria in children. Both tests showed a very good correlation with a standard method of urine culture.Testuria is easier to read and more accurate; however, it shows more “doubtful” results. Uricult presents the advantages of showing fewer “doubtful” results, and of differentiating between gram-positive and gram-negative organisms.The use of both tests should be limited to screening for bacteriuria. Certainly they are no substitute for accepted methods of urine culture.     

Comparison of multiplex PCR and standard bacteriological methods of detecting Salmonella on chicken skin

A multiplex PCR technique was compared with standard bacteriological techniques for the detection of Salmonella , and Salm. enteritidis in particular, in samples of chicken skin obtained from retail outlets. Five of 68 samples were positive by both systems (7.4%), 58 of 68 were negative by both systems (85%), five were positive by PCR only (7.4%) but none were positive by culture only. The PCR was quicker and more sensitive than the bacteriological methods, finding 15% of samples positive for Salmonella compared to 7.4%, and taking less than 24 h to reach a result as opposed to 2–3 d.     

Description and validation of a novel technique to study the bacterial flora of the teat duct of ewes

We describe a new technique that may be used for studying the bacterial flora of the teat duct in live ewes, and we present evidence regarding its accuracy, safety and repeatability. A sterile, plastic, 20 G catheter is used for sampling. The catheter stylet is taken out and the plastic catheter cut with a sterile blade to a length of 2 mm. In order to ensure accurate and consistent cutting of the catheter at the desired length, a sterilized ruler is placed beside the catheter. The whole procedure is carried out under aseptic conditions, on sterilized surgical screens. Before sampling, a thorough disinfection with povidone iodine scrub solution of the teat apex and the lower (1 cm) part of the teat skin is carried out. The catheter is held by the investigator from the cannula hub; then, it is inserted into the teat, rolled around the internal teat wall, in order to sample the mucosa and then withdrawn. Material collected on the tip of the catheter is plated onto bacteriological media. The sensitivity of the technique compared to a method that detects the bacterial flora of the teat after slaughter of the animals (scratching of the teat mucosa by means of a scalpel blade) was 99.1% for field samples and 100% for experimental samples; specificity of the technique was 90.7 and 100%, respectively, kappa coefficient between results of the two methods was 0.9045. The safety of the technique was assessed in an investigation, where ewes were repeatedly (eight times) sampled by means of this technique, throughout their lactation; in no case was clinical or subclinical mastitis recorded. Finally, the repeatability of the method was assessed in 37 paired samples, obtained 30–40 min apart; a 100% match was recorded between bacteriological results of the two samplings of each pair. We conclude that the method can be useful in research studies, in order to obtain information regarding bacterial ecology and dynamics inside the teat of ewes.     

细菌学方法和HOOF-Prints技术在绵羊种布鲁氏菌019株鉴定中的比较研究

应用细菌学常规方法和分子生物学检测方法对绵羊种布鲁氏菌非典型株019进行分类研究。利用高变8聚核苷酸DNA指纹技术(HOOF-Prints)对绵羊种布鲁氏菌019株可变数目重复片段(VNTR)的8个位点进行PCR扩增和序列测定,将测定结果与GenBank数据库比较分析,应用DNAMAN进行同源性分析,并构建系统进化树。结果表明,绵羊种布鲁氏菌019株和绵羊种布鲁氏菌参考株63/290的亲缘关系高于绵羊种布鲁氏菌019株与其他参考株的亲缘关系,该结论与细菌学常规鉴定结果一致。应用HOOF-Prints技术可以对绵羊种布鲁氏菌非典型株019进行鉴定,该技术有望弥补传统分类方法的不足。     

Screening methods for covert bacteriuria in schoolgirls.

Screening tests for bacteriuria based on two different principles were evaluated in1582 schoolgirls aged 5-11 years, and in 26 girls aged 3-16 years attending hospitalwith symptomatic urinary tract infection. Tests for hypoglucosuria, performed by a semi-automated fluorometric method and with Uriglox strips on early-morning urine samples voided after overnight fasting, gave unacceptably high false-negative rates (16.7% and 20.8% respectively). Oxoid and Uricult dipslides were immersed in fresh midstreamspecimens of urine obtained at school and read overnight incubation at 37 degrees C.Both gave comparable results, with low false-positive rates and no false-negative responses. The higher cost of screening by dipslides was halved by using the "dipstream" technique, which also gave no false-negative results. Its false-positive rate of 13.5% could be reduced to 1.8% by disregarding colony counts of 10-8 non-faecal organisms and over per litre, which appear unimportant in schoolchildren. Bacteriuria was found in 2.3% of the schoolgirls; 39% of them had symptons, compared with 7.2% of the healthy girls, and 25% showed vesicoureteric reflux, which in 17% was associated with renalscarring. Since the natural history of covert bacteriuria and its relationship withreflux and scarring remain undetermined further research is required. The dipstreamtechnique offers a simple, reliable, and comparatively cheap screening method which could also be applied in general practice.     

Detection of the tdh and trh genes in Vibrio parahaemolyticus isolates in fish and mussels from Middle Black Sea Coast of Turkey

Aim:  The aim of this study was to demonstrate the occurrence of potential pathogenic Vibrio parahaemolyticus in seafoods using DNA-based techniques in comparison with bacteriological methods.
Methods and Results:  From 120 fresh and processed fish and mussel samples collected from Middle Black Sea, 32 isolates were identified as V. parahaemolyticus by bacteriological methods and confirmed by tl gene-based conventional PCR. Of them, 13 isolates were found positive for only tdh gene, six isolates for only trh gene and 13 isolates for both genes by multiplex PCR.
Conclusions:  It is the first report demonstrating the presence of potential pathogenic V. parahaemolyticus isolates from the Black Sea seafoods by PCR detection of tl , trh and tdh genes that was found more rapid than bacteriological methods.
Significance and Impact of the Study:  This study confirmed the previous reports that characterization of potential pathogenic V. parahaemolyticus isolates based on the PCR techniques was reliable and cost-effective. These results suggest that molecular detection methods should be included in Turkish Standards of seafood control in addition to bacteriological methods.     

Eradication of Helicobacter pylori in patients with duodenal ulcer following the short course of treatment with azitromycin and amoxycillin]

The efficacy of short-term treatment with azithromycin in 17 patients with acute doudenal ulcer associated with H. pylori was evaluated. Bioptats of the gastric mucosa taken at the beginning and after one month of treatment were investigated for H. pylori presence by histological, bacteriological methods, by urease test and by PCR. All the patients with positive H. pylori test were treated with Gastrozol (omeperazole, ICN Pharmaceutical) 40 mg per day for 1 week followed by 20 mg per day for 3 weeks, Sumamed (azithromycin, peira) 0.5 g once daily for 3 days and amoxycillin 0.5 mg four times a day for 10 days. Bioptats analysis before treatment revealed H. pylori in 100% for PCR methods, in 94.1% for urease test, in 88.2% for histological test. After the treatment H. pylori was revealed in 12.5% for urease and histological test, in 18.8% for bacteriological test and in 25% for PCR test. Thus the treatment efficacy was 75%. Side effects for short-term azithromycin therapy were shown in 5.9% cases.     

Diagnosis methods for pneumococcal etiology determination in lower respiratory tract infections

A comparative study of etiological diagnosis in lower respiratory tract infections (LRTI), by conventional bacteriological methods and by pneumococcal antigen direct detection in sputum was performed. This work followed the establishing of rapid methods place, respectively of coagglutination (CoA), counterimmunoelectrophoresis (CIE), within the methodology of bacteriological diagnosis in lower respiratory tract infections presenting pneumococcal etiology. The results of investigations performed on 84 sputa from LRTI patients proved the utility of CoA method in determining a rapid etiological diagnosis, important for applying an emergence targetted antibiotherapy. CoA method, with the reagents in use, covering only 10 out of 83 serological types of S. pneumoniae in not capable of replacing conventional methods of bacteriological diagnosis; they complete each other, increasing the efficiency of etiological diagnosis in LRTI. CIE method is less sensitive and more difficult to perform, being less useful in rapid etiological diagnosis of LRTI.     

Comparison of conventional PCR, quantitative PCR, bacteriological culture and the Warthin Starry technique to detect Leptospira spp. in kidney and liver samples from naturally infected sheep from Brazil

Leptospirosis is an infectious disease of worldwide importance. The development of diagnostic techniques allows sick animals to be identified, reservoirs to be eliminated and the disease prevented and controlled. The present study aimed to compare different techniques for diagnosing leptospirosis in sheep. Samples of kidney, liver and blood were collected from 465 animals that originated from a slaughterhouse. The sera were analyzed by the Microscopic Agglutination Test (MAT), and kidney and liver samples of seropositive animals were analyzed using four techniques: bacteriological culture, the Warthin Starry (WS) technique, conventional PCR (cPCR), and quantitative PCR (qPCR). With the MAT, 21 animals were positive (4.5%) to serovars Hardjo (n=12), Hebdomadis (n=5), Sentot (n=2), Wolfii (n=1) and Shermani (n=1). Titers were 100 (n=10), 200 (n=2), 400 (n=6) and 1600 (n=3). No animal was positive by bacteriological culture; four animals were positive by the WS technique in kidney samples; six animals were positive by cPCR in kidney samples; and 11 animals were positive by qPCR, eight of which in kidney samples and three in liver. The bacterial quantification revealed a median of 4.3 bacteria/μL in liver samples and 36.6 bacteria/μL in kidney samples. qPCR presented the highest sensitivity among the techniques, followed by cPCR, the WS technique and bacteriological culture. These results indicate that sheep can carry leptospires of the Sejroe serogroup, and demonstrate the efficiency of quantitative PCR to detect Leptospira spp. in tissue samples.     

City-wide screening for urinary abnormalities in schoolboys

Screening for urinary tract infection was carried out in 27,722 schoolboys aged 5 to 14 using Uricult to perform urine cultures and Hema-combistix to detect hematuria, proteinuria and glycosuria. Cultures of 10⁵ colonies per ml or more on two occasions were found in 40 cases (0.14%), but no case was confirmed by the family physician using standard culture techniques.Proteinuria was found in 136 cases (0.49%) and confirmed in 47 (37%) of the 126 children who were seen by their family physician. In this group 8.8% had evidence of pyelonephritic scarring on intravenous pyelograms without a positive urine culture.Hematuria was found in 19 children and confirmed in 10 (59%) of the 17 children who were seen by their family physician. No abnormalities were detected on intravenous pyelography in any case.Glycosuria was found in 12 cases and confirmed in five. Three of these children had renal glycosuria and two had previously undetected diabetes.     

Evaluation of New Urinary Tract Infection Screening Devices

Several new methods for detection of bacteriuria were studied to evaluate their usefulness as screening procedures. A new filter paper device incorporating dehydrated media and tetrazolium was found to be reliable when compared with the standard pour plate method in the laboratory and with the dip-slide method in a field test. It failed to detect yeasts and slowly growing streptococci. Antibiotics blocked the test when susceptible organisms were present. An agar-cup method was found to be quite reliable, but could be improved by use of differential media. The Griess test was confirmed in a small trial to be highly specific when used in conjunction with a first morning specimen, but of little value with random specimens. Phenzopyridine was found to give false positive reactions. The subnormal glucose test, although highly sensitive and specific, gave too many false positive tests to be useful other than as a screening method.     

A dot-immunobinding assay for the laboratory diagnosis of tuberculous meningitis and its comparison with enzyme-linked immunosorbent assay.

In an attempt to establish an alternative to standard bacteriological methods in the laboratory diagnosis of tuberculous meningitis (TBM), a simple dot-immunobinding assay (Dot-Iba) was standardized to detect Mycobacterium tuberculosis antigen 5 and antimycobacterial antibody in cerebrospinal fluid (CSF) specimens of patients with TBM. Sensitivity and specificity of Dot-Iba was compared with conventional enzyme-linked immunosorbent assay (ELISA) and standard bacteriological techniques. The Dot-Iba showed excellent correlation with indirect ELISA for the detection of antimycobacterial antibody in CSF and showed 60% sensitivity and 100% specificity in culture-negative patients with TBM. However Dot-Iba was less sensitive for the detection of antigen 5 in CSFs and showed false negative results (60%) in culture-positive patients with TBM.     

Comparison of 2 methods of qualitative bacteriological urinalysis

Two techniques of the quantitative bacteriological urinalysis were compared. Hundred seventy eight samples of the urine were analysed with routine technique and paper strip test "Mast Bacteriuritest". Hundred percent conformity of both techniques was obtained in case of insignificant bacteriuria. In case of significant bacteriuria the results differed: paper tests were negative in 10% of cases. Significant bacteriuria was diagnosed in the samples in which Gould's test was positive with routine technique in 22% and with paper test in 18% of the analysed samples. It seems that paper test is valuable quantitative technique of the urinalysis because of its simplicity and low cost. It should be used, however, for the detection of the significant bacteriuria.     

多杀性巴氏杆菌分子分型方法简述

多杀性巴氏杆菌是一种能感染多种动物甚至是人的重要革兰氏阴性病原菌。目前临床上用于多杀性巴氏杆菌诊断的分型方法主要包括血清学分型方法和分子分型方法。其中血清学分型方法主要基于免疫学实验技术建立,操作过程繁琐,技术要求高,工作量大,不适用于临床上大规模快速开展多杀性巴氏杆菌流行病学调查的需要;而基于分子生物学手段建立的分子分型方法相对于传统的血清学分型方法而言具有快速、简单、灵敏、灵活等特点,特别是某些分子分型方法与传统的分型方法形成了较为精确的对应关系,因而在临床上得到了广泛的应用。目前适用于临床上开展多杀性巴氏杆菌分离鉴定的分子分型方法主要包括多重PCR方法及多位点序列分型法(MLST),其中多重PCR方法又包括基于荚膜编码区及脂多糖外核编码簇建立的PCR方法。本文将重点就这3种常用的多杀性巴氏杆菌分子分型方法进行综述,介绍其建立原理、实现手段以及各自的优缺点,为临床上开展多杀性巴氏杆菌的流行病学调查特别是分子流行病学调查提供参考。     

Usefulness of the latex test for rapid detection of Salmonella O antigen in bacterial culture fluids and clinical specimens. III. Diagnostic field studies]

The aim of this study was to evaluate a latex reagent prepared in our laboratory for a routine diagnosis of Salmonellosis in humans. Liquid cultures in selenite broth (SF) (18-24 hr), previously inoculated with faeces samples of individuals suspected of being infected with Salmonella were subjected to the study. In these cultures, after 15 min. of heating at boiling temperatures, group antigens of Salmonella with an aid of polyvalent latex reagent A-E and monovalent reagents B, C1, C2, D, and E were searched. The results of latex test were compared to the results obtained by routine bacteriological examination. Studies performed in 13 laboratories of Sanitary Epidemiological Stations included 5246 faeces samples. Out of these samples 1835 (35%) reacted with monovalent latex reagent and 1897 (36.2%) samples were positive, for Salmonella by culture technique and belonged to 14 genera of group B, C1, D, and E. S. enteritidis was the most frequently isolated and encountered for 98.6% of all isolated strains. Latex test with A-E reagent was positive in 2246 (42.8%) of culture samples in SF medium, of which 1736 were positive by culture and 510 samples were negative for Salmonella in routine bacteriological examination. The samples positive in culture and with A-E latex reagent reacted in 97.2% with one monovalent reagent. Out of bacteriologically negative samples and reacting with A-E latex reagent 28.8% were positive with monovalent latex reagents. In summary, we can conclude that latex test used in a survey studies can be an usefull test in addition to routine bacteriological examination, since after 18-24 hr it allows with high credibility of 95% to confirm or exclude Salmonella in a tested sample. Such a procedure due to a shortening of routine diagnostic course brings significant savings. Moreover, latex test makes possible rapid detection of mixed infections with Salmonella of different serological groups. The use of extremely carefully, properly prepared selenite broth constitutes a basic condition for agreement between results of latex test and routine bacteriological investigation.     

Early detection and rapid identification of Streptococcus pneumoniae and Haemophilus influenzae in acute pneumonias

The comparative study of the diagnostic value of the enzyme immunoassay (EIA), indirect immunofluorescence (IF) and countercurrent immunoelectrophoresis (CIE) was made. The serological identification of the isolated and reference pneumococci (19) and H. influenzae (38) strains revealed the possibility of using all three microanalytical methods for this purpose. The study of pneumococcal and H. influenzae antigens in native sputum obtained from 74 patients with acute pneumonia showed that EIA and indirect IF were highly sensitive, their sensitivity considerably exceeding that of the bacteriological analysis. Pneumococcal antigens were detected in 66.2% of patients by EIA and in 54.0% of patients by indirect IF, while H. influenzae antigens were detected in 58.1% of patients by EIA and in 67.6% of patients by indirect IF. The sensitivity of CIE proved to be considerably lower; in the detection of pneumococcal antigens it was level with the sensitivity of the bacteriological analysis (23.0%) and H. influenzae antigens could be detected only in 27.0% of patients.     

Typing of sheep clinical isolates and identification of enterotoxigenic Clostridium perfringens strains by classical methods and by polymerase chain reaction (PCR).

A simple procedure was developed to identify toxitypes of Clostridium perfringens of different origins. Ninety strains of C. perfringens were identified by classical bacteriological methods, typing of the strains was done by a seroneutralisation test on mice. Production of enterotoxin was tested and all strains were analysed by PCR using gene of toxin alpha, gene of toxin E, gene of toxin beta and gene of enterotoxin. Simple amplification (amplifying one gene), and duplex and triplex amplification (amplifying two and three genes simultaneously) were performed. In the conditions of the experiment, the PCR method has proved efficacious. The specificity and sensitivity are excellent and superior to those of the classical methods. The prophylaxis of enterotoxaemia in animals is achieved by vaccination, the PCR technique can thus become a first-choice tool for the identification and typing of the C. perfringens strains which initiate these diseases. In turn, this would simplify the development of vaccines adapted to the epidemiological situation.     

Microbiological diagnosis of suppurative-inflammatory processes of the E. N. T. organs and the maxillo-mandibulo-facial region under suspicion of anaerobic non-clostridial infection

The etiological role of non-sporulating anaerobic bacteria as causative agents of suppurative-inflammatory diseases (SID) of the maxillo-mandibulo-facial region and the E. N. T. organs as studied. Express diagnosis of anaerobic infection was carried out by means of gas-liquid chromatography. The species-specific composition of the microflora of the suppurative focus was investigated. It was established that only obligate anaerobes in monoculture or in association were isolated from patients with so-called "sterile" inoculations when strictly anaerobic bacteriological technique was used. The use of anaerobic bacteriological technique of investigation enlarged the spectrum of the microflora isolated from the suppurative focus from 5 to 26 species. Results of chromatographic and bacteriological examinations were compared; the main causes of obtaining false-positive and false-negative results of chromatography were analysed. Statistical processing using factor analysis has shown that the information power of chromatographic examination of the metabolites of anaerobic bacteria is higher in comparison with the main clinical-laboratory indices, but statistical processing using cluster analysis and correlation analysis has revealed that an index like metabolic activity reflects the degree of real participation of anaerobic microflora in the development of the pathological process, and can be used in the clinic for the evaluation of the degree of severity of the course of the SID and of the effectiveness of treatment.     

Improved method for sanitary bacteriological control in the epidemiological surveillance of suppurative-inflammatory diseases in maternity hospitals

To increase the information content of sanitary bacteriological control, the methods of planning laboratory investigations and their organization have been developed and the scheme of the bacteriological analysis of washings has been improved. On the basis of the improved method of bacteriological control, greatly varying data indicating the contamination of environmental objects in children's wards with indicator microorganisms (12.0-64.0%) have been obtained. The qualitative and quantitative characteristics of the microflora existing in the biotope of infants' skin have been studied on a new methodological level, and close correlation between the degrees of the microbial contamination of the skin and the incidence of purulent inflammatory diseases in newborn infants has been established. For the dynamic surveillance of the bacteriological situation in maternity hospitals a signal test, viz. the determination of the microbial contamination of the inner surface of swaddling clothes, has been proposed. This "swaddling clothes test" has made it possible to establish, for the first time, the microbiological characteristics indicating the degree of epidemic well-being in obstetric institutions.     

Utilization of an auxotrophic strain of the yeast Yarrowia lipolytica to improve γ-decalactone production yields

Here, hybridization assay of amplified products is described which detect Salmonella sp. from chicken fillets and other food homogenates within 24 h. This technique is composed of four steps : (1) sample is pre-enriched overnight in phosphate buffered peptone water ; (2) total DNA is extracted ; (3) a Salmonella spp. specific DNA target sequence is amplified by polymerase chain reaction ; (4) amplified products are captured by a probe covalently bound onto NH-Covalink^TM (Nunc, Danemark) microwells and detected by a chemiluminescent enzymatic reaction. This hybridization of amplified products was demonstrated as sensitive as their analysis on agarose gel. Compared to a bacteriological method for Salmonella spp. detection, its specificity was estimated at 100% and its sensitivity was 93·2% from analysis of 207 naturally contaminated chicken fillets samples.