Antifungal susceptibility and virulence of Aspergillus fumigatus environmental strains from a public tertiary hospital in Metro Manila, Philippines

@#<p><strong>Background and Objectives:</strong> The increase in the number of invasive Aspergillus infections has been observed among immunocompromised and hospitalized patients. In the Philippines to date, no published data focused on the prevalence of Aspergillus species or any other thermotolerant fungal species in a hospital environment. This research served as a primary study to characterize the antifungal susceptibility of environmental strains of Aspergillus fumigatus from a hospital facility against three antifungal agents and to determine the virulence of these isolates on BALB/c mice using an animal survival assay.<br /><strong>Methodology:</strong> Ten environmental strains of A. fumigatus were isolated from three air-conditioned wards in a medical facility using Andersen Air Sampler. The antifungal susceptibility profile of the isolates was determined against Voriconazole, Amphotericin B and Caspofungin. The virulence of these isolates was also tested on BALB/c mice using an animal survival assay. Moreover, the lung tissues of infected BALB/c mice were subjected to histopathological analyses using Gomori Methenamine Silver stain (GMS) and Hematoxylin & Eosin (H&E) stains.<br /><strong>Results:</strong> Etest result for antifungal susceptibility testing showed that two of the ten isolates were resistant to Amphotericin B (AF2-A and AF-3A); one isolate resistant to Voriconazole (AF2-A) and an isolate that manifested non- susceptibility to Caspofungin m(AF2-A). Epidemiological cut-off values were determined for each antifungal following the M38-A2 CLSI guidelines. BALB/c mice median survival analysis revealed that the isolate with the highest Minimum Inhibitory Concentration (MIC= 4.89 ?g/ml) for Voriconazole resulted in the most number of mortality with the least number of observation days. GMS AND H&E histopathology slides showed fungal elements embedded on left lung lobe of mice.<br /><strong>Conclusion:</strong> This study showed that there were strains of Aspergillus fumigatus from a hospital indoor air which were considered as resistant strains to Voriconazole, Amphotericin B, and Caspofungin (AF2-A and AF3-A). Lung tissues of infected mice showed characteristics of bronchopneumonia.</p>

Evaluation of Real-time PCR and Gomori-Methenamine Silver Stain for Diagnosing of Pneumocystis Pneumonia / 现代检验医学杂志

Objective To compare real-time PCR and gomori-methenamine silver stain in the diagnosis of pneumocystis peumonia (PCP).Methods 2 525 unrepeated specimens from suspected PCP patient admitted in Peaking Union Medical College Hospital were collected in 2014.2 492 samples were detected by gomori-methenamine silver stain,33 samples were detected by real-time PCR,and 429 samples were detected by both methods at the meanwhile.With clinical diagnosis as reference standard,the sensitivity,specificity,positive predictive value and negative predictive value of the two methods were analysised.Results Positive rate of gomori-methenamine silver stain was 1.2 ％ (30/2 492).The first three specimen types were sputum,tracheal intubation suction and bronchoalveolar lavage fluid,the positive rate was 0.70 ％ (13/1 845),4.00％ (10/250) and 2.72％ (7/257) respectively.Positive rate of realtime PCR was 34.20％ (158/462),and the positive rate of sputum and bronchoalveolar lavage fluid was 30.61％ (105/343) and 44.54％ (53/119) respectively.The sensitivity were 13.97％ vs 72.07％,specificity were 100％ vs 94.24％,positive predictive value were 100％ vs 92.14％ and negative predictive value were 55.36％ vs 78.26％ for gomori-methenamine silver stain and real-time PCR respectively.All of which were statistically significant analysed by x2 test for paired data.The x2 value and P alue were x2 =68.625,P＜0.01;x2 =4.296,P＜0.05;x2 =6.380,P＜0.01 and x2 =11.873,P＜0.01.Conclusion The real-time PCR had higher sensitivity,fewer interference factors and more clinical diagnostic value,so clinicians should make more use of real-time PCR to diagnose PCP earlier.

The Evaluation of Usefulness of PAS Stain and GMS Stain as Clinical Diagnostic Methods of Tinea unguium / 대한의진균학회지

BACKGROUND: Tinea unguium is a common problem seen in clinical practice. Considering the many differential diagnoses of dystrophic nails, it is important to make a definitive diagnosis of dermatophyte infection before the initiation of antifungal therapy. Potassium hydroxide (KOH) preparation and fungal culture, which are commonly used in the diagnosis of these infections, often yield false-negative results. Recent reports have suggested that nail plate biopsy (Bx) using periodic acid-Schiff (PAS) (Bx/PAS) stain may be a very sensitive technique for the diagnosis of tinea unguium. OBJECTIVE: The purpose of this study was to evaluate the usefulness of PAS and Grocott's methenamine silver (GMS) staining of nail specimen in the diagnosis of tinea unguium as a standard method. METHODS: We evaluated 75 nail specimens from suspected tinea unguium using KOH preparation, biopsy using periodic acid-Schiff stain, and Grocott's methenamine silver stain. RESULTS: Of the 75 nails which were negative on potassium hydroxide mounting, 43 and 39 cases were tested positive respectively on periodic acid-Schiff stain and Grocott's methenamine silver stain. CONCLUSION: Bx/PAS and Bx/GMS are the sensitive methods for the diagnosis of tinea unguium. They are indicated if clinical suspicion of onychomycosis is high and KOH preparation shows no fungal elements.

Transferrin Analysis by Immunofixation for The Diagnosis of Cerebrospinal Fluid Leakage / 대한임상병리학회지

BACKGROUND: CSF can be leaked from the nose or ear due to fractures, tumors or surgical procedures in the skull base region, and the threat of impending meningitis necessitates early identification of it. Since 2-transferrin occurs practically in cerebrospinal fluid (CSF) and not in other body fluid, its detection from the rhinorrhea or otorrhea can be used for the diagnosis of CSF leakage. We carried out immunofixation-silver stain (IF-SS) method for detection of 2-transferrin in the CSF in order to know optimal identification condition of specific cerebrogenic marker. METHODS: The fresh CSF sample was collected by spinal tapping. 2-Transferrin was estimated by quantifying the total transferrin by nephelomertry (Behring, Germany). 2-Transferrin of CSF was identified by electrophoresis using Titan gel high resolution protein system (Beckman, USA), immunofixation with anti-human transferrin antibody (Dako, Denmark) and then stained with silver nitrate. Serial dilutions of CSF were performed to know the detection limit of 2-transferrin. To know the influence of blood mixing, tests for mixed specimen of serum and hemolysate in CSF were performed. To evaluate the specimen storage condition, tests for different temperature and storage time were performed . RESULTS: By IF-SS method, identification limit of 2-transferrin was 0.5 mg/dL in 1:4 diluted CSF with distilled water. And 2-transferrin could be detected in condition of mixing serum protein (7.5 g/dL) or hemoglobin (13 g/dL) with CSF up to 6 : 4. At various sample storage condition, such as 37degrees C, room temperature, and 4degrees C, band intensity decreased abruptly after 1 day, and it was not detected 5 days later. Mean while, in -20degrees C and -70degrees C, 2-transferin band was detected after 10 days. CONCLUSIONS: IF-SS method was sufficiently sensitive and specific for invalidation by blood contamination, and seems to be used as effective identification of 2-transferrin in the CSF without sample concentration, less diagnostic test for CSF leakage.

Histochemical and Immunohistochemical Stain of Helicobacter pylori from the Gastric Mucosa / 대한내과학회지

No abstract available.

Studies on Un-fluorescent Staining Methods in Comet Assay / 环境与健康杂志

Objective To develop an un_fluorescent staining method in Comet Assay. Methods Some special un_fluorescent DNA staining methods, Feulgen, Methyl Green, Stains_all and Silver_Stain were tested in Comet Assay and the comparison studies with EB stain were carried out. Results Among the stain methods tested, just the silver_stain could give a satisfactory effect: legible, ease to judge the stained area of the comets head and tail, permanent, sensitive, could make little segment of DNA be stained, which couldnt be stained by EB stain, the length of comets head and tail, were over 2 times of that of EB stain, safety and economical. Conclusion The silver_stain method discarded the defects of EB stain, so that it could replace EB stain and make comet assay possible to be extended.

Nucleolar organizer regions of lymphomas in Korea

Nucleolar organizer regions (NORs) are loops of DNA which occur in the nucleoli of cells and which possess ribosomal RNA (rRNA) genes. The numbers and/or configurations of NORs have been thought to be related to cellular activities. To assess the applicability of NORs associated protein (Ag-NORs) in the field of diagnostic histopathology, a silver staining was done in paraffin sections of malignant lymphomas, tonsils and reactive lymph nodes and the numbers of Ag-NORs in the nuclei of low-grade and those of high-grade lymphomas were compared. A significant difference was found between the numbers of Ag-NORs in the nuclei of low-grade lymphoma (a mean of 1.3 per nucleus) and those of high-grade lymphomas (a mean of 4.2 to 8.3 per nucleus). The Ag-NORs were often observed in nuclei in areas where nucleoli themselves were not visible in H and E stain. It is suggested that this method would be of great value in the field of tumor histopathology.

Nucleolar organizer regions of lymphomas in Korea

Nucleolar organizer regions (NORs) are loops of DNA which occur in the nucleoli of cells and which possess ribosomal RNA (rRNA) genes. The numbers and/or configurations of NORs have been thought to be related to cellular activities. To assess the applicability of NORs associated protein (Ag-NORs) in the field of diagnostic histopathology, a silver staining was done in paraffin sections of malignant lymphomas, tonsils and reactive lymph nodes and the numbers of Ag-NORs in the nuclei of low-grade and those of high-grade lymphomas were compared. A significant difference was found between the numbers of Ag-NORs in the nuclei of low-grade lymphoma (a mean of 1.3 per nucleus) and those of high-grade lymphomas (a mean of 4.2 to 8.3 per nucleus). The Ag-NORs were often observed in nuclei in areas where nucleoli themselves were not visible in H and E stain. It is suggested that this method would be of great value in the field of tumor histopathology.

AMPLIFIED FRAGMENT LENGTH POLYMORPHISM ANALYSIS OF pYNZ22 LOCUS IN 120 CHINESES AND ITS APPLIFICATION TO FORENSIC SCIENCE PRACTICE / 中国法医学杂志

A region of the D17S30(pYNZ22) locus was amplified in DNA from 120 unrelated chinese individuals was carried out. The amplified products were analysed by mini-polyacrylamide gel electrophoresis followed by silver stain. 11 alleles, ranging from 170bp to 870bp in size and from 0. 4 % to 30. 4 % in frequency, was detected. The heterozygosity was 73%, Dp value was 0.938. The family study showed that the Amp-FLP in pYNZ22 locus is inherited according to the Mendelian law. The correct genotyping results can be obtained from very little amount of biological material, such as mixed stains, blood stains, single hair and saliva.

INNERVATION IN THE RAT THYMUS——AN INVESTIGATION BY SILVER STAIN AND ACETYLCHOLINESTERASE HISTOCHEMICAL METHOD / 解剖学报

The innervation in the thymus was observed by silver stain and acetylcho linesterase (AChE) histochemical method. It was shown by silver stain that nerve fibers constituted a complex network in the thymic medulla. In the cortex and medulla there were AChE-containing nerve fibers. which terminated near or encircled the thymocytes. AChE-containing nerve fibers contacted with mast cells. Some mast cells had AChE-containing substance. This study suggests that the cortex and medulla of thymus are innervated by parasympathetic cholinergic nerve fibers, which may regulate the thymocytes and mast cells.

THE ARGYROPHIL CELLS IN RAT GASTROINTESTINAL TRACT——A COMPARATIVE STUDY BETWEEN IMMUNOHISTOCHEMICAL AND SLIVER IMPREGNATION TECHNIQUES / 解剖学报

The aim of this study is to investigate comparatively the relationship between the endocrine and argyrophil cells in the rat gastrointestinal tract by means of the IGS, PAP immunohistochemical and Grimelius technique in the same sections. The results showed that the SS cells and almost G ceils can not be identified by Grimelius technique. The Grimelius technique had high specifity to reveal EC cells in the antrum of stomach and the intestine. Our technique is relatively simple and reliable, and may be used for consecutive demonstration of argyrophil cells and bioactive peptide-containing cells.