scholarly journals Investigation of Mycobacterium paratuberculosis in Arabian dromedary camels (Camelus dromedarius)

2019 ◽  
Vol 12 (2) ◽  
pp. 218-223 ◽  
Author(s):  
Mohamed A. Salem ◽  
Wael M. El-Deeb ◽  
Ahmed A. Zaghawa ◽  
Fadel M. Housawi ◽  
Ahmed M. Alluwaimi
Keyword(s):  
Mycobacterium Avium Subspecies Paratuberculosis ◽  
Culture Method ◽  
Staining Technique ◽  
Camelus Dromedarius ◽  
Mycobacterium Paratuberculosis ◽  
Tissue Samples ◽  
Specificity And Sensitivity ◽  
Severe Infections ◽  
Tissue Specimens ◽  
Direct Recognition

Aim: Mycobacterium avium subspecies paratuberculosis (MAP) causes Johne's disease in ruminants. This study aimed to investigate Mycobacterium paratuberculosis infection in clinically infected camels on the immunological, conventional bacteriological, and molecular biological basis. Materials and Methods: A total of 30 Arabian camels (Camelus dromedarius) were examined in this study. The camels were suffering from signs ranging from mild to severe infections (that did not respond to antibiotic treatment) to chronic or intermittent diarrhea. Camels were grouped into three groups based on their age, sex, and breed. Detection of anti-MAP antibodies in camels' serum, Ziehl-Neelsen (ZN) staining technique on rectal scraps, direct recognition of MAP in stool and tissue specimens by IS900 polymerase chain reaction (PCR) assay, and finally isolation and molecular description of MAP from fecal and tissue samples were carried out. Results: Five MAP isolates were recovered from these investigated camel samples giving an isolation rate of 16.6%, while eight camels were identified by PCR (26.6%). Five camels yielded MAP in their feces by ZN fecal staining (16.6%), whereas ELISA detected anti-MAP antibodies in nine camels only (30%). Conclusion: From the obtained results, we concluded that the gold standard for the diagnosis of MAP is the culture method despite its limitations. Molecular diagnosis (PCR) could be a useful tool in the identification of truly positive and negative camels; however, great care should be given regarding the primers specificity and sensitivity.

scholarly journals High sensitivity sanger sequencing detection of BRAF mutations in metastatic melanoma FFPE tissue specimens

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Lauren Y. Cheng ◽  
Lauren E. Haydu ◽  
Ping Song ◽  
Jianyi Nie ◽  
Michael T. Tetzlaff ◽  
...  
Keyword(s):  
Sanger Sequencing ◽  
Limit Of Detection ◽  
False Negative ◽  
High Sensitivity ◽  
Ffpe Tissue ◽  
Braf Mutations ◽  
Tissue Samples ◽  
Ffpe Samples ◽  
Droplet Pcr ◽  
Tissue Specimens

AbstractMutations in the BRAF gene at or near the p. V600 locus are informative for therapy selection, but current methods for analyzing FFPE tissue DNA generally have a limit of detection of 5% variant allele frequency (VAF), or are limited to the single variant (V600E). These can result in false negatives for samples with low VAFs due to low tumor content or subclonal heterogeneity, or harbor non-V600 mutations. Here, we show that Sanger sequencing using the NuProbe VarTrace BRAF assay, based on the Blocker Displacement Amplification (BDA) technology, is capable of detecting BRAF V600 mutations down to 0.20% VAF from FFPE lymph node tissue samples. Comparison experiments on adjacent tissue sections using BDA Sanger, immunohistochemistry (IHC), digital droplet PCR (ddPCR), and NGS showed 100% concordance among all 4 methods for samples with BRAF mutations at ≥ 1% VAF, though ddPCR did not distinguish the V600K mutation from the V600E mutation. BDA Sanger, ddPCR, and NGS (with orthogonal confirmation) were also pairwise concordant for lower VAF mutations down to 0.26% VAF, but IHC produced a false negative. Thus, we have shown that Sanger sequencing can be effective for rapid detection and quantitation of multiple low VAF BRAF mutations from FFPE samples. BDA Sanger method also enabled detection and quantitation of less frequent, potentially actionable non-V600 mutations as demonstrated by synthetic samples.

scholarly journals Evaluation of the Xpert MTB/RIF Performance on Tissues: Potential Impact on Airborne Infection Isolation at a Tertiary Cancer Care Center

2018 ◽  
Vol 39 (4) ◽  
pp. 462-466 ◽  
Author(s):  
Tracy McMillen ◽  
Shauna C. Usiak ◽  
Liang Hua Chen ◽  
Luz Gomez ◽  
Peter Ntiamoah ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Limit Of Detection ◽  
Tertiary Care ◽  
Ffpe Tissue ◽  
Cancer Center ◽  
Oncology Patients ◽  
Tissue Samples ◽  
Airborne Infection ◽  
Potential Impact ◽  
Tissue Specimens

OBJECTIVESIn this study, we sought to evaluate the performance of the Xpert MTB/RIF (Cepheid) assay for the detection of Mycobacterium tuberculosis (MTB) complex DNA on fresh and formalin-fixed, paraffin-embedded (FFPE) tissue specimens from oncology patients in an area with a low prevalence of tuberculosis. We also aimed to retrospectively assess the potential impact of Xpert MTB/RIF on the duration of airborne infection isolation (AII).SETTINGA 473-bed, tertiary-care cancer center in New York City.DESIGNA total of 203 tissue samples (101 FFPE and 102 fresh) were tested using Xpert MTB/RIF, including 133 pulmonary tissue samples (65.5%) and 70 extrapulmonary tissue samples (34.5%). Acid-fast bacilli (AFB) culture was used as the diagnostic gold standard. The limit of detection (LOD) and reproducibility were also evaluated for both samples types using contrived specimens. The potential impact of the Xpert MTB PCR assay on tissue samples from AII patients on AII duration was retrospectively assessed.RESULTSUsing the Xpert MTB/RIF for fresh tissue specimens, the sensitivity was 50% (95% CI, 1.3%–98.7%) and the specificity was 99% (95% CI, 94.5%–99.9%). For FFPE tissue specimens, the sensitivity was 100% (95% CI, 63.1%–100%) and the specificity was 98.3% (95% CI, 95.5%–100%. The LOD was 103 colony-forming units (CFU)/mL for both fresh and FFPE tissue specimens, and the Xpert MTB/RIF was 100% reproducible at concentrations 10 times that of the LOD. With an expected turnaround time of 24 hours, the Xpert MTB PCR could decrease the duration of AII from a median of 8 days to a median of 1 day.CONCLUSIONSThe Xpert MTB/RIF assay offers a valid option for ruling out Mycobacterium tuberculosis complex (MTBC) on tissue samples from oncology patients and for minimizing AII resource utilization.Infect Control Hosp Epidemiol 2018;39:462–466

scholarly journals Occult Pathologic Findings in Reduction Mammaplasty in 5781 Patients—An International Multicenter Study

2020 ◽  
Vol 9 (7) ◽  
pp. 2223
Author(s):  
Britta Kuehlmann ◽  
Florian D. Vogl ◽  
Tomas Kempny ◽  
Gabriel Djedovic ◽  
Georg M. Huemer ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Risk Factors ◽  
Histopathological Examination ◽  
Reduction Mammaplasty ◽  
Developed Countries ◽  
Preoperative Imaging ◽  
Breast Size ◽  
Tissue Samples ◽  
International Multicenter Study ◽  
Tissue Specimens

Breast cancer is among the most commonly diagnosed cancers in the world, affecting one in eight women in their lifetimes. The disease places a substantial burden on healthcare systems in developed countries and often requires surgical correction. In spite of this, much of the breast cancer pathophysiology remains unknown, allowing for the cancer to develop to later stages prior to detection. Many women undergo reduction mammaplasties (RM) to adjust breast size, with over 500,000 operations being performed annually. Tissue samples from such procedures have drawn interest recently, with studies attempting to garner a better understanding of breast cancer’s development. A number of samples have revealed nascent cancer developments that were previously undetected and unexpected. Investigating these so-called “occult” findings of cancer in otherwise healthy patients may provide further insight regarding risk factors and countermeasures. Here, we detail occult findings of cancer in reduction mammaplasty samples provided from a cohort of over 5000 patients from 16 different institutions in Europe. Although the majority of our resected breast tissue specimens were benign, our findings indicate that there is a continued need for histopathological examination. As a result, our study suggests that preoperative imaging should be routinely performed in patients scheduled for RM, especially those with risk factors of breast cancer, to identify and enable a primary oncologic approach.

Evaluation of Environmental Sampling Methods and Rapid Detection Assays for Recovery and Identification of Listeria spp. from Meat Processing Facilities

2009 ◽  
Vol 72 (4) ◽  
pp. 696-701 ◽  
Author(s):  
JOVANA KOVAČEVIĆ ◽  
VALERIE M. BOHAYCHUK ◽  
PABLO ROMERO BARRIOS ◽  
GARY E. GENSLER ◽  
DEANA L. ROLHEISER ◽  
...  
Keyword(s):  
Sampling Methods ◽  
Culture Method ◽  
Detection Methods ◽  
Lower Sensitivity ◽  
Meat Processing ◽  
Cotton Swab ◽  
Conventional Culture ◽  
Specificity And Sensitivity ◽  
Highly Sensitive ◽  
Labor Inputs

Studies that isolated Listeria spp. from the environment of two meat processing facilities were conducted. Samples were collected in the processing environment of the facilities with three different sampling methods (cotton swab, sterile sponge, and composite-ply tissues) to evaluate their ability to recover Listeria spp. A total of 240 samples for each sampling method were collected and tested. The cotton swab method of sampling was significantly (P < 0.01) less efficient in recovery of Listeria spp. than the sterile-sponge and composite-ply tissue methods, which were similar (P > 0.05) in their ability to recover Listeria spp. The specificity and sensitivity of four detection methods (conventional culture, Petrifilm Environmental Listeria Plates [ELP], lateral-flow immunoprecipitation [LFI], and automated PCR) were evaluated for identification of Listeria spp. Facilities were visited until a minimum of 100 positive and 100 negative samples per detection method were collected. The LFI and PCR methods were highly sensitive (95.5 and 99.1%, respectively) and specific (100%) relative to the culture method. The ELP method was significantly less efficient (P < 0.01) than LFI and PCR in detection of Listeria spp., with lower sensitivity (50.6%) and specificity (91.5%). Kappa values indicated excellent agreement of the LFI and PCR assays and moderate agreement of the ELP method to the culture method. Overall, ELP was easy to use but less efficient in detection of Listeria spp. from environmental samples, while the LFI and PCR methods were found to be excellent alternatives to culture, considering performance and time and labor inputs.

scholarly journals Comparison of a Commercial DNA Probe Test and Three Cultivation Procedures for Detection of Mycobacterium Paratuberculosis in Bovine Feces

1992 ◽  
Vol 4 (1) ◽  
pp. 23-27 ◽  
Author(s):  
Diana L. Whipple ◽  
Paul A. Kapke ◽  
Phil R. Andersen
Keyword(s):  
Positive Culture ◽  
Culture Method ◽  
Dna Probe ◽  
Fecal Culture ◽  
Mycobacterium Paratuberculosis ◽  
Culture Methods ◽  
Colony Forming Units ◽  
Probe Test ◽  
Centrifugation Method ◽  
Bovine Feces

Diagnosis of paratuberculosis using the IDEXX DNA probe test and 3 methods for cultivation of Mycobacterium paratuberculosis from fecal specimens were compared. Twenty-one of 170 fecal specimens were DNA probe test positive, whereas 35 specimens were positive by 1 or more of the cultivation methods evaluated. Four specimens were DNA probe test positive but were negative by fecal culture. The probe test detected M. paratuberculosis DNA in 62.9% of the specimens positive by a sedimentation culture method, in 56.6% of those positive by a centrifugation culture method, and in 65.4% of the specimens positive by the Cornell culture method. Specificity of the DNA probe test was approximately 97% relative to all culture methods. Generally, the probe test detected M. paratuberculosis DNA in fecal specimens from animals shedding at least 104 M. paratuberculosis colony forming units per gram of feces. Although the probe test did not detect all of the cattle shedding M. paratuberculosis, it was possible to identify cattle shedding the greatest number of organisms in 3 days compared with a minimum of 6 weeks required for positive culture results. The centrifugation method resulted in the most isolations of M. paratuberculosis after 12 weeks of incubation. However, contamination also was greatest when the centrifugation method was used. Contamination was best controlled using the Cornell method. The sedimentation method was the least time consuming and yielded results similar to those of the other 2 methods.

Enhanced Gross Visualization of Chicken Peyer's Patch: Novel Staining Technique Applied to Fresh Tissue Specimens

2006 ◽  
Vol 50 (2) ◽  
pp. 298-302 ◽  
Author(s):  
L. E. Vaughn ◽  
P. S. Holt ◽  
R. W. Moore ◽  
R. K. Gast
Keyword(s):  
Staining Technique ◽  
Peyer’S Patch ◽  
Peyer's Patch ◽  
Fresh Tissue ◽  
Tissue Specimens

scholarly journals Sonication versus Tissue Sampling for Diagnosis of Prosthetic Joint and Other Orthopedic Device-Related Infections

2018 ◽  
Vol 56 (12) ◽  
Author(s):  
Maria Dudareva ◽  
Lucinda Barrett ◽  
Mel Figtree ◽  
Matthew Scarborough ◽  
Masanori Watanabe ◽  
...  
Keyword(s):  
Tissue Culture ◽  
Relative Yield ◽  
Tissue Samples ◽  
Orthopedic Device ◽  
Joint Infections ◽  
Prosthetic Joint ◽  
Prosthetic Joint Infections ◽  
Tissue Specimens ◽  
Sonication Culture ◽  
Culture Yield

ABSTRACTCurrent guidelines recommend collection of multiple tissue samples for diagnosis of prosthetic joint infections (PJI). Sonication of explanted devices has been proposed as a potentially simpler alternative; however, reported microbiological yield varies. We evaluated sonication for diagnosis of PJI and other orthopedic device-related infections (DRI) at the Oxford Bone Infection Unit between October 2012 and August 2016. We compared the performance of paired tissue and sonication cultures against a “gold standard” of published clinical and composite clinical and microbiological definitions of infection. We analyzed explanted devices and a median of five tissue specimens from 505 procedures. Among clinically infected cases the sensitivity of tissue and sonication culture was 69% (95% confidence interval, 63 to 75) and 57% (50 to 63), respectively (P< 0.0001). Tissue culture was more sensitive than sonication for both PJI and other DRI, irrespective of the infection definition used. Tissue culture yield was higher for all subgroups except less virulent infections, among which tissue and sonication culture yield were similar. The combined sensitivity of tissue and sonication culture was 76% (70 to 81) and increased with the number of tissue specimens obtained. Tissue culture specificity was 97% (94 to 99), compared with 94% (90 to 97) for sonication (P= 0.052) and 93% (89 to 96) for the two methods combined. Tissue culture is more sensitive and may be more specific than sonication for diagnosis of orthopedic DRI in our setting. Variable methodology and case mix may explain reported differences between centers in the relative yield of tissue and sonication culture. Culture yield was highest for both methods combined.

Rapid Detection of Mycoplasma pneumoniae by Loop-Mediated Isothermal Amplification (LAMP) in Clinical Respiratory Specimens

2019 ◽  
Author(s):  
Maryam ARFAATABAR ◽  
Narjes NOORI GOODARZI ◽  
Davoud AFSHAR ◽  
Hamed MEMARIANI ◽  
Ghasem AZIMI ◽  
...  
Keyword(s):  
Mycoplasma Pneumoniae ◽  
Rapid Detection ◽  
Lamp Assay ◽  
Culture Method ◽  
Isothermal Amplification ◽  
Lamp Method ◽  
Culture Methods ◽  
Loop Mediated Isothermal Amplification ◽  
Specificity And Sensitivity ◽  
Respiratory Specimens

  Background: Mycoplasma pneumoniae is a common cause of community-acquired pneumonia (CAP) worldwide, especially among children and debilitated populations. The present study aimed to investigate a loop-mediated isothermal amplification (LAMP) technique for rapid detection of M. pneumoniae in clini-cal specimens collected from patients with pneumonia. Methods: Throat swabs were collected from 110 outpatients who suffered from pneumonia. Throat swab samples were obtained from patients referred to the hospital outpatient clinics of Tehran University hospitals, Iran in 2017. The presence of M. pneumoniae in the clinical specimens was evaluated by LAMP, PCR and culture methods. Sensitivity and specificity of the LAMP and PCR assays were also determined. Results: Out of 110 specimens, LAMP assay detected M. pneumoniae in 35 specimens. Detection limit of the LAMP assay was determined to be 33fg /μL or ~ 40 genome copies/reaction. Moreover, no cross-reaction with genomic DNA from other bacteria was observed. Only 25 specimens were positive by the culture method. The congruence between LAMP assay and culture method was ‘substantial’ (κ=0.77). Specificity and sensitivity of LAMP assay were 88.2%, 100% in compare with culture. However, the con-gruence between LAMP assay and PCR assay was ‘almost perfect’ (κ=0.86). Specificity and sensitivity of LAMP assay were 92.5%, 100% in compare with PCR. Conclusion: Overall, the LAMP assay is a rapid and cost-efficient laboratory test in comparison to other methods including PCR and culture. Therefore, the LAMP method can be applied in identification of M. pneumoniae isolates in respiratory specimens.

FEATURES OF THE PATHOMORPHOLOGICAL DIAGNOSIS OF MICROCRYSTALLINE ARTHROPATHIES IN THE PRACTICE OF SURGICAL MATERIAL EXAMINATION

2020 ◽  
Vol 58 (3) ◽  
pp. 286-289
Author(s):  
N. S. Migalkin ◽  
T. A. Stupina ◽  
A. V. Kaminsky ◽  
D. S. Mokhovikov ◽  
D. A. Shabalin ◽  
...  
Keyword(s):  
Staining Technique ◽  
Calcium Pyrophosphate ◽  
Alcohol Solution ◽  
Formalin Fixation ◽  
Histological Techniques ◽  
Tissue Sections ◽  
Stereo Microscope ◽  
Hematoxylin And Eosin ◽  
Tissue Specimens ◽  
Msu Crystals

The development of microcrystalline arthritides is most frequently associated with the formation of monosodium urate (MSU) and calcium pyrophosphate (CPP) crystals. Their identification is of crucial importance in recognizing these diseases. Objective: to determine the possibilities of histological techniques in identifying MSU and CPP crystals and to evaluate the effectiveness of the techniques. Subjects and methods. Twenty-four tissue blocks (fragments of the affected areas of the elbow joint, the interphalangeal joint of the index finger, and hip joint) from 7 patients were examined. Paraffin sections were stained with a 0.5% alcohol solution of eosin, as well as with hematoxylin and eosin. Tissue specimens were examined and digitized using an AxioScope.A1 stereo microscope with Zenblue software (Carl Zeiss MicroImaging GmbH, Germany). Results and discussion. When staining the tissue sections with hematoxylin and eosin, microcrystals were not visualized; the major portions of MSU crystals was dissolved during fixation and staining, whereas CPP crystals were masked with hematoxylin as focal basophilic aggregates. The staining technique with an alcohol solution of eosin and short formalin fixation (within 12 hours) made it possible to avoid dissolution of MSU crystals and to visualize both MSU and CPP crystals, and to determine their shape and color. Conclusion. Light microscopy of the tissue sections stained with an alcohol solution of eosin along with short formalin fixation is a reliable method to differentiate MSU and CPP crystals. In patients undergoing endoprosthetic replacement, the significance of this technique for the pathomorphological study of surgical material consists in assessing inflammatory activity and in eliminating a disease, such as microcrystalline arthropathy.

scholarly journals The Equine Endometrial Mast Cell during the Puerperal Period: Evaluation of Mast Cell Numbers and Types in Comparison to Other Inflammatory Changes

1997 ◽  
Vol 34 (1) ◽  
pp. 23-30 ◽  
Author(s):  
M. M. Welle ◽  
L. Audigé ◽  
J.-P. Belz
Keyword(s):  
Mast Cell ◽  
Mast Cells ◽  
Immunohistochemical Staining ◽  
Staining Technique ◽  
Cell Types ◽  
Postnatal Period ◽  
Double Labeling ◽  
Tissue Samples ◽  
Cell Numbers ◽  
Significant Difference

Endometrial biopsies of 44 broodmares were histologically examined on days 3, 6, and 9 postpartum. The mares were subdivided into three groups according to the course of the puerperal period. In 29 mares, parturition and expulsion of the placenta was normal, six mares showed dystocia, and in nine mares, the placenta was retained for >2 hours. Tissue samples were evaluated histologically, and the average numbers of granulocytes, lymphocytes, macrophages, siderophages, and mast cells was determined. Protease content of mast cells was examined with a double-enzyme immunohistochemical staining technique, using a histochemical reaction for chloroacetate esterase and fast blue to detect chymase activity and an immunohistochemical staining method with a polyclonal antibody and fast red for the detection of tryptase. Analyzing the cell numbers using the statistical software Statistica, a marked inflammatory reaction was observed in the endometrium postpartum. Although the number of granulocytes decreased during the first 9 days postpartum, the number of lymphocytes, macrophages, and siderophages increased. No significant difference in the number of any of these cell types could be demonstrated in the three different courses of the puerperal period, although the numbers of these cells seemed to be lower in mares with dystocia. In contrast with other cells, no change in the number of endometrial mast cells was observed during the puerperal period, but a significantly lower number were found in the endometrium of mares with retained placenta. The enzyme immunohistochemical double-labeling technique could demonstrate only tryptase-positive mast cells; no chymase activity was detectable in any endometrial mast cells. The number of mast cells detected with the metachromatic staining technique was significantly higher than that detected with double labeling. These results support the hypothesis that a sufficient number of mast cells may be necessary for a normal postnatal period and suggest a mast cell subtype in the equine endometrium that is tryptase and chymase negative.

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