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Tsung C Chang - One of the best experts on this subject based on the ideXlab platform.

  • bacterial bioburden decrease in orthokeratology lens storage cases after forewarning assessment by the dna Dot Hybridization assay
    Eye & Contact Lens-science and Clinical Practice, 2017
    Co-Authors: Pochiung Fang, Jung Lo, Tsung C Chang, Chunchih Chien, Changchun Hsiao, Shinling Tseng
    Abstract:

    BACKGROUND: The aim of this study was to measure the changes in the bacterial bioburden in orthokeratology (OK) lens storage cases using the DNA Dot Hybridization assay (DHA) after forewarning patients about their bacterial contamination severity. METHODS: Thirty-one OK lens wearers were prospectively enrolled in this study. Dot Hybridization assay was used for serial measurements of bacterial bioburden in OK storage cases after lenses had been soaked for approximately 6 hr. After the first assessment, the lens wearers were informed of the extent of case contamination and the possible risk of microbial keratitis (MK), and best practices for lens care and lens case hygiene were reviewed and reinforced. A second assessment by the same DHA method was performed after approximately 6 months. RESULTS: Two universal bacterial probes confirmed a significant decrease in bacterial bioburden at the second assessment (P<0.01 and P<0.001). Genus-specific probes showed significant reductions in Acinetobacter and Klebsiella (P=0.02 and P=0.01), but not in Pseudomonas (P=0.42). CONCLUSIONS: Making OK lens wearers aware of the bacterial bioburden in their lens cases resulted in improved quality of case care and reduced bioburden. Our results suggest that a strategy of bioburden assessment with forewarning could be a useful method to decrease the incidence of OK-related MK.

  • a multiplex Dot Hybridization assay for detection and differentiation of acanthamoeba and herpes keratitis
    Investigative Ophthalmology & Visual Science, 2016
    Co-Authors: Mingtse Kuo, Chunchih Chien, Shinling Tseng, Changchun Hsiao, Pochiung Fang, Yuhsuan Lai, Tsae Ling Chao, Shun Hua Chen, Jen Ren Wang, Tsung C Chang
    Abstract:

    PURPOSE We verified a multiplex Dot Hybridization (MDH) assay for the rapid detection and differentiation of Acanthamoeba keratitis (AK) and herpes simplex keratitis (HSK). METHODS Molecular detection of Acanthamoeba and herpes simplex virus in corneal scrapes was performed with the MDH assay and standard diagnostic methods. The experimental group included corneal scrapes (n = 33) from patients with culture- or pathology-confirmed AK (n = 15) and real-time PCR-confirmed HSK (n = 16). The control group included 50 samples from cases of bacterial keratitis (n = 15), fungal keratitis (n = 15), and initially presumed AK (n = 5) or HSK (n = 17) which finally were excluded by culture for Acanthamoeba or by real-time PCR for herpes simplex virus, respectively. Discrepant results between methods were resolved by DNA sequencing of the PCR amplicons. RESULTS After discrepant analysis, the sensitivity for the diagnosis of AK and HSK was both 93.3% by the MDH assay, while the specificity was 100% for the two types of keratitis. The turnaround time of MDH assay was within a working day using an already prepared array. Two false-negatives (one AK case and one HSK case) were obtained by the MDH assay. CONCLUSIONS The MDH assay could effectively prevent missed or delayed diagnosis of AK and HSK and has a potential to be adopted in routine clinical practice if the test is commercialized.

  • a dna Dot Hybridization model for assessment of bacterial bioburden in orthokeratology lens storage cases
    Investigative Ophthalmology & Visual Science, 2015
    Co-Authors: Chunchih Chien, Jung Lo, Shinling Tseng, Changchun Hsiao, Pochiung Fang, Tsung C Chang
    Abstract:

    PURPOSE: The aim of this study was to evaluate a DNA Dot Hybridization assay (DHA) for assessing bacterial bioburden in orthokeratology lens (OK) storage cases. METHODS: Forty-one OK wearers participated in this study. The Dot Hybridization assay was used to assess the bacterial bioburden of OK after removal and 6-hour soaking in a storage case. Signals of the DHA were standardized after gray image transformation. The correlations between the Hybridization intensities of three universal bacteria probes (BP1, BP2, and BP3) and bacterial bioburden determined by culture (colony forming units per milliliter) was analyzed by Pearson's correlation coefficient and receiver operating characteristic plots. In addition, three genus-specific probes for Pseudomonas, Acinetobacter, and Klebsiella were used to detect potentially hazardous bacterial contamination regardless of bacterial viability status. RESULTS: Among the three universal probes, there were good correlations between probe BP2 (r2 = 0.31, P = 9.5 × 10(-5)) and probe BP3 (r2 = 0.35, P = 3.1 × 10(-5)) with bacterial bioburden, but no correlation was found between probe BP1 and bacterial bioburden (r2 = 0.04, P = 0.11). In 41 samples, one was Pseudomonas-positive by both DHA and culture, while 10 were Pseudomonas-positive by DHA but negative by culture. Furthermore, nine samples tested positive for Acinetobacter (n = 7) and Klebsiella (n = 2) by DHA only. CONCLUSIONS: The Dot Hybridization assay provides a novel way to assess the bacterial bioburden of OK storage cases. Lens care quality can be assessed with universal bacteria probes, while potentially hazardous bacterial contamination can be traced with genus-specific probes.

  • a highly sensitive method for molecular diagnosis of fungal keratitis a Dot Hybridization assay
    Ophthalmology, 2012
    Co-Authors: Mingtse Kuo, Chunchih Chien, Pochiung Fang, Hsienchang Chang, Chao Kun Cheng, Tsung C Chang
    Abstract:

    Purpose Fungal keratitis (FK) is an important cause of ocular morbidity, especially for people living in the agricultural communities of the developing world. Current diagnostic methods may lack sensitivity (direct microscopy) or are time consuming (culture). The aim of this study was to develop a Dot Hybridization assay for sensitive and rapid diagnosis of FK. Design Evaluation of diagnostic test or technology. Participants and Controls Fifty corneal scrapes (49 patients) from consecutive cases of clinically suspected microbial keratitis were analyzed prospectively. Methods Molecular detection of fungi in the scrapes was performed by amplification of the internal transcribed spacer region (ITS) that contained the target gene (5.8S rRNA gene) by polymerase chain reaction (PCR), followed by Hybridization of the PCR product to a fungus-specific oligonucleotide probe immobilized on a nylon membrane. The results were compared with those obtained by gram-stain microscopy, culture, and gel electrophoresis of the PCR products. Discrepant results were resolved by cloning and resequencing of the amplified ITS fragments. Main Outcome Measures Performance of the Dot Hybridization assay, including sensitivity, specificity, and positive and negative predictive values, was evaluated. Results Ten scrapes demonstrated positive results by both the Dot Hybridization assay and culture. However, 11 scrapes demonstrated positive results by the Dot Hybridization assay, but demonstrated negative results by culture, and 10 of the 11 samples were considered to be positive for FK by cloning and resequencing of the amplified ITS fragment and by a pathologic examination or clinical course review. The sensitivities for diagnosis of FK by the Dot Hybridization assay and culture were 100% and 50%, respectively, whereas the specificities were 96.7% and 100%, respectively. Conclusions The Dot Hybridization assay is a highly sensitive and specific diagnostic tool for FK. The method provides a much higher sensitivity than that of culture (100% vs. 50%; P Financial Disclosure(s) The author(s) have no proprietary or commercial interest in any materials discussed in this article.

Pochiung Fang - One of the best experts on this subject based on the ideXlab platform.

  • molecular bioburden of the lens storage case for contact lens related keratitis
    Cornea, 2018
    Co-Authors: Yuting Hsiao, Jiunnliang Chen, Tsailing Chao, Hunju Yu, Pochiung Fang, Yuting Huang
    Abstract:

    PURPOSE: To elucidate the bioburden of a lens care system in patients with contact lens (CL)-related keratitis. METHODS: In this prospective case-control study, the CL storage case was used as the target of bioburden assessment. Participants were CL wearers with (n = 26) or without (n = 24) keratitis in southern Taiwan. The case fluid sample was consecutively collected and assessed using a Dot Hybridization assay (DHA) for bioburden assessment. The DHA was composed of 3 universal bacterial probes for detecting all bacteria, genus-specific probes for Pseudomonas and Acinetobacter, and 1 probe for Acanthamoeba. The receiver operating characteristic curve was used to analyze the differentiating performance of a probe. RESULTS: The storage case bioburden was significantly higher in those with CL-related keratitis than in control subjects; patients with CL-related keratitis had stronger standardized signals in all universal bacterial probes. Moreover, in the cases of these patients, the bioburden was significantly heavier in confirmed infectious keratitis than in presumed noninfectious keratitis. Receiver operating characteristic analysis showed that stronger signals in all 3 universal bacterial probes and the Pseudomonas probe might suggest that the wearers have infectious keratitis. Only the storage case of the Acanthamoeba keratitis case showed positive detection by the Acanthamoeba probe. CONCLUSIONS: A heavier bioburden in the lens storage case was associated with a higher risk of CL-related keratitis and infectious keratitis. Inappropriate maintenance of the CL will lead to microbial contamination and transfer the pathogen onto the ocular surface causing keratitis accordingly. The DHA assessment for the lens storage case might provide an alternative way to differentiate infectious from noninfectious CL-related keratitis.

  • bacterial bioburden decrease in orthokeratology lens storage cases after forewarning assessment by the dna Dot Hybridization assay
    Eye & Contact Lens-science and Clinical Practice, 2017
    Co-Authors: Pochiung Fang, Jung Lo, Tsung C Chang, Chunchih Chien, Changchun Hsiao, Shinling Tseng
    Abstract:

    BACKGROUND: The aim of this study was to measure the changes in the bacterial bioburden in orthokeratology (OK) lens storage cases using the DNA Dot Hybridization assay (DHA) after forewarning patients about their bacterial contamination severity. METHODS: Thirty-one OK lens wearers were prospectively enrolled in this study. Dot Hybridization assay was used for serial measurements of bacterial bioburden in OK storage cases after lenses had been soaked for approximately 6 hr. After the first assessment, the lens wearers were informed of the extent of case contamination and the possible risk of microbial keratitis (MK), and best practices for lens care and lens case hygiene were reviewed and reinforced. A second assessment by the same DHA method was performed after approximately 6 months. RESULTS: Two universal bacterial probes confirmed a significant decrease in bacterial bioburden at the second assessment (P<0.01 and P<0.001). Genus-specific probes showed significant reductions in Acinetobacter and Klebsiella (P=0.02 and P=0.01), but not in Pseudomonas (P=0.42). CONCLUSIONS: Making OK lens wearers aware of the bacterial bioburden in their lens cases resulted in improved quality of case care and reduced bioburden. Our results suggest that a strategy of bioburden assessment with forewarning could be a useful method to decrease the incidence of OK-related MK.

  • a Dot Hybridization assay for the diagnosis of bacterial keratitis
    Molecular Vision, 2017
    Co-Authors: Pochiung Fang, Chunchih Chien, Shinling Tseng, Yuhsuan Lai, Mingtse Kuo
    Abstract:

    PURPOSE To evaluate a bacterial Dot Hybridization (BDH) assay for the diagnosis of bacterial keratitis (BK). METHODS Sixty-one qualified corneal scrapings from 61 patients with suspected microbial keratitis were collected consecutively and prospectively. Among the 61 patients, 16 cases were BK and 45 cases were non-BK, including fungal keratitis, viral keratitis, parasitic keratitis, and non-microbial keratitis. Molecular diagnosis of BK in these corneal scrapes was performed using the BDH assay with three universal bacterial probes (PB1, PB2, and PB3) and three genus-specific probes (Aci, Klb, and Psu) to detect Acinetobacter, Klebsiella, and Pseudomonas, respectively. Signals were standardized after grayscale image transformation for objective validation using receiver operating characteristic (ROC) curves. RESULTS The standardized intensities for the three universal probes differed statistically significantly between the BK group and the non-BK group. Based on the ROC curves, the sensitivities of PB1, PB2, and PB3 were 81.3%, 81.3%, and 93.8%, and the specificities were 71.1%, 88.9%, and 91.1%, respectively. The sensitivity and specificity of the Psu probe were 92% and 100%, respectively, while those of the Aci and Klb probes could not be estimated because there were no BK cases caused by Acinetobacter spp. or Klebsiella spp. CONCLUSIONS The BDH assay is an effective molecular approach to improve the diagnosis of BK. Because the bias from bacterial contamination on the ocular surface can be minimized with signal standardization, the assay has the potential to be adopted for routine clinical practice.

  • a multiplex Dot Hybridization assay for detection and differentiation of acanthamoeba and herpes keratitis
    Investigative Ophthalmology & Visual Science, 2016
    Co-Authors: Mingtse Kuo, Chunchih Chien, Shinling Tseng, Changchun Hsiao, Pochiung Fang, Yuhsuan Lai, Tsae Ling Chao, Shun Hua Chen, Jen Ren Wang, Tsung C Chang
    Abstract:

    PURPOSE We verified a multiplex Dot Hybridization (MDH) assay for the rapid detection and differentiation of Acanthamoeba keratitis (AK) and herpes simplex keratitis (HSK). METHODS Molecular detection of Acanthamoeba and herpes simplex virus in corneal scrapes was performed with the MDH assay and standard diagnostic methods. The experimental group included corneal scrapes (n = 33) from patients with culture- or pathology-confirmed AK (n = 15) and real-time PCR-confirmed HSK (n = 16). The control group included 50 samples from cases of bacterial keratitis (n = 15), fungal keratitis (n = 15), and initially presumed AK (n = 5) or HSK (n = 17) which finally were excluded by culture for Acanthamoeba or by real-time PCR for herpes simplex virus, respectively. Discrepant results between methods were resolved by DNA sequencing of the PCR amplicons. RESULTS After discrepant analysis, the sensitivity for the diagnosis of AK and HSK was both 93.3% by the MDH assay, while the specificity was 100% for the two types of keratitis. The turnaround time of MDH assay was within a working day using an already prepared array. Two false-negatives (one AK case and one HSK case) were obtained by the MDH assay. CONCLUSIONS The MDH assay could effectively prevent missed or delayed diagnosis of AK and HSK and has a potential to be adopted in routine clinical practice if the test is commercialized.

  • a dna Dot Hybridization model for assessment of bacterial bioburden in orthokeratology lens storage cases
    Investigative Ophthalmology & Visual Science, 2015
    Co-Authors: Chunchih Chien, Jung Lo, Shinling Tseng, Changchun Hsiao, Pochiung Fang, Tsung C Chang
    Abstract:

    PURPOSE: The aim of this study was to evaluate a DNA Dot Hybridization assay (DHA) for assessing bacterial bioburden in orthokeratology lens (OK) storage cases. METHODS: Forty-one OK wearers participated in this study. The Dot Hybridization assay was used to assess the bacterial bioburden of OK after removal and 6-hour soaking in a storage case. Signals of the DHA were standardized after gray image transformation. The correlations between the Hybridization intensities of three universal bacteria probes (BP1, BP2, and BP3) and bacterial bioburden determined by culture (colony forming units per milliliter) was analyzed by Pearson's correlation coefficient and receiver operating characteristic plots. In addition, three genus-specific probes for Pseudomonas, Acinetobacter, and Klebsiella were used to detect potentially hazardous bacterial contamination regardless of bacterial viability status. RESULTS: Among the three universal probes, there were good correlations between probe BP2 (r2 = 0.31, P = 9.5 × 10(-5)) and probe BP3 (r2 = 0.35, P = 3.1 × 10(-5)) with bacterial bioburden, but no correlation was found between probe BP1 and bacterial bioburden (r2 = 0.04, P = 0.11). In 41 samples, one was Pseudomonas-positive by both DHA and culture, while 10 were Pseudomonas-positive by DHA but negative by culture. Furthermore, nine samples tested positive for Acinetobacter (n = 7) and Klebsiella (n = 2) by DHA only. CONCLUSIONS: The Dot Hybridization assay provides a novel way to assess the bacterial bioburden of OK storage cases. Lens care quality can be assessed with universal bacteria probes, while potentially hazardous bacterial contamination can be traced with genus-specific probes.

Chunchih Chien - One of the best experts on this subject based on the ideXlab platform.

  • bacterial bioburden decrease in orthokeratology lens storage cases after forewarning assessment by the dna Dot Hybridization assay
    Eye & Contact Lens-science and Clinical Practice, 2017
    Co-Authors: Pochiung Fang, Jung Lo, Tsung C Chang, Chunchih Chien, Changchun Hsiao, Shinling Tseng
    Abstract:

    BACKGROUND: The aim of this study was to measure the changes in the bacterial bioburden in orthokeratology (OK) lens storage cases using the DNA Dot Hybridization assay (DHA) after forewarning patients about their bacterial contamination severity. METHODS: Thirty-one OK lens wearers were prospectively enrolled in this study. Dot Hybridization assay was used for serial measurements of bacterial bioburden in OK storage cases after lenses had been soaked for approximately 6 hr. After the first assessment, the lens wearers were informed of the extent of case contamination and the possible risk of microbial keratitis (MK), and best practices for lens care and lens case hygiene were reviewed and reinforced. A second assessment by the same DHA method was performed after approximately 6 months. RESULTS: Two universal bacterial probes confirmed a significant decrease in bacterial bioburden at the second assessment (P<0.01 and P<0.001). Genus-specific probes showed significant reductions in Acinetobacter and Klebsiella (P=0.02 and P=0.01), but not in Pseudomonas (P=0.42). CONCLUSIONS: Making OK lens wearers aware of the bacterial bioburden in their lens cases resulted in improved quality of case care and reduced bioburden. Our results suggest that a strategy of bioburden assessment with forewarning could be a useful method to decrease the incidence of OK-related MK.

  • a Dot Hybridization assay for the diagnosis of bacterial keratitis
    Molecular Vision, 2017
    Co-Authors: Pochiung Fang, Chunchih Chien, Shinling Tseng, Yuhsuan Lai, Mingtse Kuo
    Abstract:

    PURPOSE To evaluate a bacterial Dot Hybridization (BDH) assay for the diagnosis of bacterial keratitis (BK). METHODS Sixty-one qualified corneal scrapings from 61 patients with suspected microbial keratitis were collected consecutively and prospectively. Among the 61 patients, 16 cases were BK and 45 cases were non-BK, including fungal keratitis, viral keratitis, parasitic keratitis, and non-microbial keratitis. Molecular diagnosis of BK in these corneal scrapes was performed using the BDH assay with three universal bacterial probes (PB1, PB2, and PB3) and three genus-specific probes (Aci, Klb, and Psu) to detect Acinetobacter, Klebsiella, and Pseudomonas, respectively. Signals were standardized after grayscale image transformation for objective validation using receiver operating characteristic (ROC) curves. RESULTS The standardized intensities for the three universal probes differed statistically significantly between the BK group and the non-BK group. Based on the ROC curves, the sensitivities of PB1, PB2, and PB3 were 81.3%, 81.3%, and 93.8%, and the specificities were 71.1%, 88.9%, and 91.1%, respectively. The sensitivity and specificity of the Psu probe were 92% and 100%, respectively, while those of the Aci and Klb probes could not be estimated because there were no BK cases caused by Acinetobacter spp. or Klebsiella spp. CONCLUSIONS The BDH assay is an effective molecular approach to improve the diagnosis of BK. Because the bias from bacterial contamination on the ocular surface can be minimized with signal standardization, the assay has the potential to be adopted for routine clinical practice.

  • a multiplex Dot Hybridization assay for detection and differentiation of acanthamoeba and herpes keratitis
    Investigative Ophthalmology & Visual Science, 2016
    Co-Authors: Mingtse Kuo, Chunchih Chien, Shinling Tseng, Changchun Hsiao, Pochiung Fang, Yuhsuan Lai, Tsae Ling Chao, Shun Hua Chen, Jen Ren Wang, Tsung C Chang
    Abstract:

    PURPOSE We verified a multiplex Dot Hybridization (MDH) assay for the rapid detection and differentiation of Acanthamoeba keratitis (AK) and herpes simplex keratitis (HSK). METHODS Molecular detection of Acanthamoeba and herpes simplex virus in corneal scrapes was performed with the MDH assay and standard diagnostic methods. The experimental group included corneal scrapes (n = 33) from patients with culture- or pathology-confirmed AK (n = 15) and real-time PCR-confirmed HSK (n = 16). The control group included 50 samples from cases of bacterial keratitis (n = 15), fungal keratitis (n = 15), and initially presumed AK (n = 5) or HSK (n = 17) which finally were excluded by culture for Acanthamoeba or by real-time PCR for herpes simplex virus, respectively. Discrepant results between methods were resolved by DNA sequencing of the PCR amplicons. RESULTS After discrepant analysis, the sensitivity for the diagnosis of AK and HSK was both 93.3% by the MDH assay, while the specificity was 100% for the two types of keratitis. The turnaround time of MDH assay was within a working day using an already prepared array. Two false-negatives (one AK case and one HSK case) were obtained by the MDH assay. CONCLUSIONS The MDH assay could effectively prevent missed or delayed diagnosis of AK and HSK and has a potential to be adopted in routine clinical practice if the test is commercialized.

  • a dna Dot Hybridization model for assessment of bacterial bioburden in orthokeratology lens storage cases
    Investigative Ophthalmology & Visual Science, 2015
    Co-Authors: Chunchih Chien, Jung Lo, Shinling Tseng, Changchun Hsiao, Pochiung Fang, Tsung C Chang
    Abstract:

    PURPOSE: The aim of this study was to evaluate a DNA Dot Hybridization assay (DHA) for assessing bacterial bioburden in orthokeratology lens (OK) storage cases. METHODS: Forty-one OK wearers participated in this study. The Dot Hybridization assay was used to assess the bacterial bioburden of OK after removal and 6-hour soaking in a storage case. Signals of the DHA were standardized after gray image transformation. The correlations between the Hybridization intensities of three universal bacteria probes (BP1, BP2, and BP3) and bacterial bioburden determined by culture (colony forming units per milliliter) was analyzed by Pearson's correlation coefficient and receiver operating characteristic plots. In addition, three genus-specific probes for Pseudomonas, Acinetobacter, and Klebsiella were used to detect potentially hazardous bacterial contamination regardless of bacterial viability status. RESULTS: Among the three universal probes, there were good correlations between probe BP2 (r2 = 0.31, P = 9.5 × 10(-5)) and probe BP3 (r2 = 0.35, P = 3.1 × 10(-5)) with bacterial bioburden, but no correlation was found between probe BP1 and bacterial bioburden (r2 = 0.04, P = 0.11). In 41 samples, one was Pseudomonas-positive by both DHA and culture, while 10 were Pseudomonas-positive by DHA but negative by culture. Furthermore, nine samples tested positive for Acinetobacter (n = 7) and Klebsiella (n = 2) by DHA only. CONCLUSIONS: The Dot Hybridization assay provides a novel way to assess the bacterial bioburden of OK storage cases. Lens care quality can be assessed with universal bacteria probes, while potentially hazardous bacterial contamination can be traced with genus-specific probes.

  • a highly sensitive method for molecular diagnosis of fungal keratitis a Dot Hybridization assay
    Ophthalmology, 2012
    Co-Authors: Mingtse Kuo, Chunchih Chien, Pochiung Fang, Hsienchang Chang, Chao Kun Cheng, Tsung C Chang
    Abstract:

    Purpose Fungal keratitis (FK) is an important cause of ocular morbidity, especially for people living in the agricultural communities of the developing world. Current diagnostic methods may lack sensitivity (direct microscopy) or are time consuming (culture). The aim of this study was to develop a Dot Hybridization assay for sensitive and rapid diagnosis of FK. Design Evaluation of diagnostic test or technology. Participants and Controls Fifty corneal scrapes (49 patients) from consecutive cases of clinically suspected microbial keratitis were analyzed prospectively. Methods Molecular detection of fungi in the scrapes was performed by amplification of the internal transcribed spacer region (ITS) that contained the target gene (5.8S rRNA gene) by polymerase chain reaction (PCR), followed by Hybridization of the PCR product to a fungus-specific oligonucleotide probe immobilized on a nylon membrane. The results were compared with those obtained by gram-stain microscopy, culture, and gel electrophoresis of the PCR products. Discrepant results were resolved by cloning and resequencing of the amplified ITS fragments. Main Outcome Measures Performance of the Dot Hybridization assay, including sensitivity, specificity, and positive and negative predictive values, was evaluated. Results Ten scrapes demonstrated positive results by both the Dot Hybridization assay and culture. However, 11 scrapes demonstrated positive results by the Dot Hybridization assay, but demonstrated negative results by culture, and 10 of the 11 samples were considered to be positive for FK by cloning and resequencing of the amplified ITS fragment and by a pathologic examination or clinical course review. The sensitivities for diagnosis of FK by the Dot Hybridization assay and culture were 100% and 50%, respectively, whereas the specificities were 96.7% and 100%, respectively. Conclusions The Dot Hybridization assay is a highly sensitive and specific diagnostic tool for FK. The method provides a much higher sensitivity than that of culture (100% vs. 50%; P Financial Disclosure(s) The author(s) have no proprietary or commercial interest in any materials discussed in this article.

Shinling Tseng - One of the best experts on this subject based on the ideXlab platform.

  • bacterial bioburden decrease in orthokeratology lens storage cases after forewarning assessment by the dna Dot Hybridization assay
    Eye & Contact Lens-science and Clinical Practice, 2017
    Co-Authors: Pochiung Fang, Jung Lo, Tsung C Chang, Chunchih Chien, Changchun Hsiao, Shinling Tseng
    Abstract:

    BACKGROUND: The aim of this study was to measure the changes in the bacterial bioburden in orthokeratology (OK) lens storage cases using the DNA Dot Hybridization assay (DHA) after forewarning patients about their bacterial contamination severity. METHODS: Thirty-one OK lens wearers were prospectively enrolled in this study. Dot Hybridization assay was used for serial measurements of bacterial bioburden in OK storage cases after lenses had been soaked for approximately 6 hr. After the first assessment, the lens wearers were informed of the extent of case contamination and the possible risk of microbial keratitis (MK), and best practices for lens care and lens case hygiene were reviewed and reinforced. A second assessment by the same DHA method was performed after approximately 6 months. RESULTS: Two universal bacterial probes confirmed a significant decrease in bacterial bioburden at the second assessment (P<0.01 and P<0.001). Genus-specific probes showed significant reductions in Acinetobacter and Klebsiella (P=0.02 and P=0.01), but not in Pseudomonas (P=0.42). CONCLUSIONS: Making OK lens wearers aware of the bacterial bioburden in their lens cases resulted in improved quality of case care and reduced bioburden. Our results suggest that a strategy of bioburden assessment with forewarning could be a useful method to decrease the incidence of OK-related MK.

  • a Dot Hybridization assay for the diagnosis of bacterial keratitis
    Molecular Vision, 2017
    Co-Authors: Pochiung Fang, Chunchih Chien, Shinling Tseng, Yuhsuan Lai, Mingtse Kuo
    Abstract:

    PURPOSE To evaluate a bacterial Dot Hybridization (BDH) assay for the diagnosis of bacterial keratitis (BK). METHODS Sixty-one qualified corneal scrapings from 61 patients with suspected microbial keratitis were collected consecutively and prospectively. Among the 61 patients, 16 cases were BK and 45 cases were non-BK, including fungal keratitis, viral keratitis, parasitic keratitis, and non-microbial keratitis. Molecular diagnosis of BK in these corneal scrapes was performed using the BDH assay with three universal bacterial probes (PB1, PB2, and PB3) and three genus-specific probes (Aci, Klb, and Psu) to detect Acinetobacter, Klebsiella, and Pseudomonas, respectively. Signals were standardized after grayscale image transformation for objective validation using receiver operating characteristic (ROC) curves. RESULTS The standardized intensities for the three universal probes differed statistically significantly between the BK group and the non-BK group. Based on the ROC curves, the sensitivities of PB1, PB2, and PB3 were 81.3%, 81.3%, and 93.8%, and the specificities were 71.1%, 88.9%, and 91.1%, respectively. The sensitivity and specificity of the Psu probe were 92% and 100%, respectively, while those of the Aci and Klb probes could not be estimated because there were no BK cases caused by Acinetobacter spp. or Klebsiella spp. CONCLUSIONS The BDH assay is an effective molecular approach to improve the diagnosis of BK. Because the bias from bacterial contamination on the ocular surface can be minimized with signal standardization, the assay has the potential to be adopted for routine clinical practice.

  • a multiplex Dot Hybridization assay for detection and differentiation of acanthamoeba and herpes keratitis
    Investigative Ophthalmology & Visual Science, 2016
    Co-Authors: Mingtse Kuo, Chunchih Chien, Shinling Tseng, Changchun Hsiao, Pochiung Fang, Yuhsuan Lai, Tsae Ling Chao, Shun Hua Chen, Jen Ren Wang, Tsung C Chang
    Abstract:

    PURPOSE We verified a multiplex Dot Hybridization (MDH) assay for the rapid detection and differentiation of Acanthamoeba keratitis (AK) and herpes simplex keratitis (HSK). METHODS Molecular detection of Acanthamoeba and herpes simplex virus in corneal scrapes was performed with the MDH assay and standard diagnostic methods. The experimental group included corneal scrapes (n = 33) from patients with culture- or pathology-confirmed AK (n = 15) and real-time PCR-confirmed HSK (n = 16). The control group included 50 samples from cases of bacterial keratitis (n = 15), fungal keratitis (n = 15), and initially presumed AK (n = 5) or HSK (n = 17) which finally were excluded by culture for Acanthamoeba or by real-time PCR for herpes simplex virus, respectively. Discrepant results between methods were resolved by DNA sequencing of the PCR amplicons. RESULTS After discrepant analysis, the sensitivity for the diagnosis of AK and HSK was both 93.3% by the MDH assay, while the specificity was 100% for the two types of keratitis. The turnaround time of MDH assay was within a working day using an already prepared array. Two false-negatives (one AK case and one HSK case) were obtained by the MDH assay. CONCLUSIONS The MDH assay could effectively prevent missed or delayed diagnosis of AK and HSK and has a potential to be adopted in routine clinical practice if the test is commercialized.

  • a dna Dot Hybridization model for assessment of bacterial bioburden in orthokeratology lens storage cases
    Investigative Ophthalmology & Visual Science, 2015
    Co-Authors: Chunchih Chien, Jung Lo, Shinling Tseng, Changchun Hsiao, Pochiung Fang, Tsung C Chang
    Abstract:

    PURPOSE: The aim of this study was to evaluate a DNA Dot Hybridization assay (DHA) for assessing bacterial bioburden in orthokeratology lens (OK) storage cases. METHODS: Forty-one OK wearers participated in this study. The Dot Hybridization assay was used to assess the bacterial bioburden of OK after removal and 6-hour soaking in a storage case. Signals of the DHA were standardized after gray image transformation. The correlations between the Hybridization intensities of three universal bacteria probes (BP1, BP2, and BP3) and bacterial bioburden determined by culture (colony forming units per milliliter) was analyzed by Pearson's correlation coefficient and receiver operating characteristic plots. In addition, three genus-specific probes for Pseudomonas, Acinetobacter, and Klebsiella were used to detect potentially hazardous bacterial contamination regardless of bacterial viability status. RESULTS: Among the three universal probes, there were good correlations between probe BP2 (r2 = 0.31, P = 9.5 × 10(-5)) and probe BP3 (r2 = 0.35, P = 3.1 × 10(-5)) with bacterial bioburden, but no correlation was found between probe BP1 and bacterial bioburden (r2 = 0.04, P = 0.11). In 41 samples, one was Pseudomonas-positive by both DHA and culture, while 10 were Pseudomonas-positive by DHA but negative by culture. Furthermore, nine samples tested positive for Acinetobacter (n = 7) and Klebsiella (n = 2) by DHA only. CONCLUSIONS: The Dot Hybridization assay provides a novel way to assess the bacterial bioburden of OK storage cases. Lens care quality can be assessed with universal bacteria probes, while potentially hazardous bacterial contamination can be traced with genus-specific probes.

Changchun Hsiao - One of the best experts on this subject based on the ideXlab platform.

  • bacterial bioburden decrease in orthokeratology lens storage cases after forewarning assessment by the dna Dot Hybridization assay
    Eye & Contact Lens-science and Clinical Practice, 2017
    Co-Authors: Pochiung Fang, Jung Lo, Tsung C Chang, Chunchih Chien, Changchun Hsiao, Shinling Tseng
    Abstract:

    BACKGROUND: The aim of this study was to measure the changes in the bacterial bioburden in orthokeratology (OK) lens storage cases using the DNA Dot Hybridization assay (DHA) after forewarning patients about their bacterial contamination severity. METHODS: Thirty-one OK lens wearers were prospectively enrolled in this study. Dot Hybridization assay was used for serial measurements of bacterial bioburden in OK storage cases after lenses had been soaked for approximately 6 hr. After the first assessment, the lens wearers were informed of the extent of case contamination and the possible risk of microbial keratitis (MK), and best practices for lens care and lens case hygiene were reviewed and reinforced. A second assessment by the same DHA method was performed after approximately 6 months. RESULTS: Two universal bacterial probes confirmed a significant decrease in bacterial bioburden at the second assessment (P<0.01 and P<0.001). Genus-specific probes showed significant reductions in Acinetobacter and Klebsiella (P=0.02 and P=0.01), but not in Pseudomonas (P=0.42). CONCLUSIONS: Making OK lens wearers aware of the bacterial bioburden in their lens cases resulted in improved quality of case care and reduced bioburden. Our results suggest that a strategy of bioburden assessment with forewarning could be a useful method to decrease the incidence of OK-related MK.

  • a multiplex Dot Hybridization assay for detection and differentiation of acanthamoeba and herpes keratitis
    Investigative Ophthalmology & Visual Science, 2016
    Co-Authors: Mingtse Kuo, Chunchih Chien, Shinling Tseng, Changchun Hsiao, Pochiung Fang, Yuhsuan Lai, Tsae Ling Chao, Shun Hua Chen, Jen Ren Wang, Tsung C Chang
    Abstract:

    PURPOSE We verified a multiplex Dot Hybridization (MDH) assay for the rapid detection and differentiation of Acanthamoeba keratitis (AK) and herpes simplex keratitis (HSK). METHODS Molecular detection of Acanthamoeba and herpes simplex virus in corneal scrapes was performed with the MDH assay and standard diagnostic methods. The experimental group included corneal scrapes (n = 33) from patients with culture- or pathology-confirmed AK (n = 15) and real-time PCR-confirmed HSK (n = 16). The control group included 50 samples from cases of bacterial keratitis (n = 15), fungal keratitis (n = 15), and initially presumed AK (n = 5) or HSK (n = 17) which finally were excluded by culture for Acanthamoeba or by real-time PCR for herpes simplex virus, respectively. Discrepant results between methods were resolved by DNA sequencing of the PCR amplicons. RESULTS After discrepant analysis, the sensitivity for the diagnosis of AK and HSK was both 93.3% by the MDH assay, while the specificity was 100% for the two types of keratitis. The turnaround time of MDH assay was within a working day using an already prepared array. Two false-negatives (one AK case and one HSK case) were obtained by the MDH assay. CONCLUSIONS The MDH assay could effectively prevent missed or delayed diagnosis of AK and HSK and has a potential to be adopted in routine clinical practice if the test is commercialized.

  • a dna Dot Hybridization model for assessment of bacterial bioburden in orthokeratology lens storage cases
    Investigative Ophthalmology & Visual Science, 2015
    Co-Authors: Chunchih Chien, Jung Lo, Shinling Tseng, Changchun Hsiao, Pochiung Fang, Tsung C Chang
    Abstract:

    PURPOSE: The aim of this study was to evaluate a DNA Dot Hybridization assay (DHA) for assessing bacterial bioburden in orthokeratology lens (OK) storage cases. METHODS: Forty-one OK wearers participated in this study. The Dot Hybridization assay was used to assess the bacterial bioburden of OK after removal and 6-hour soaking in a storage case. Signals of the DHA were standardized after gray image transformation. The correlations between the Hybridization intensities of three universal bacteria probes (BP1, BP2, and BP3) and bacterial bioburden determined by culture (colony forming units per milliliter) was analyzed by Pearson's correlation coefficient and receiver operating characteristic plots. In addition, three genus-specific probes for Pseudomonas, Acinetobacter, and Klebsiella were used to detect potentially hazardous bacterial contamination regardless of bacterial viability status. RESULTS: Among the three universal probes, there were good correlations between probe BP2 (r2 = 0.31, P = 9.5 × 10(-5)) and probe BP3 (r2 = 0.35, P = 3.1 × 10(-5)) with bacterial bioburden, but no correlation was found between probe BP1 and bacterial bioburden (r2 = 0.04, P = 0.11). In 41 samples, one was Pseudomonas-positive by both DHA and culture, while 10 were Pseudomonas-positive by DHA but negative by culture. Furthermore, nine samples tested positive for Acinetobacter (n = 7) and Klebsiella (n = 2) by DHA only. CONCLUSIONS: The Dot Hybridization assay provides a novel way to assess the bacterial bioburden of OK storage cases. Lens care quality can be assessed with universal bacteria probes, while potentially hazardous bacterial contamination can be traced with genus-specific probes.