XIV
h
International Conference on Molecular Spectroscopy, Białka Tatrzańska 2017
74
T2: O–6
Rapid detection and identification of bacterial meningitis pathogens
in ex vivo clinical samples by SERS method and principal component
analysis
Agnieszka Kamińska
1
, Evelin Witkowska
1
, Aneta Kowalska
1
, Anna Skoczyńska
2
,
Patrycja Ronkiewicz
2
, Tomasz Szymborski
1
, and Jacek Waluk
1,3
1
Institute of Physical Chemistry, Polish Academy of Sciences, Kasprzaka 44/52, 01-224 Warsaw,
Poland, e-mail: akamin@ichf.edu.pl
2
National Medicines Institute, Chełmska 30/34, 00-725 Warsaw, Poland
3
Faculty of Mathematics and Natural Sciences, College of Science, Cardinal Stefan Wyszyński
University, Dewajtis 5, 01-815 Warsaw, Poland
The three of the most common meningitis pathogens, Neisseria meningitidis, Haemophilus
influenzae, and Streptococcus pneumoniae have been successfully detected and identified in
clinical cerebrospinal fluid (CSF) samples using a new class of a surface-enhanced Raman
scattering (SERS) assay. Bacterial meningitis is a disease of a nervous system which is
extremely serious and often fatal (an inflammation comprises the lining around the brain and
spinal cord). The approach presented in this study challenges the current SERS-based method of
microorganisms detection in terms of sensitivity and more importantly reveals a simple, quick
(on a time scale of seconds), label-free detection of multiple components from very small
volume of clinical samples. This new SERS class of assay based on combination of two types of
Au-Ag coated nuclepore track-etched polycarbonate membranes, which allows simultaneous
filtration of CSF, immobilization of CSF components, enhancing their Raman signals, and
finally enabling detection of spectra of single bacteria present in the analyzed CSF samples. The
multivariate statistical method, principal component analysis (PCA) was applied (i) to extract
the biochemical information from the recorded bacterial spectra; (ii) to perform the statistical
classification of analyzed microorganisms, and, finally (iii) to identify the spectrum of an
unknown sample by comparing it to library of spectra of known bacteria.
The three meningitis pathogens: N. meningitidis, H. influenzae, and S. pneumoniae were
detected and identified simultaneously using a label-free SERS method. This method of
detection produces consistent results faster and cheaper than traditional laboratory techniques
and demonstrates the powerful potential of SERS technique in medical applications.
Additionally, the present study was undertaken to evaluate the CSF neopterin level in patients
with diagnosed meningococcal meningitis. The results of this study confirmed that bacterial
meningitis caused by N. meningitidis, H. influenzae, and S. pneumoniae are associated with
elevated cerebrospinal fluid neopterin levels compared with control CSF samples. The value of
neopterin concentration can be used to predict meningitis, but cannot be applied to qualify the
species of bacteria inducing the meningitis infection.
Keywords: surface-enhanced Raman spectroscopy (SERS), neopterin, bacterial infections, Neisseria meningitidis,
Haemophilus influenzae, Streptococcus pneumoniae
Acknowledgment.
We would like to thank Dorota Korsak from University of Warsaw (Applied Microbiology, Institute of
Microbiology, Faculty of Biology) for preparation of sBHI agar medium and cultivation of H. influenza.
The authors would like to acknowledge the support from NCN under grant UMO-2015/17/B/ST4/04128.
XIV
h
International Conference on Molecular Spectroscopy, Białka Tatrzańska 2017
75
T2: O–7
Spectropathology of adrenal gland – is this possible?
Joanna Dudała
1
, and Magdalena Bialas
2
1
AGH University of Science and Technology, Faculty of Physics and Applied Computer Science,
Mickiewicza Av. 30, 30-059 Krakow, Poland, e-mail: Joanna.Dudala@fis.agh.edu.pl
2
Chair and Department of Pathomorphology, Jagiellonian University, Medical College,
Grzegorzecka st. 16, 31-531 Krakow, Poland
The adrenal glands are paired, small endocrine organs located in the retroperitoneum on the
kidney surface. They consist of cortex and medulla which differ in their development and
function. Nowadays, when the image diagnostic methods develop rapidly, and they are easy
accessible, the number of detected adrenal gland neoplastic lesions constantly increase. The
chirurgical intervention might be done due to oncological (when the imaging examinations
suggest the neoplastic lesion ) or endocrinological (when the hormonal activity of tumor was
confirmed) indications [1]. Although imaging and hormonal studies give a basic knowledge of
the tumor nature, the final diagnosis after adrenalectomy is based on the histopthological
examination. Classification of adrenal tumors due to their potential malice is based on
multivariate evaluation systems based on morphological, clinical and biochemical features, as
well as on the evaluation of the tumor proliferative activity using Ki-67. Nevertheless none of
the proposed methods for the differentiation between the benign and malignant tumors is not
ideal and fully objective. Therefore it is reasonable to undertake research aimed at finding the
complementary methods which could be helpful in differentiating the pathological alterations
within the adrenals. The literature reviews shows that it is possible to support diagnosis by
comparing the composition of biomolecular and elemental of healthy and pathologically altered
tissues. Nowadays the Fourier Transformed Infrared spectroscopy is regarded as a promising
complementary diagnostic method [2].
The presented studies are the continuation of research towards the possibility of using FTIR
spectroscopy for the biomolecular characterization of the adrenal gland. The preliminary studies
have been done for archival, fresh-frozen tissue samples. The results showed the clear difference
between the biomolecular pattern for the lesion derived from adrenal cortex and adrenal medulla
as well as between malignant and benign adrenal cortex lesion.
The next step of the investigations is the attempt to use formalin fixed and paraffin
embedded (FFPE) tissue samples which constitute a valuable source of clinical material for
retrospective studies [3]. It is particularly important in case of studies of the rare samples as it is
in case of malignant lesion of adrenal gland. The aim of presented study is the answer of two
questions. First – is it possible to use deparaffinised adrenal samples for FTIR measurements,
second – is it possible to differentiate the pathological alterations on the basis of obtained
results.
The studies were carried out at the Faculty of Physics and Applied Computer Science at the
AGH University of Science and Technology in Krakow in co-operation with the Department of
Pathomorfology, Collegium Medicum, Jagiellonian University in Krakow.
Keywords: FTIR spectroscopy; adrenal gland; adrenal cortical carcinoma; adrenal cortical adenoma; FFPE
samples
Acknowledgements
This work was supported by the Polish Ministry of Science and Higher Education and its grants for
Scientific Research.
References
[1] T. Bednarczuk, M. Bolanowski, K. Sworczyk et al., Endokrynologia Polska 67(2) (2016) 234.
[2] C. Petibois, G. Deleris, Trends in Biotechnology 24(10) (2006) 455.
[3] V. Zohdi, D.R. Whelan et al., PLoS ONE 10(2) (2015) pp 1–11.
Dostları ilə paylaş: |