快速筛选菌尿症-尿涂片镜检法探讨

目的　探讨一种快速筛选菌尿的方法。　方法　用10μl尿液直接涂片,经革兰染色后镜检,同时取10μl尿液做细菌培养及菌落计数。　结果　总计检测1155份尿标本,其中显微镜检查阳性(每个油镜视野平均菌数≥2)且尿培养阳性(菌落计数≥10     

Rapid screening of urine for significant bacteriuria by Gram stain,acridine orange stain,and the Autobac MTS system

Urine specimens submitted for microbiologic examination were screened for evidence of bacteriuria by three rapid methods: Gram staining, acridine orange staining, and the Autobac MTS system. The screening results were compared with those obtained by semiquantitative colony counts on agar plates. In this comparative study 1055 urine specimens were examined, of which 146 (13.8%) had colony counts of ≥1 × 10⁵/ml. All thre urine screening methods detected this level of bacteriuria at a sensitivity of 98% and a specificity of 55.2% (acridine orange), 66.0% (Gram stain), and 83.2% (Autobac), respectively. Of the 1055 urine specimens examined, 185 (17.5%) had colony counts of ≥1 × 10⁴/ml, at which level the sensitivity of the three methods was 93% and the specificity was 56.7% (acridine orange), 68.0% (Gram stain), and 86.0% (Autobac), respectively. For any level of sensitivity, the Autobac urine screen was shown to be more specific than either the Gram stain or the acridine orange method. The acridine orange stain was the least specific urine screen, especially at the upper limits of sensitivity.     

阴道分泌物的细菌检查方法探讨

目的探讨妇产科阴道分泌物细菌检查的实验室检查方法,为阴道炎患者提供准确可靠的诊断依据。方法对昆明市第一人民医院门诊15 810例18～50岁已婚妇女的阴道分泌物进行生化快速酶法和革兰染色显微镜检查法。结果在15 810例妇女中细菌性阴道炎生化快速酶法占总例数的31.0%;其中唾液酸苷酶法占8.8%;β-葡萄糖醛酸苷酶法占11.0%;凝固酶法占11.2%。革兰染色显微镜检测法阳性占总例数的31.9%。革兰染色显微镜检查法分别与上述3种酶法检测对比的阳性率分别为9.0%、11.6%和11.3%。革兰染色镜检法分别与唾液酸苷酶法、β-葡萄糖醛酸苷酶法、凝固酶法相比较差异无统计学意义(P>0.05)。结论阴道分泌物细菌检查采用生化快速酶法与革兰染色显微镜检查法可以同时检测相互补充,以提高阴道分泌物细菌检查的准确率和阳性率。     

Is it meaningful to apply midstream urine culture to urine specimens with negative Gram stain results?

IntroductionGram staining is a convenient method for bacterial estimation. Urine culture is typically used to diagnose urinary tract infections. Therefore, urine culture is also performed on Gram stain-negative urine specimens. However, the frequency of uropathogen identification in these samples remains unclear.MethodsFrom 2016 to 2019, we retrospectively compared the results of Gram staining and urine culture tests on midstream urine specimens submitted for the diagnosis of urinary tract infections to confirm the significance of urine culture on Gram stain-negative specimens. Analysis was performed according to the patients’ sex and age, and the frequency of uropathogen identification in the culture was examined.ResultsA total of 1763 urine specimens (women, 931; men, 832) were collected. Of these, 448 (25.4%) were not positive on Gram staining but were positive on culture. In specimens without bacteria on Gram staining, the frequencies of specimens with uropathogens detected on culture were 20.8% (22/106) in women aged <50 years, 21.4% (71/332) in women aged ≥50 years, 2.0% (2/99) in men aged <50 years, and 7.8% (39/499) in men aged ≥50 years.ConclusionsIn men aged <50 years, the frequency of uropathogenic bacteria identification by urine culture was low in Gram stain-negative specimens. Therefore, urine cultures may be excluded from this group. In contrast, in women, a small number of Gram stain-negative specimens showed significant culture results for the diagnosis of urinary tract infection. Therefore, urine culture should not be omitted in women without careful consideration.     

Modified Field's staining—a rapid stain for Trichomonas vaginalis

Trichomonas vaginalis, a flagellate protozoan parasite commonly found in the human genitourinary tract, is transmitted primarily by sexual intercourse. Diagnosis is usually by in vitro culture method and staining with Giemsa stain. There are laboratories that use Gram stain as well. We compared the use of modified Field's (MF), Giemsa, and Gram stains on 2 axenic and xenic isolates of T. vaginalis, respectively. Three smears from every sediment of spun cultures of all 4 isolates were stained, respectively, with each of the stains. We showed that MF staining, apart from being a rapid stain (20 s), confers sharper staining contrast, which differentiates the nucleus and the cytoplasm of the organism when compared to Giemsa and Gram staining especially on parasites from spiked urine samples. The alternative staining procedure offers in a diagnostic setting a rapid stain that can easily visualize the parasite with sharp contrasting characteristics between organelles especially the nucleus and cytoplasm. Vacuoles are more clearly visible in parasites stained with MF than when stained with Giemsa.     

Evaluation of a rapid bacteriuria screening instrument

A newly modified, semiautomated instrument (Bacteriuria detection device (BDD) designed to detect the presence of bacteriuria in less than 3 min was compared to quantitative urine culture plating techniques. The instrument consists of a self-enclosed vacuum—filtration—staining system in which a 1-ml urine filtrate is stained on a filter. The resulting color determines the quantitation. Of the total of 525 clinical urine specimens tested, 66 (12.6%) were uninterpretable due to pigment deposition or inability to complete the filtering process (clogging of the filter). Of the remaining 459 specimens, 93 (20.3%) had a plate quantitation colony count of 10⁵ colony-forming units (CFU)/ml or more. The BDD detected 94.2% of these positive specimens if only significant pathogens were included (85% if specimens that were probably contaminated were also included). For specimens containing significant pathogens at 10⁴–10⁵ CFU/ml, the BDD detection rate was 41%. The device detected most (94.3%) gram-negative bacilli and enterococci at colony counts of 10⁵ CFU/ml or more. In addition, the BDD accurately detected 95.6% of specimens with no growth or fewer than 10⁴ CFU/ml. With several proposed modifications, these results suggest that this instrument is potentially useful as a urine screening device in a select population.     

Rhodamine-auramine O versus Kinyoun-carbolfuchsin acid-fast stains for detection of Cryptosporidium oocysts.

The rhodamine-auramine O stain was compared with the Kinyoun carbolfuchsin acid-fast stain for detection of Cryptosporidium oocysts in samples from patients infected with the human immunodeficiency virus (HIV). A total of 283 fecal specimens from HIV-infected patients were examined for the presence of Cryptosporidium oocysts. Duplicate smears of the fecal concentrates, prepared by the formalin ethyl acetate procedure, were stained by the Kinyoun carbolfuchsin and fluorescent rhodamine-auramine O acid-fast methods. The Kinyoun stain detected 13 positive specimens, while the rhodamine-auramine O stain detected 14 positive specimens. The average time required to survey a stained smear was 2.5 minutes with the fluorescent method, compared with 6.0 minutes with the Kinyoun technique. The rhodamine-auramine O stain is a dependable and efficient method of examining fecal smears for the presence of Cryptosporidium oocysts in a high-risk population.     

Comparison of Gram and Kopeloff stains in the diagnosis of bacterial vaginosis in pregnancy

Bacterial vaginosis (BV) is commonly diagnosed by using the Nugent score, a semiquantitative scoring system to evaluate bacterial morphotypes on Gram stain of vaginal secretions. Some authors have suggested using the Kopeloff modification of the Gram stain. Asymptomatic BV in pregnancy has been associated with adverse outcomes. We performed both stains on simultaneously collected vaginal smears from 2652 women at 24-26 weeks of gestation. Gram staining gave significantly higher (more abnormal) Nugent scores than Kopeloff staining. Compared to the Kopeloff stain, the number of specimens graded as indeterminate or consistent with BV by Gram stain increased by 29% (469 versus 364, P<.001). Interrater reliability of the Nugent score (n=413) for Kopeloff staining was significantly better than Gram staining (agreement=74% versus 63%, intraclass correlation coefficient=0.87 versus 0.79, P<.05, 95% confidence intervals 0.85-0.89 and 0.75-0.82, respectively).     

加德纳杆菌性阴道炎的细胞学诊断

应用单克隆抗体直接免疫荧光染色，在妇科门诊２４７例异常阴道分泌物涂片中检出加德纳菌感染者６例（２．４％），诊断为加德纳杆菌性阴道炎。阴道涂片中，乳酸杆菌缺少，炎细胞亦少见，经革兰、ＨＥ、巴氏、姬姆萨染色，这些涂片中均可查见＂线索细胞－ｃｌｕｅｃｅｌｌ，这是加德纳杆菌性阴道炎的特征性细胞学变化。     

The Ames Clinitek 200/Multistix 9 urinalysis method compared with manual and microscopic methods

We assayed 315 urine specimens by the Ames Clinitek 200/Multistix 9 semi-automated method and by corresponding standard methods (listed in parentheses) for the following analytes: protein (sulfosalicylic acid precipitation), glucose (Lilly Tes-Tape), ketones (Boehringer Mannheim Chemstrip K), leukocyte esterase (microscopic examination for leukocytes), blood (microscopic examination for erythrocytes), nitrite (microscopic examination for bacteria, Gram stain, or culture), and pH (pH meter). The Clinitek and standard methods agreed well at all concentrations for protein, ketones, and glucose. The Clinitek leukocyte esterase, nitrite, and blood methods were less sensitive than microscopic methods for detection of pyuria, bacteriuria, and hematuria, respectively. The Clinitek pH method produced falsely low pH values for urines with true pH less than 6.5, and falsely high values for urines with true pH greater than 6.5.     

The diagnosis of bacteriuria during pregnancy

Three diagnostic tests, Nitur, Urobact, and Uricult, were evaluated in the detection of bacteriuria in 865 pregnant women. As reference method agar culture was performed. Heavy growth (greater than 10(5) CFU/ml) of urinary tract bacteria was considered a true positive result and demonstrated in 58 (6.7%) of the women, 14 of whom had gram-negative rods. The sensitivity of the nitrite test was extremely low (0.13). The test gave negative results in eight of 17 specimens yielding heavy growth of Escherichia coli or Proteus mirabilis. Although the Urobact test was highly sensitive as regards gram-negative infection, it had an unacceptably low (0.27) predictive value in positive tests. The sensitivity of the Uricult test was low (0.35) in this study. The predictive value (0.50) of a positive test result may be acceptable, since just over half of the false positive results were explainable by moderate growth of urinary tract pathogens (10(4)-10(5) CFU/ml). It is argued that semi-quantitative urine culture may be preferable to the rapid diagnostic methods studied for the screening of bacteriuria in pregnant women.     

体液标本生理盐水涂片镜检在微生物学检验中的意义

目的探讨生理盐水涂片镜检在微生物学检验中的意义。方法用生理盐水涂片直接镜检结合革兰染色与培养结果进行综合分析。结果171例标本中，细菌培养的总阳性率为21．6％（37／171），革兰染色找到细菌的阳性率为27．5％（47／171），有较多活动细粒和白细胞的阳性率为75．4％（129／171）；在129例观察到有较多活动细粒和自细胞的标本中，革兰染色找到细菌的阳性率为36．4％（47／129），细菌培养阳性率为29．5％（37／129）；在革兰染色未找到细菌的82例标本中，细菌培养的阳性率为1．2％（1／82）。结论涂片直接观察结合革兰染色检查，可以对标本的性质（是细菌性的还是非细菌性的）作出快速的初步判断；涂片直接观察还可以对黏附力差的标本的革兰染色是否成功作出判断；盐水涂片直接镜检还有利于发现异形细胞、活动的原虫、寄生虫等。     

A method for improved fluorescent staining for acid fast smear microscopy by incorporating an acetone rinse step

Microscopic examination of the specimen smear for acid fast bacilli (AFB) provides a simple and rapid means of detecting AFB using fluorescent stain methods and remains a valuable diagnostic test used worldwide to identify and manage suspect cases of tuberculosis (TB). Methods to improve AFB smear staining protocols could provide better detection of suspect TB cases. In particular, decreasing background debris may improve the detection of smears with low numbers of bacilli. We assessed staining by the standard rack method compared to bulk container staining using an acetone rinse step to decrease background debris. No cross-contamination was observed in the bulk container staining, and higher accuracy with less reading time was achieved with the acetone rinse. Most importantly, more bacilli were detected per positive smear using the acetone rinse method.     

Utility of Gram stain in the clinical management of suspected ventilator-associated pneumonia. Secondary analysis of a multicenter randomized trial

PURPOSE: Gram stains of endotracheal aspirates (EA) and bronchoalveolar lavages (BAL) may guide empiric antibiotic therapy in critically ill patients with suspected ventilator-associated pneumonia (VAP). Previous studies differ regarding the ability of the Gram stain to predict final culture results. The aim of the present study was to evaluate the relationship between EA or BAL Gram stains and final culture results in intensive care unit patients with a suspected VAP. MATERIAL AND METHODS: We retrospectively analyzed data from the Canadian multicenter VAP study to correlate EA or BAL Gram stain and final culture results. We categorized Gram stains as Gram positive (GP) and Gram negative (GN) if any GP or GN organisms respectively were seen on staining. Cultures were considered "positive" if they yielded pathogenic organisms on final results. RESULTS: Seven hundred forty patients were enrolled in the study; 35 did not have a Gram stain done leaving 350 BALs and 355 EAs from 705 patients. Pooling BAL and EA results, we found the overall agreement between Gram stain class and pathogenic bacteria culture results to be poor (kappa = 0.36; 95% CI, 0.31-0.40). Among specimens with Gram stains showing no organisms, 99 (30%) of 331 grew pathogenic organisms. Among specimens with Gram stains showing no GN organisms, 113 (25%) of 452 grew pathogenic GN organisms. Among specimens with Gram stains showing no GP organisms, 45 (11%) of 428 grew pathogenic GP organisms. CONCLUSIONS: Gram stains performed for clinically suspected VAP poorly predict the final culture result and thus have a limited role in guiding initial empiric antibiotic therapy in such patients.     

Erroneous diagnosis of meningitis due to false-positive gram stains.

During a two-month period, 6% of Gram stains of predominantly CSF specimens revealed gram-negative bacilli with no growth. The source of the false-positive Gram stain results was an alcohol storage bath from which slides were taken and flamed to remove residual alcohol. All 11 patients in the outbreak had further diagnostic tests or changes in therapy. In the laboratory, false-positive slides could be created by contaminating a slide bath with 10(5) bacilli/ml, flame drying, and staining. In addition, contaminated crystal violet caused a false-positive result when applied to a warm but not to a cool slide. To prevent false-positive Gram stain results, the storage of slides in alcohol baths should be abandoned, slides should not be flamed to remove alcohol, and specimens should be Gram-stained only when the heat-fixed slide has thoroughly cooled.     

阳性血培养中阴性杆菌快速鉴定方法探讨

目的：结合荧光原位杂交法和直接VITEK法，快速鉴定阳性血培养中的阴性杆菌。方法：当BacT／ALERT3D120报警后，涂片革兰氏染色，若发现阴性杆菌，首先与大肠杆菌特异性的探针杂交，结果阳性者可直接报告为大肠杆菌；结果阴性者，将阳性血培养标本中的细菌离心后制成混悬液，采用VITEK直接法鉴定。所有菌株同时按标准法分纯后，采用VITEK标准法鉴定，比较两法的鉴定结果。结果：分离到的86株阴性杆菌中，在6h内可鉴定94％的细菌，90％为正确鉴定。结论：结合荧光原位杂交法和直接VITEK法，可在一天时间内报告鉴定阴性杆菌结果，并可将其鉴定至种的水平，有助于病人的早期选用抗生素。     

Performance of a rapid yeast test in detecting Candida spp. in the vagina

We compared the performance of a rapid vaginal yeast assay (Savvycheck) with that of microscopic examination of a Gram-stained smear and culture of vaginal discharge in detecting Candida spp. Two hundred thirty-one women with vaginal symptoms were studied prospectively. Vaginal specimens obtained from all participants were studied by the Savvycheck rapid yeast test, microscopic evaluation of Gram-stained vaginal smears, and yeast culture. Savvycheck rapid yeast test was positive in 79% of women with positive cultures and in 3.6% of women with negative cultures. The Savvycheck test detected yeasts in 93% of subjects with positive Gram stain and in 5.5% of subjects with negative Gram stain. The Savvycheck rapid yeast test showed 93% sensitivity, 95% specificity, and a 97% negative predictive value compared with the Gram stain. It revealed 79% sensitivity, 96% specificity, and an 87% negative predictive value compared with culture. The Savvycheck rapid yeast test can be used in the busy office instead of microscopy as a point-of-care tool for diagnosing vulvovaginal candidiasis. It can also reduce the need for yeast cultures in patients with vaginitis.     

眼科术后感染患者病原菌分布及其药敏性分析

目的研究眼科术后感染患者病原菌的分布情况,并分析其对各抗菌药物的药敏性,为临床及时、合理使用抗菌药物提供医学依据。方法选取2010年2月至2014年12月就诊于本院眼科的术后感染患者60例,通过涂片获得患者眼部病原菌,经革兰染色和药敏试验,针对病原菌的分布进行分析。结果检出的病原菌为革兰阴性菌、革兰阳性菌和真菌。有50.00%患者被检出革兰阳性菌,其中表皮葡萄球菌占21.67%;25.00%的患者被检出革兰阴性菌,其中铜绿假单胞菌占10.00%;有25.00%的患者被检出真菌。对病原菌进行药敏试验,革兰阴性菌中铜绿假单胞菌对阿米卡星、舒巴坦钠、他唑巴坦、亚胺培南、美罗培南的敏感性均大于或等于83.30%;革兰阳性菌中表皮葡萄球菌和金黄色葡萄球菌对利奈唑安、万古霉素和环丙沙星的敏感性均大于或等于75.00%。结论眼科术后患者感染的病原菌,革兰阴性菌、革兰阳性菌和真菌均有,对常用青霉素等有较高的耐药性,但不同的病原菌都有各自敏感性较高的药物,临床上有针对性地、及时地选择有效的抗菌药物进行预防或治疗,从而降低眼科术后的感染率。     

泌尿系感染中肠杆菌科细菌的分布及耐药性

目的 分析本院泌尿系感染中肠杆菌科细菌的分布及耐药性,指导临床医生合理使用抗菌药物.方法 按照《全国临床检验操作规程》的要求操作,细菌鉴定采用常规方法和API鉴定系统,药敏试验采用K-B纸片扩散法.结果 大肠埃希菌是泌尿系感染的主要病原菌,多数肠杆菌科细菌对亚胺培南、头孢他啶、头孢哌酮/舒巴坦敏感,而对氟喹诺酮类抗菌药物敏感率较低.结论 临床医生应重视细菌培养和耐药性检测,根据药敏报告合理选用抗生素,防止和减少耐药菌株的产生和流行.     

Strategies for the avoidance of bacterial contamination of blood components

Gram staining and bacterial culturing methods were used to determine the incidence of bacterial contamination of cellular blood components at the time of transfusion reactions. Over a 5-year period, 2208 (4.3%) of 51,278 transfusions were complicated by reactions. Overall bacterial contamination occurred in 5 (0.03%) of 17,928 transfusions of single- donor apheresis platelets, 1 (0.14%) of 712 transfusions of pooled random-donor platelet concentrates, 1 (0.003%) of 31,385 transfusions of red cells, and 0 of 1253 transfusions of fresh-frozen plasma. Gram staining done at the time of positive cultures was positive in three of six cases. Although six of seven recipients of contaminated components suffered no clinical sequelae, contaminated transfusions may have been a contributing cause of death in one case. Attempts were made to avoid the transfusion of contaminated cellular blood components by performing routine bacterial cultures: 0 of 341 quality control cultures were positive. To avoid the transfusion of contaminated platelets by identifying bacteria, Gram staining was performed in all single-donor apheresis platelet units collected on open systems and daily in platelets stored > 48 hours: 8 (0.15%) of 5334 smears done on 3829 platelet units were interpreted as positive, and those units were not transfused, but only two of eight units were culture positive. These studies suggest that bacterial contamination can result in adverse clinical sequelae in transfusion recipients and that both culturing and Gram staining are poor methods of screening for contaminated units. More sensitive and specific methods of generalized screening for bacterial contamination are needed.