• Parasites, Hosts and Diseases
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• v.56 no.2
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• pp.113-119
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• 2018

Cryptosporidium species is an important cause of gastrointestinal infections globally. This study aimed to shed light on its role in diarrheic immunocompetent patients in Beni-Suef, Egypt and to compare three diagnostic methods. Two hundred diarrheic patients, $37{\pm}16.8year$ old, were enrolled. Stool samples were examined by light microscopy, using modified Ziehl-Neelsen stain (MZN) for Cryptosporidium spp. oocysts. Coproantigens were detected by sandwich ELISA. DNA molecular diagnosis was done by nested PCR. PCR yielded the highest detection rates (21.0%), compared to ELISA (12.5%) and MZN staining method (9.5%). The higher infection rates were in 20-40 year-old group, followed by 40-60 year-old. Association between epidemiologic factors was statistically not significant; positivity and gender, clinical manifestations, residence, source or water, or contact with animals. Cryptosporidiosis is an important enteric parasitic infection in Beni-Suef and PCR remains the gold standard for diagnosis.

• Journal of Biomedical Engineering Research
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• v.23 no.3
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• pp.235-241
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• 2002

The detection of tubercle bacilli (TB) from sputum smear is one of the fast and inexpensive methods for diagnosis of tuberculosis. For this method. sputum smears are usually flexed by heating and stained by acid-fast staining method, and then examined under an optical microscope. Two Procedures are commonly used fur TB staining. One is hot staining and the other is cold staining method. The Ziehl-Neelsen method which is a hot staining method is widely used in Korea because its stained color is more vivid However, the conventional automated stainer has to fix the sputum smear on a slide manually and the stain is not so vivid because it has not heating function. In an effort to save labor and minimize variations in manual staining Procedure. we developed an automated stainer with heating function. The entire staining process is fully automated. from fixation to final washing and drying. With the automated methods, five slides can be flexed and stained in 21 minutes at consistent high quality We compared the concordance rate between the two methods for 91 sputum samples to validate the stain quality of the developed automated stainer. As the results, the concordant rate between the two methods was 95% and there was no significant difference (p>0.05)

• Biomedical Science Letters
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• v.23 no.2
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• pp.96-103
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• 2017

Nocardia species (spp.) are opportunistic pathogen in immunocompromised hosts. The genus Nocardia contains more than 70 species. Nocardia takedensis has been recently reported as a new species of the genus Nocardia. In this study, we describes the first clinical isolate of N. takedensis from the skin lesion in Busan, Korea. For the identification of clinical isolate to the species level as N. takedensis, classical methods (colony morphology, biochemical characteristics, and antimicrobial susceptibility), molecular method (16S rRNA gene sequencing), and MS (mass spectrometry) analysis were conducted. Clinical isolates grew slowly on the culture media (5% sheep blood agar and chocolate agar) under 5% $CO_2$ condition. Especially, carotene pigmentation was detected well on the media. Using mass spectrometry, Nocardia isolate was not identified to the species level. However, molecular method based on 16S rRNA sequencing confirmed the isolate as N. takedensis correctly. N. takedensis isolate was partial positive for acid-fast bacilli on the Ziehl-Neelsen method. And it was observed to be resistance to amoxicillin/clavulanic acid and ciprofloxacin. Our results provide useful information to develop optimal identification protocol of N. takedensis in clinical diagnostic laboratories.

• Parasites, Hosts and Diseases
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• v.50 no.1
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• pp.57-62
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• 2012

A total of 450 stool samples were collected from inpatient and outpatient clinics of Pediatric Department, Minia University Hospital, Minia District, Egypt. Two groups of patients were studied, including 200 immunosuppressed and 250 immunocompetent children. Stool samples were subjected to wet saline and iodine mounts. A concentration technique (formol-ether sedimentation method) was carried out for stool samples diagnosed negative by wet saline and iodine mounts. Samples were stained by 2 different methods; acid fast stain (modified Ziehl-Neelsen stain) and Giemsa stain. Total 188 cases (94%) were diagnosed positive for parasitic infections among immunosuppressed children, whereas 150 cases (60%) were positive in immunocompetent children ($P$<0.0001). The most common protozoan infection in immunosuppressed group was $Cryptosporidium$ $parvum$ (60.2%), followed by $Blastocystis$ $hominis$ (12.1%), $Isospora$ $belli$ (9.7%), and $Cyclospora$ $caytenensis$ (7.8%). On the other hand, $Entamoeba$ $histolytica$ (24.6%) and $Giardia$ $lamblia$ (17.6%) were more common than other protozoans in immunocompetent children.

• Korean Journal of Veterinary Research
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• v.47 no.4
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• pp.443-447
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• 2007

A 7-year-old, female snow goose (Anser caerulescens hyperboreus) with history of decreased activity for 2 month died in Daejeon Zoo Land in September 2006. At necropsy, granulomatous pneumonia and hepatomegaly with multiple cysts were observed. Small masses were found in the spleen. Microscopically, fibrinous pneumonia distributed in most of the lung lobe with pulmonary edema and congestion. Especially, granulomatous inflammation with numerous multinucleated giant cells was observed around the dilated bronchi. To confirm the diagnosis, acid-fast (Ziehl-Neelsen method) and periodic acid-Schiff (PAS) staining was performed. Acid-fast staining showed red bacterial colony indicating tuberculosis. PAS staining was also positive enough to diagnose aspergillus spp. co-infection that was an opportunistic fungi occurring in immuno-compromised animals. Based on the above results, we confirmed that the case submitted was diagnosed as avian tuberculosis.

• Biomedical Science Letters
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• v.22 no.1
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• pp.24-28
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• 2016

Tuberculosis (TB) remains an unsolved community health problem since identification of its causing microorganism called Mycobacterium tuberculosis (MTB) by Robert Koch in 1882. Annually, eight million TB cases are newly reported and 2~3 million patients die from TB. Pulmonary TB is highly infectious and untreated pulmonary TB patients are believed to infect >10 people in a year. The conventional methods for diagnosis of TB are chest X-ray and isolation of the causing microorganisms from patient specimens. Screening of TB is conducted with smeared sputum in slides, and TB is confirmed by identification of MTB in cultured specimens. One of the fatal pitfalls of screening detection for smeared sputum is that it is impossible to distinguish MTB and other acid-fast bacilli (AFB) because they are stained equally with Ziehl-Neelsen (ZN) stain. Culture of MTB is the most reliable method for diagnosis of TB but it takes 4~8 weeks. In this report, we suggest a fast and highly-reliable MTB detection method that distinguishes AFB in sputum samples. Purified DNA from the AFB stained slide samples offered by The Korean Institute of Tuberculosis were used to detect infected MTB in patients. PCR, real-time PCR and reverse blot hybridization assay (REBA) methods were applied to purified DNA. Conclusively, the real-time PCR method was confirmed to produce high sensitivity and we were able to further detect drug-resistant MTB with REBA.

• Biomedical Science Letters
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• v.19 no.2
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• pp.153-157
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• 2013

Although culture is the gold standard method to identify mycobacteria, its use in tuberculous lymphadenitis (TBL) is limited due to formalin fixation of the submitted specimens. We evaluated the performance of quantitative real-time PCR (q-PCR) for Mycobacterium Tuberculosis (MTB) in granulomatous lymphadenitis using formalin-fixed paraffin-embedded (FFPE) tissues. From 2000 to 2010, a total number of 117 cases of lymph node samples with granulomatous inflammation which were surgically removed and fixed in formalin were studied. Hematoxylin & Eosin (H&E) and Ziehl-Neelsen-stained (ZN) slides were reviewed. qPCR using Real TB-Taq$^{(R)}$ was performed for all cases to identify Mycobacterium tuberculosis. Thirteen non-tuberculous lymphadenopathy cases were used as negative control. Cervical lymph nodes were more frequently affected (60%, 70/117) than other sites. ZN stain for acid fast bacilli was positive in 19 (16.24%) cases. qPCR for tuberculosis was positive in 92 (78.63%) cases. Caseous necrosis was found in 103 (88.03%) cases. While the ZN stain and qPCR were both negative in all control cases, the qPCR showed a significantly higher positive rate (78.63% vs. 16.24%) compared to ZN stain in histologically diagnosed TBL. Quantitative real-time PCR proves to be more sensitive than ZN stain for diagnosis of tuberculous lymphadenitis.

• Korean Journal of Head & Neck Oncology
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• v.12 no.2
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• pp.169-176
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• 1996

Tuberculous cervical lymphadenitis is still an important cause of neck mass in Korea. Tuberculosis is an important differential diagnosis in patients of cervical lymphadenopathy. Rapid and sensitive test for the diagnosis of tuberculosis is essential for the approapiate treatment. Up to now, conventional diagnostic methods for M. tuberculosis were acid-fast bacilli(AFB) stain and culture of M. tuberculosis. The direct microscopic examination of AFB by Ziehl-Neelsen stain is rapid, but often negative. The culture for M. tuberculosis is time-consuming, taking 4 to 8 weeks. Recently various methods to detect Mycobacterial DNA, including PCR and restriction fragment length polymorphism(RFLP) analysis have been reported. Here we represent a simple method for the confirmation of M. tuberculosis and exclusion of the other Mycobacterial species by RFLP analysis and silver staining of polyacrylamide gel electrophoresis after nested PCR for a repetitive DNA sequence(IS986) specific for M. tuberculosis from fresh or paraffin-embedded biopsy specimens. This result leads us to conclude that this method is simple, rapid and possibly applicable to confirm M. tuberculosis and rule out the other Mycobacteria species from the clinical specimens in the clinical laboratories.

• Tuberculosis and Respiratory Diseases
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• v.79 no.4
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• pp.282-288
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• 2016

Background: Tuberculosis (TB) is a major health problem, and accurate and rapid diagnosis of multidrug-resistant (MDR) and extended drug-resistant (XDR) TB is important for appropriate treatment. In this study, performances of solid and liquid culture methods were compared with respect to MDR- and XDR-TB isolate recovery and drug susceptibility testing. Methods: Sputum specimens from 304 patients were stained with Ziehl-Neelsen method. Mycobacterium tuberculosis (Mtb) isolates were tested for recovery on $L{\ddot{o}wenstein$-Jensen (LJ) medium and the BacT Alert 3D system. For drug susceptibility testing of Mtb, isolates were evaluated on M-KIT plates and the BacT Alert 3D system. Results: The recovery rates were 94.9% (206/217) and 98.2% (213/217) for LJ medium and the BacT Alert 3D system, respectively (kappa coefficient, 0.884). The rate of drug resistance was 13.4% for at least one or more drugs, 6.0% for MDR-TB and 2.3% for XDR-TB. M-KIT plate and BacT 3D Alert 3D system were comparable in drug susceptibility testing for isoniazid (97.7%; kappa coefficient, 0.905) and rifampin (98.6%; kappa coefficient, 0.907). Antibiotic resistance was observed using M-KIT plates for 24 of the total 29 Mtb isolates (82.8%). Conclusion: The liquid culture system showed greater reduction in the culture period, as compared with LJ medium; however, drug susceptibility testing using M-KIT plates was advantageous for simultaneous testing against multiple drug targets.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.12 no.11
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• pp.5012-5018
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• 2011

According to the nationwide survey of tuberculosis from 1965 to 1995, the incidence and drug resistance rate of tuberculosis have been decreased in Korea, but the prevalence of multidrug resistance of Mycobacterium tuberculosis is still a serious problem. The purpose of this study is to investigate the drug resistance rate and pattern of tuberculosis in Daejeon from 2001 to 2008. Of the total 581cases where the drug susceptibility test was performed, resistance to at least one anti-TB drug was found in 104 cases(17.9%) of these, 68(11.7%) were resistant to at least INH and 41(7.1%) were resistant to at least RFP. Single-drug resistance was found for isolates from 37(6.4%) ; 18(3.1%) of these were resistant to INH and 5(0.9%) to RFP. Multidrug resistance, where TB was resistant to at least isoniazid and refampin, was found in 35 cases(6.0%). and Factors associated with MDR-TB included age under 40-60.The drug-resistance rate of pulmonary TB, especially MDR-TB, is higher in the initial treated patients at a private referral hospital than in those in the pubulic sector. Initial drug resistance is common and the drug susceptibility test is informative for pulmonary TB patients who have not received previous TB treatment. The need for an improved control program, coupled with early diagnosis of MDR-TB, to reduce the spread and development of resistance. Multidrug resistance rate is still problem in korea. Efforts to decrease multidrug resistance rate either independently or in cooperation with the pubulic sector will be needed.