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Specificity and Strain-Typing Capabilities of Nanorod Array-Surface Enhanced Raman Spectroscopy for Mycoplasma pneumoniae Detection.

Henderson KC, Benitez AJ, Ratliff AE, Crabb DM, Sheppard ES, Winchell JM, Dluhy RA, Waites KB, Atkinson TP, Krause DC - PLoS ONE (2015)

Bottom Line: At present the most effective means for detection and strain-typing is quantitative polymerase chain reaction (qPCR), which can exhibit excellent sensitivity and specificity but requires separate tests for detection and genotyping, lacks standardization between available tests and between labs, and has limited practicality for widespread, point-of-care use.Here we demonstrate using partial least squares- discriminatory analysis (PLS-DA) of sample spectra that NA-SERS correctly identified M. pneumoniae clinical isolates from globally diverse origins and distinguished these from a panel of 12 other human commensal and pathogenic mycoplasma species with 100% cross-validated statistical accuracy.Furthermore, PLS-DA correctly classified by strain type all 30 clinical isolates with 96% cross-validated accuracy for type 1 strains, 98% cross-validated accuracy for type 2 strains, and 90% cross-validated accuracy for type 2V strains.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, University of Georgia, Athens, GA, United States of America.

ABSTRACT
Mycoplasma pneumoniae is a cell wall-less bacterial pathogen of the human respiratory tract that accounts for > 20% of all community-acquired pneumonia (CAP). At present the most effective means for detection and strain-typing is quantitative polymerase chain reaction (qPCR), which can exhibit excellent sensitivity and specificity but requires separate tests for detection and genotyping, lacks standardization between available tests and between labs, and has limited practicality for widespread, point-of-care use. We have developed and previously described a silver nanorod array-surface enhanced Raman Spectroscopy (NA-SERS) biosensing platform capable of detecting M. pneumoniae with statistically significant specificity and sensitivity in simulated and true clinical throat swab samples, and the ability to distinguish between reference strains of the two main genotypes of M. pneumoniae. Furthermore, we have established a qualitative lower endpoint of detection for NA-SERS of < 1 genome equivalent (cell/μl) and a quantitative multivariate detection limit of 5.3 ± 1 cells/μl. Here we demonstrate using partial least squares- discriminatory analysis (PLS-DA) of sample spectra that NA-SERS correctly identified M. pneumoniae clinical isolates from globally diverse origins and distinguished these from a panel of 12 other human commensal and pathogenic mycoplasma species with 100% cross-validated statistical accuracy. Furthermore, PLS-DA correctly classified by strain type all 30 clinical isolates with 96% cross-validated accuracy for type 1 strains, 98% cross-validated accuracy for type 2 strains, and 90% cross-validated accuracy for type 2V strains.

No MeSH data available.


Related in: MedlinePlus

Principle component analysis of M. pneumoniae strain typing and other human commensal and pathogenic Mollicutes species.For all panels, each individual shape represents a single sample spectrum. PC scores plots of 1 vs. 2 vs. 3 of: (A)M. pneumoniae reference strains and all 13 other type 1 clinical isolates; (B)M. pneumoniae reference strains and all 11 other type 2 clinical isolates; (C)M. pneumoniae type 1 reference strain, type 2 reference strain, and all six type 2V clinical isolates; and (D) growth medium control, all M. pneumoniae strains, and all 12 other human commensal and pathogenic Mollicutes species. For panels A-C, dark gray diamonds represent type 1 sample spectra whereas open squares represent the type 2 sample spectra. In panel C, type 2V clinical isolate spectra are represented by light gray triangles. For panel D, growth medium control spectra are represented by dark gray diamonds, M. pneumoniae spectra by open squares, and all 12 other Mollicutes species by light gray triangles. For panels A-D, clustering of samples is indicated by black circles or brackets.
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pone.0131831.g007: Principle component analysis of M. pneumoniae strain typing and other human commensal and pathogenic Mollicutes species.For all panels, each individual shape represents a single sample spectrum. PC scores plots of 1 vs. 2 vs. 3 of: (A)M. pneumoniae reference strains and all 13 other type 1 clinical isolates; (B)M. pneumoniae reference strains and all 11 other type 2 clinical isolates; (C)M. pneumoniae type 1 reference strain, type 2 reference strain, and all six type 2V clinical isolates; and (D) growth medium control, all M. pneumoniae strains, and all 12 other human commensal and pathogenic Mollicutes species. For panels A-C, dark gray diamonds represent type 1 sample spectra whereas open squares represent the type 2 sample spectra. In panel C, type 2V clinical isolate spectra are represented by light gray triangles. For panel D, growth medium control spectra are represented by dark gray diamonds, M. pneumoniae spectra by open squares, and all 12 other Mollicutes species by light gray triangles. For panels A-D, clustering of samples is indicated by black circles or brackets.

Mentions: Pre-processed SERS spectra from M. pneumoniae reference strain type 1 and 2 controls and all other type 1 clinical isolates were used to generate a PCA plot comparing principle components 1, 2, and 3, which captured 54.3% of the total variance present in the 180 spectra used to build the model (Fig 7A). Type 2 control strain FH clustered in the bottom right corner, and the clustering pattern for all type 1 strains was predominately below and to the left, though some overlap between the two strain types was present. The PCA model of the type 1 clinical isolates supports the PLS-DA modeling of the spectra shown in Fig 4A.


Specificity and Strain-Typing Capabilities of Nanorod Array-Surface Enhanced Raman Spectroscopy for Mycoplasma pneumoniae Detection.

Henderson KC, Benitez AJ, Ratliff AE, Crabb DM, Sheppard ES, Winchell JM, Dluhy RA, Waites KB, Atkinson TP, Krause DC - PLoS ONE (2015)

Principle component analysis of M. pneumoniae strain typing and other human commensal and pathogenic Mollicutes species.For all panels, each individual shape represents a single sample spectrum. PC scores plots of 1 vs. 2 vs. 3 of: (A)M. pneumoniae reference strains and all 13 other type 1 clinical isolates; (B)M. pneumoniae reference strains and all 11 other type 2 clinical isolates; (C)M. pneumoniae type 1 reference strain, type 2 reference strain, and all six type 2V clinical isolates; and (D) growth medium control, all M. pneumoniae strains, and all 12 other human commensal and pathogenic Mollicutes species. For panels A-C, dark gray diamonds represent type 1 sample spectra whereas open squares represent the type 2 sample spectra. In panel C, type 2V clinical isolate spectra are represented by light gray triangles. For panel D, growth medium control spectra are represented by dark gray diamonds, M. pneumoniae spectra by open squares, and all 12 other Mollicutes species by light gray triangles. For panels A-D, clustering of samples is indicated by black circles or brackets.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4487258&req=5

pone.0131831.g007: Principle component analysis of M. pneumoniae strain typing and other human commensal and pathogenic Mollicutes species.For all panels, each individual shape represents a single sample spectrum. PC scores plots of 1 vs. 2 vs. 3 of: (A)M. pneumoniae reference strains and all 13 other type 1 clinical isolates; (B)M. pneumoniae reference strains and all 11 other type 2 clinical isolates; (C)M. pneumoniae type 1 reference strain, type 2 reference strain, and all six type 2V clinical isolates; and (D) growth medium control, all M. pneumoniae strains, and all 12 other human commensal and pathogenic Mollicutes species. For panels A-C, dark gray diamonds represent type 1 sample spectra whereas open squares represent the type 2 sample spectra. In panel C, type 2V clinical isolate spectra are represented by light gray triangles. For panel D, growth medium control spectra are represented by dark gray diamonds, M. pneumoniae spectra by open squares, and all 12 other Mollicutes species by light gray triangles. For panels A-D, clustering of samples is indicated by black circles or brackets.
Mentions: Pre-processed SERS spectra from M. pneumoniae reference strain type 1 and 2 controls and all other type 1 clinical isolates were used to generate a PCA plot comparing principle components 1, 2, and 3, which captured 54.3% of the total variance present in the 180 spectra used to build the model (Fig 7A). Type 2 control strain FH clustered in the bottom right corner, and the clustering pattern for all type 1 strains was predominately below and to the left, though some overlap between the two strain types was present. The PCA model of the type 1 clinical isolates supports the PLS-DA modeling of the spectra shown in Fig 4A.

Bottom Line: At present the most effective means for detection and strain-typing is quantitative polymerase chain reaction (qPCR), which can exhibit excellent sensitivity and specificity but requires separate tests for detection and genotyping, lacks standardization between available tests and between labs, and has limited practicality for widespread, point-of-care use.Here we demonstrate using partial least squares- discriminatory analysis (PLS-DA) of sample spectra that NA-SERS correctly identified M. pneumoniae clinical isolates from globally diverse origins and distinguished these from a panel of 12 other human commensal and pathogenic mycoplasma species with 100% cross-validated statistical accuracy.Furthermore, PLS-DA correctly classified by strain type all 30 clinical isolates with 96% cross-validated accuracy for type 1 strains, 98% cross-validated accuracy for type 2 strains, and 90% cross-validated accuracy for type 2V strains.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, University of Georgia, Athens, GA, United States of America.

ABSTRACT
Mycoplasma pneumoniae is a cell wall-less bacterial pathogen of the human respiratory tract that accounts for > 20% of all community-acquired pneumonia (CAP). At present the most effective means for detection and strain-typing is quantitative polymerase chain reaction (qPCR), which can exhibit excellent sensitivity and specificity but requires separate tests for detection and genotyping, lacks standardization between available tests and between labs, and has limited practicality for widespread, point-of-care use. We have developed and previously described a silver nanorod array-surface enhanced Raman Spectroscopy (NA-SERS) biosensing platform capable of detecting M. pneumoniae with statistically significant specificity and sensitivity in simulated and true clinical throat swab samples, and the ability to distinguish between reference strains of the two main genotypes of M. pneumoniae. Furthermore, we have established a qualitative lower endpoint of detection for NA-SERS of < 1 genome equivalent (cell/μl) and a quantitative multivariate detection limit of 5.3 ± 1 cells/μl. Here we demonstrate using partial least squares- discriminatory analysis (PLS-DA) of sample spectra that NA-SERS correctly identified M. pneumoniae clinical isolates from globally diverse origins and distinguished these from a panel of 12 other human commensal and pathogenic mycoplasma species with 100% cross-validated statistical accuracy. Furthermore, PLS-DA correctly classified by strain type all 30 clinical isolates with 96% cross-validated accuracy for type 1 strains, 98% cross-validated accuracy for type 2 strains, and 90% cross-validated accuracy for type 2V strains.

No MeSH data available.


Related in: MedlinePlus