Добавил:
Sekretar
kiopkiopkiop18@yandex.ru
t.me/Prokururor I Вовсе не секретарь, но почту проверяю
Опубликованный материал нарушает ваши авторские права? Сообщите нам.
Вуз:
Предмет:
Файл:Ординатура / Хирургия / Библиотека им академика М.И. Перельмана / Книга_5539_Библиотеки_им_академика_М_И_Перельмана.pdf
Recent Advances in Molecular Diagnostics and Treatment of Heart...
97
[Citation Time(s):1]
7. I. Kindermann, M. Kindermann, R. Kandolf, K. Klingel, B. Bültmann,
T. Müller, A. Lindinger and M. Böhm, “Predictors of Outcome in
Patients with Suspected Myocarditis,” Circulation, Vol. 118, No. 6,
2008,pp. 639-648. doi:10.1161/CIRCULATIONAHA.108.769489
[Citation Time(s):1]
8. A. Gulati, A. Jabbour, T. F. Ismail, K. Guha, J. Khwaja, S. Raza, K.
Morarji, T. D. Brown, N. A. Ismail, M. R. Dweck, E. Di Pietro, M.
Roughton, R. Wage, Y. Daryani, R. O’Hanlon, M. N. Sheppard, F.
Alpendurada, A. R. Lyon, S. A. Cook, M. R. Cowie, R. G. Assomull,
D. J. Pennell and S. K. Prasad, “Association of Fibrosis with
Mortality and Sudden Cardiac Death in Patients with Nonischemic
Dilated Cardiomyopathy,” JAMA, Vol. 309, No. 9, 2013, pp. 896-
908. doi:10.1001/jama.2013.1363 [Citation Time(s):1]
9. U. Kühl, D. Lassner, M. Pauschinger, U. M. Gross, B. Seeberg,
M. Noutsias, W. Poller and H.-P. Schultheiss, “Prevalence of
Erythrovirus Genotypes in the Myocardium of Patients with Dilated
Cardiomyopathy,” Journal of Medical Virology, Vol. 80, No. 8,
2008, pp. 1243- 1251. doi:10.1002/jmv.21187 [Citation Time(s):1]
10. U. Kühl, M. Rohde, D. Lassner, U. M. Gross, F. Escher and H. P.
Schultheiss, “miRNA as Activity Markers in Parvo B19 Associated
Heart Disease,” Herz, Vol. 37, No. 6, 2012, pp. 637-643. doi:10.10
07/s00059-012-3656-3 [Citation Time(s):4]
11. U. Kuhl, M. Pauschinger, M. Noutsias, et al., “High Prevalence of
Viral Genomes and Multiple Viral Infections in the Myocardium
of Adults with ‘Idiopathic’ Left Ventricular Dysfunction,”
Circulation, Vol. 111, 2005, pp. 887-893. doi:10.1161/01.
CIR.0000155616.07901.35 [Citation Time(s):1]
12. P. E. Pellett, D. V. Ablashi, P. F. Ambros, et al., “Chromosomally
Integrated Human Herpesvirus 6: Questions and Answers,” Reviews
in Medical Virology, Vol. 22, No. 3, 2012, pp. 144-155. doi:10.1002/
rmv.715 [Citation Time(s):1]
13. C. T. Bock, K. Klingel and R. Kandolf, “Human Parvovirus B19-
Associated Myocarditis,” The New England Journal of Medicine,
Vol. 362, No. 13, 2010, pp. 1248- 1249.
14. A. L. Caforio, F. Calabrese, A. Angelini, et al., “A Prospective Study
98
Advances in Molecular Diagnostics
of Biopsy-Proven Myocarditis: Prognostic Relevance of Clinical and
Aetiopathogenetic Features at Diagnosis,” European Heart Journal,
Vol. 28, No. 11, 2007, pp. 1326-1333. doi:10.1093/eurheartj/ehm076
15. U. Kühl, D. Lassner, J. von Schlippenbach, W. Poller and H. P.
Schultheiss “Interferon-Beta Improves Survival in EnterovirusAssociated Cardiomyopathy,” Journal of the American College of
Cardiology, Vol. 60, No. 14, 2012, pp. 1295-1296. doi:10.1016/j.
jacc.2012.06.026 [Citation Time(s):2]
16. A. Willitzki, R. Hiemann, V. Peters, U. Sack, P. Schierack, S.
Rödiger, U. Anderer, K. Conrad, D. P. Bogdanos, D. Reinhold and
D. Roggenbuck, “New Platform Technology for Comprehensive
Serological Diagnostics of Autoimmune Diseases,” Clinical and
Developmental Immunology, Vol. 2012, 2012, Article ID: 284740.
17. N. Deubner, D. Berliner, A. Schlipp, G. Gelbrich, A. L. Caforio, S.
B. Felix, M. Fu, H. Katus, C. E. Angermann, M. J. Lohse, G. Ertl, S.
Störk, R. Jahns, Etiology, Titre-Course and Survival-Study Group,
“Cardiac Beta1- Adrenoceptor Autoantibodies in Human Heart
Disease: Rationale and Design of the Etiology, Titre-Course, and
Survival (ETiCS) Study,” European Journal of Heart Failure, Vol.
12, No. 7, 2010, pp. 753-762. doi:10.1093/eurjhf/hfq072 [Citation
Time(s):1]
18. C. Skurk, F. Wittchen, L. Suckau, H. Witt, M. Noutsias, H.
Fechner, H.-P. Schultheiss and W. Poller, “Description of a Local
Cardiac Adiponectin System and Its Deregulation in Dilated
Cardiomyopathy,” European Heart Journal, Vol. 29, No. 9, 2008,
pp. 1168-1180. doi:10.1093/eurheartj/ehn136 [Citation Time(s):1]
19. K. L. Kellar, R. R. Kalwar, K. A. Dubois, D. Crouse, W. D.
Chan and B. E. Kane, “Multiplexed Fluorescent Bead-Based
Immunoassays for Quantitation of Human Cytokines in Serum
and Culture Supernatants,” Cytometry, Vol. 45, No. 1, 2001,
pp. 27-36. doi:10.1002/1097-0320(20010901)45:1<27::AIDCYTO1141>3.0.CO;2-I
20. F. Wittchen, L. Suckau, H. Witt,C. Skurk, D. Lassner, H. Fechner, I.
Sipo, U. Ungethüm, P. Ruiz, M. Pauschinger, C. Tschope, U. Rauch
U, Kühl, H.-P. Schultheiss and W. Poller, “Genomic Expression
Proling of Human Inammatory Cardiomyopathy (DCMi)
Recent Advances in Molecular Diagnostics and Treatment of Heart...
99
Suggests Novel Therapeutic Targets,” European Heart Journal,
Vol. 85, No. 3, 2007, pp. 257-271. doi:10.1007/s00109-006-0122-9
[Citation Time(s):1]
21. S. V. Naga Prasad, Z. H. Duan, M. K. Gupta, et al., “Unique microRNA
Prole in End-Stage Heart Failure Indicates Alterations in Specic
Cardiovascular Signaling Networks,” The Journal of Biological
Chemistry, Vol. 284, 2009, pp. 27487-27499. doi:10.1074/jbc.
M109.036541
22. D. Baek, J. Villen, C. Shin, F. D. Camargo, S. P. Gygi and D. P.
Bartel, “The Impact of microRNAs on Protein Output,” Nature,
Vol. 4355, No. 7209, 2008, pp. 64-71. doi:10.1038/nature07242
[Citation Time(s):1]
23. P. S. Mitchell, R. K. Parkin, et al., “Circulating microRNAs as
Stable Blood-Based Markers for Cancer Detection,” PNAS, Vol.
105, 2008, pp. 10513-10518. [Citation Time(s):1]
24. G. Wang, E. S. Chan, B. C. Kwan, P. K. Li, S. K. Yip, C. C. Szeto
and C. F. Ng, “Expression of miRNAs in the Urine of Patients with
Bladder Cancer,” Clinical Genitourinary Cancer, Vol. 10, No. 2,
2012, pp. 106-113. doi:10.1016/j.clgc.2012.01.001
25. P. Puerta-Gil, R. Garcia-Baquero, A. Y. Jia, S. Ocana, M. Alvarez-
Mugica, J. L. Alvarez-Ossorio, C. Cordon-Cardo, F. Cava and M.
Sanchez-Carbayo, “miR-143, miR-222, and miR-452 Are Useful as
Tumor Stratication and Noninvasive Diagnostic Biomarkers for
Bladder Cancer,” American Journal of Pathology, Vol. 180, No. 5,
2012, pp. 1808-1815. doi:10.1016/j.ajpath.2012.01.034
26. E. V. Grigorenko, E. Ortenberg, J. Hurley, A. Bond and K. Munnelly,
“miRNA Proling on High-Throughput OpenArray™ System,”
Methods in Molecular Biology, Vol. 676, 2011, pp. 101-110.
doi:10.1007/978-1-60761-863-8_8 [Citation Time(s):1]
27. R. E. Hershberger, J. Lindenfeld, L. Mestroni, C. E. Seidman, M. R.
Taylor and J. A. Towbin, “Genetic Evaluation of Cardiomyopathy:
A Heart Failure Society of America Practice Guideline,” Journal
of Cardiac Failure, Vol. 15, No. 2, 2009, pp. 83-97. doi:10.1016/j.
cardfail.2009.01.006 [Citation Time(s):1]
28. C. Andreasen, J. B. Nielsen, L. Refsgaard, A. G. Holst, A. H.
Christensen, L. Andreasen, A. Sajadieh, S. Haunsø, J. H. Svendsen
100
Advances in Molecular Diagnostics
and M. S. Olesen, “New Population-Based Exome Data Are
Questioning the Pathogenicity of Previously CardiomyopathyAssociated Genetic Variants,” European Journal of Human Genetics,
2013. doi:10.1038/ejhg.2012.283 [Citation Time(s):1]
29. D. Lassner, M. Rohde, U. M. Gross, F. Escher, H. P. Schultheiss, R.
P. Linke and U. Kühl, “Classication of Four Chemically Different
Amyloid Types in Routine Endomyocardial Biopsies by Advanced
Immunohistochemistry,” Amyloid, Vol. 18, Suppl. 1, 2011, pp. 76-
78. doi:10.3109/13506129.2011.574354027 [Citation Time(s):1]
30. D. A. Wheeler, M. Srinivasan, M. Egholm, et al., “The Complete
Genome of an Individual by Massively Parallel DNA Sequencing,”
Nature, Vol. 452, No. 7189. 2008, pp. 872-876. doi:10.1038/
nature06884 [Citation Time(s):1]
31. K. Hopp, K. Weber, D. Bellissimo, S. T. Johnson and B. Pietz,
“High-Throughput Red Blood Cell Antigen Genotyping Using a
Nanouidic Real-Time Polymerase Chain Reaction Platform,”
Transfusion, Vol. 50, No. 1, 2010, pp. 40-46. doi:10.1111/j.1537-
2995.2009.02377.x [Citation Time(s):1]
32. L. J. Cooper, J. M. Hare, H. D. Tazelaar, et al., “Usefulness of
Immunosupp. Ression for Giant Cell Myocarditis,” American
Journal of Cardiology, Vol. 102, 2008, pp. 1535-1539. doi:10.1016/j.
amjcard.2008.07.041
33. A. Frustaci, C. Chimenti, F. Calabrese, et al., “Immunosupp.
Ressive Therapy for Active Lymphocytic Myocarditis: Virological
and Immunologic Prole of Responders versus Nonresponders,”
Circulation, Vol. 107, 2003, pp. 857-863. doi:10.1161/01.
CIR.0000048147.15962.31
34. A. Frustaci, M. A. Russo and C. Chimenti, “Randomized Study
on the Efcacy of Immunosupp. Ressive Therapy in Patients with
Virus-Negative Inammatory Cardiomyopathy: The TIMIC Study,”
European Heart Journal, Vol. 30, No. 16, 2009, pp. 1995-2002.
doi:10.1093/eurheartj/ehp249 [Citation Time(s):1]
35. U. Kuhl, M. Pauschinger, P. L. Schwimmbeck, B. Seeberg, C.
Lober and M. Noutsias, “Interferon-Beta Treatment Eliminates
Cardiotropic Viruses and Improves Left Ventricular Function in
Patients with Myocardial Persistence of Viral Genomes and Left
Recent Advances in Molecular Diagnostics and Treatment of Heart...
101
Ventricular Dysfunction,” Circulation, Vol. 107, No. 22, 2003, pp.
2793- 2798. doi:10.1161/01.CIR.0000072766.67150.51
36. C. Schmidt-Lucke, F. Spillmann, T. Bock, U. Kühl, S. Van Linthout,
H.-P. Schultheiss and C. Tschöpe “Interferon Beta Modulates
Endothelial Damage in Patients with Cardiac Persistence of Human
Parvovirus b19 Infection,” The Journal of Infectious Diseases,
Vol. 201 No. 6, 2010, pp. 936-945. doi:10.1086/650700 [Citation
Time(s):1]
37. T. M. Wheeler, A. J. Leger, S. K. Pandey, A. R. MacLeod, M.
Nakamori, S. H. Cheng, B. M. Wentworth, C. F. Bennett and C.
A. Thornton, “Targeting Nuclear RNA for in Vivo Correction of
Myotonic Dystrophy,” Nature, Vol. 488, No. 7409, 2012, pp. 111-
115. doi:10.1038/nature11362 [Citation Time(s):1]
38. T. G. Hullinger, R. L. Montgomery, A. G. Seto, B. A. Dickinson, et
al., “Inhibition of miR-15 Protects against Cardiac Ischemic Injury,”
Circulation Research, Vol. 110, No. 1, 2012, pp. 71-81. doi:10.1161/
CIRCRESAHA.111.244442 [Citation Time(s):1]
39. L. T. Cooper, K. L. Baughman, A. M. Feldman, A. Frustaci, M.
Jessup, U. Kuhl, G. N. Levine, J. Narula, R. C. Starling, J. Towbin
and R. Virmani, American Heart Association, American College
of Cardiology, European Society of Cardiology, Heart Failure
Society of America and Heart Failure Association of the European
Society of Cardiology, “The Role of Endomyocardial Biopsy in
the Management of Cardiovascular Disease: A Scientic Statement
from the American Heart Association, the American College of
Cardiology, and the European Society of Cardiology. Endorsed
by the Heart Failure Society of America and the Heart Failure
Association of the European Society of Cardiology,” Journal of the
American College of Cardiology, Vol. 50, No. 19, 2007, pp. 1914-
1931. doi:10.1016/j.jacc.2007.09.008 [Citation Time(s):1]
MOLECULAR DIAGNOSIS OF
BRUCELLOSIS: A BRIEF
6
REPORT
Panagiotis Andriopoulos and Maria Tsironi
Department of Nursing, University of Peloponnese, Valioti and Plataion, Sparta, Greece
ABSTRACT
Amplification of nucleic acid by polymerase chain reaction (PCR)
assays in order to diagnose infection by Brucella spp. has been used
for more than two decades. Human Brucellosis is an endemic disease
in many countries worldwide and often poses diagnostic puzzles. The
implementation of PCR (standard, real-time and multiplex) can help
in the accurate differential diagnosis and distinguish between acute,
subacute and chronic cases. PCR has also been used for follow up of
patients and serotype identification of Brucella spp. Overall PCR is a
promising and reliable technique for the diagnosis of Human Brucellosis.
Purpose of this brief report is to identify possible alternatives for rapid
and accurate diagnosis of Brucellosis using PCR.
104
Keywords: Polymerase chain reaction; Human Brucellosis; Brucella
spp; Molecular diagnosis
Advances in Molecular Diagnostics
INTRODUCTION
Brucellosis is a commonzoonoses of global distribution [1]. New incident
cases are estimated around 500.000 each year worldwide, whereas
prevalence is approximately 10/100.000 people in endemic areas. The
disease is caused by Brucella spp. and most usual isolated serotypes are
Melitensis, Abortus and Suis. Brucellae are Gram negative intracellular
bacteria that can multiply within phagocytic cells with human beings
acting as end hosts. Pathogens enter the human body through ingestion,
inhalation, the conjunctiva or skin abrasions. The pathogens reside in
the reticuloendothelial system, granulomas are formed and finally
bacteraimia may follow. The mechanisms leading to intracellular killing
of the host immune system are unknown [2]. Brucellosis can present
as acute, subacute or chronic disease with symptoms according to the
various affected organs [3]. Common symptoms include arthralgias, fever,
sweating, lack of appetite, weight loss and low back pain. On clinical
presentation patients often present with splenomegaly, hepatomegaly
or both, cervical lymphadenopathy and peripheral arthritis [2,3].
Neurobrucellosis and endocarditis are the two potentially life threatening
localizations of brucellosis, but osteoarticular, genitourinary and
gastrointestinal involvement are far more common. Laboratory diagnosis
is usually made either by isolation and culture of the microorganism or
by serology tests [4]. Rarely, some patients with brucellosis will have a
positive blood culture in the absence of positive serology. The automated
continuously monitored blood culture systems have shortened the time to
diagnosis to a maximum of 2 weeks (mean days to results 4-5). Serology
methods include the well-studied and established Rose Bengal and Wight
Coombs agglutination tests; both have limitations due to low predictive
values [4-6]. Newer tests such as Brucellacapt and detection of IgG
and IgM antibodies by ELISAare not available worldwide due to lack
of resources [6]. However, diagnostic puzzles are always present and
difficulties have been noted: Cultures require level 3 biocontainment
facilities and highly skilled technical personnel to handle samples and
live bacteria for eventual identification and biotyping. Serology cannot
Molecular Diagnosis of Brucellosis: A Brief Report
be used for diagnosis alone in endemic areas and a verification test is
often required, either cultures or repeating the tests a few days later [27]. Polymerase chain reaction (PCR) methods have been implemented in
order to enhance sensitivity and produce quicker results [8]. Nucleic acid
amplification techniques are now quite widely used, although no single
standardized procedure has been adopted. More than 400 reports have
been published describing various PCR based methods for the diagnosis
of Brucella infections [7]. We provide here a brief review of the proposed
methods and an appraisal of their usage in clinical settings for diagnosing
Brucellosis.
105
MOLECULAR METHODS
Standard PCR
The first reports for implementation of PCR for diagnosis of Brucellosis
date from the early 90’s. [9-11]. Blood is the preferred tissue for the
extraction of DNA, [12] however various others tissues such as serum,
semen, or synovial fluid have been used [13]. New approaches include the
washing of blood a few times with water in order to avoid contamination
with hemoglobin. Such techniques improve the sensitivity of the
procedure. [14]. The standard PCR assays include one pair of primers
which is used to amplify the target genomic sequence of Brucella
spp. Pairs used include the primers for sequences encoding 16S rRNA
[15,16], outer membrane protein (omp2a, omp2b and omp31) [17,18],
31kDa immunogenic Brucellaabortus protein (BCSP 31 B4/B5) [19-21],
16S-23S ribosomal DNA interspace region (ITS66/ITS279) [22,23] and
insertion sequence (IS711) [24,25]. These reports showed an excellent
sensitivity for the diagnosis not only of the acute disease but also for the
follow up pf patients and the detection of relapses where serology and
cultures are often negative [26-28].
Real time PCR
Real time PCR for brucellosis has also been used. The ability to measure
DNA copy number and mRNA expression levels together with the
rapid detection and differentiation of Brucella spp. and the decreasing
prices have made the procedure attractive and accessible [29] Real-
106
Advances in Molecular Diagnostics
time PCR seems to be highly reproducible, rapid, sensitive and specific.
Additionally, this assay is easily standardized and the risk of infection
in laboratory workers is minimal [8] Samples that have been tested by
real time PCR include cultured Brucella cells [30], serum [31], bloodand
paraffin-embedded tissues [32]. Some researchers even propose the use
of real-time PCR for the diagnosis of human brucellosis in everyday
clinical practice since various reports give a time to final results of only
30 minutes [30]. It is also suggested that it is the method of choice for the
discrimination among inactive, seropositive and active states in testing
serum samples for subjects whose clinical findings are known [32].The
predominance of real-time PCR in terms of sensitivity and specificity is
well documented in various studies. Queipo-Ortuno et al.[31] performed
real-time PCR with SYBR LightCycler Green I in blood cultures of serum
samples and whole blood of patients with brucellosis using primers B4
and B5 (targeting bspc31) and compared their results with PCR-enzymelinked immunosorbent assay [32]. Real-time PCR in serum samples had
better sensitivity. Surucuoglu et al. [33] used theTaqMan real time PCR
technique which targeted the IS711, bcsp31 and per genes in patients
with various clinical forms of brucellosis and compared the results of
their method with other conventional methods using serum samples.
The IS711-based assay was the most sensitive, specific, efficient,
and reproducible method to detect Brucella spp. Further reports have
documented the specificity, sensitivity and rapid results of real-time PCR
[34,35].
Multiplex PCR
The multiplex PCRs that have been developed the previous years have
included also Brucella spp. with the first report dating from 1994
[36]. Many reports can be found in the literature; the most interesting
studies used multiplex PCR to simultaneous detect Brucellaspp. and
Mycobacterium Tuberculosis complex [37-39]. The procedure targeted
the IS711, bcsp31 and omp2agenes for Brucella spp. and the IS6110,
senX3-regX3 and cfp31 genes for M. tuberculosis complex. Since
brucellosis is endemic in countries that are also endemic for tuberculosis
[40], a rapid diagnostic test, especially in chronic and atypical cases is of
great importance.