Accessibility / Report Error

Prevalence and association of mycoplasma infection in the development of coronary artery disease

Prevalência e associação de infecção por micoplasma no desenvolvimento de doença arterial coronariana

Abstract

Coronary heart disease (CHD) has been associated with significant morbidity and mortality worldwide. Although remain controversial, several studies have demonstrated the association of M. pneumoniae infections with atherosclerosis. We evaluated the possible association of mycoplasma infections in patients diagnosed with atherosclerosis by ELISA and PCR methods. Atherosclerotic tissue samples and blood samples were collected for the detection of mycoplasma antibodies (IgA) by ELISA from the 97 patients with coronary artery disease (CAD). M. pneumoniae specific IgA, IgG and IgM were measured by using the Anti-M. pneumoniae IgA/IgG/IgM ELISA. Detection of M. pneumoniae targeting the P1 adhesion gene was performed by PCR Acute infection of M. pneumoniae was diagnosed in 43.3% (42) of patients by PCR. The M. pneumoniae specific antibodies were detected in 36.1% (35) of patients. Twenty-five (25.8%) cases had IgG antibodies, 15 (15.5%) cases had IgM antibodies, 3 (3.1%) cases had IgA antibodies, 10 (10.3%) cases had both IgM + IgG antibodies and 1 (1%) case of each had IgM + IgA and IgG + IgA antibodies. None of the cases was positive for all three antibodies. A Pearson correlation coefficient analysis revealed an excellent correlation between the PCR and the serological results (r=0.921, p<0.001). A majority (17, 40.5%) of the M. pneumoniae positive patients are within the 41-50 years of age group, followed by 10 (23.8%) patients in the age group of 61-70 years and 2 (4.8%) patients were >70 years of age. Our study reported an unusually higher prevalence of M. pneumoniae by serological tests (36.1%) and PCR (43.3%). Although the hypothesis of the association of M. pneumoniae and CAD is yet to be proven, the unusually high prevalence of M. pneumoniae in CAD patients indicates an association, if not, in the development of atherosclerosis.

Keywords:
coronary heart disease; mycoplasma; atherosclerosis; ELISA; PCR

Resumo

A doença coronariana (DCC) tem sido associada a significativa morbidade e mortalidade em todo o mundo. Embora ainda sejam controversos, vários estudos têm demonstrado a associação de infecções por M. pneumoniae com aterosclerose. Avaliamos a possível associação de infecções por micoplasma em pacientes com diagnóstico de aterosclerose pelos métodos ELISA e PCR. Amostras de tecido aterosclerótico e amostras de sangue foram coletadas para a detecção de anticorpos contra micoplasma (IgA) por ELISA de 97 pacientes com doença arterial coronariana (DAC). IgA, IgG e IgM específicos para M. pneumoniae foram medidos usando o Anti-M. pneumoniae IgA / IgG / IgM ELISA. A detecção de M. pneumoniae visando o gene de adesão P1 foi realizada por PCR. A infecção aguda por M. pneumoniae foi diagnosticada em 43,3% (42) dos pacientes pela PCR. Os anticorpos específicos para M. pneumoniae foram detectados em 36,1% (35) dos pacientes. Vinte e cinco (25,8%) casos tinham anticorpos IgG, 15 (15,5%) casos tinham anticorpos IgM, 3 (3,1%) casos tinham anticorpos IgA, 10 (10,3%) casos tinham anticorpos IgM + IgG e 1 (1%) caso de cada um tinha anticorpos IgM + IgA e IgG + IgA. Nenhum dos casos foi positivo para os três anticorpos. A análise do coeficiente de correlação de Pearson revelou uma excelente correlação entre o PCR e os resultados sorológicos (r = 0,921, p < 0,001). A maioria (17, 40,5%) dos pacientes positivos para M. pneumoniae está na faixa etária de 41-50 anos, seguida por 10 (23,8%) pacientes na faixa etária de 61-70 anos e 2 (4,8%) pacientes tinham > 70 anos de idade. Nosso estudo relatou uma prevalência incomumente maior de M. pneumoniae por testes sorológicos (36,1%) e PCR (43,3%). Embora a hipótese da associação de M. pneumoniae e DAC ainda não tenha sido comprovada, a prevalência incomumente alta de M. pneumoniae em pacientes com DAC indica uma associação, se não, no desenvolvimento de aterosclerose.

Palavras-chave:
doença cardíaca coronariana; micoplasma; aterosclerose; ELISA; PCR

1. Introduction

Coronary heart disease (CHD) has been associated with significant morbidity and mortality worldwide. Several well-recognized risk factors include smoking, diet, hypercholesterolemia, hypertension, diabetes and sedentary habits (Alviar et al., 2011ALVIAR, C.L., ECHEVERRI, J.G., JARAMILLO, N.I., FIGUEROA, C.J., CORDOVA, J.P., KORNIYENKO, A., SUH, J. and PANIZ-MONDOLFI, A., 2011. Infectious atherosclerosis: is the hypothesis still alive? A clinically based approach to the dilemma. Medical Hypotheses, vol. 76, no. 4, pp. 517-521. http://dx.doi.org/10.1016/j.mehy.2010.12.006. PMid:21216537.
http://dx.doi.org/10.1016/j.mehy.2010.12...
). Atherosclerosis is a disease in which plaques build up inside the arteries and limits the flow of oxygen-rich blood to various parts of the body. The association of infectious agents with the atherosclerosis was first demonstrated in 1978 (Fabricant et al., 1978FABRICANT, C.G., FABRICANT, J., LITRENTA, M.M. and MINICK, C.R., 1978. Virus-induced atherosclerosis. The Journal of Experimental Medicine, vol. 148, no. 1, pp. 335-340. http://dx.doi.org/10.1084/jem.148.1.335. PMid:209124.
http://dx.doi.org/10.1084/jem.148.1.335...
). It has been hypothesized that during atherosclerosis, several common bacteria and virus contributes to the inflammation of vascular heart wall and leads to the formation of atheroma (Ross, 1999ROSS, R., 1999. Atherosclerosis--an inflammatory disease. The New England Journal of Medicine, vol. 340, no. 2, pp. 115-126. http://dx.doi.org/10.1056/NEJM199901143400207. PMid:9887164.
http://dx.doi.org/10.1056/NEJM1999011434...
; Ohayon et al., 2011OHAYON, J., GHARIB, A.M., GARCIA, A., HEROUX, J., YAZDANI, S.K., MALVE, M., TRACQUI, P., MARTINEZ, M.A., DOBLARE, M., FINET, G. and PETTIGREW, R.I., 2011. Is arterial wall-strain stiffening an additional process responsible for atherosclerosis in coronary bifurcations?: An in vivo study based on dynamic CT and MRI. American Journal of Physiology. Heart and Circulatory Physiology, vol. 301, no. 3, pp. H1097-H1106. http://dx.doi.org/10.1152/ajpheart.01120.2010. PMid:21685261.
http://dx.doi.org/10.1152/ajpheart.01120...
). Although the role of infectious agents remains controversial, several studies demonstrated the association of viruses and bacteria in the development of atherosclerosis (Góis et al., 2006GÓIS, J., HIGUCHI, M., REIS, M., DIAMENT, J., SOUSA, J., RAMIRES, J. and OLIVEIRA, S., 2006. Infectious agents, inflammation, and growth factors: how do they interact in the progression or stabilization of mild human atherosclerotic lesions? Annals of Vascular Surgery, vol. 20, no. 5, pp. 638-645. http://dx.doi.org/10.1007/S10016-006-9076-1. PMid:16983590.
http://dx.doi.org/10.1007/S10016-006-907...
). Even though there is no usually acknowledged group of bacteria or technique available for evaluating infectious atherosclerosis, reports have associated Mycoplasma pneumoniae (Momiyama et al., 2004MOMIYAMA, Y., OHMORI, R., TANIGUCHI, H., NAKAMURA, H. and OHSUZU, F., 2004. Association of mycoplasma pneumoniae infection with coronary artery disease and its interaction with chlamydial infection. Atherosclerosis, vol. 176, no. 1, pp. 139-144. http://dx.doi.org/10.1016/j.atherosclerosis.2004.04.019. PMid:15306186.
http://dx.doi.org/10.1016/j.atherosclero...
), Chlamydia pneumoniae (Hauer et al., 2006HAUER, A.D., DE VOS, P., PETERSE, N., TEN CATE, H., VAN BERKEL, T.J., STASSEN, F.R. and KUIPER, J., 2006. Delivery of chlamydia pneumoniae to the vessel wall aggravates atherosclerosis in LDLr-/- mice. Cardiovascular Research, vol. 69, no. 1, pp. 280-288. http://dx.doi.org/10.1016/j.cardiores.2005.07.011. PMid:16112098.
http://dx.doi.org/10.1016/j.cardiores.20...
) and influenza viruses as a potential risk factor of CHD (Corrales-Medina et al., 2010CORRALES-MEDINA, V.F., MADJID, M. and MUSHER, D.M., 2010. Role of acute infection in triggering acute coronary syndromes. The Lancet. Infectious Diseases, vol. 10, no. 2, pp. 83-92. http://dx.doi.org/10.1016/S1473-3099(09)70331-7. PMid:20113977.
http://dx.doi.org/10.1016/S1473-3099(09)...
).

M. pneumoniae are auto-replicating, the smallest microorganism that is devoid of the cell wall and requires cholesterol for its growth (Razin et al., 1998RAZIN, S., YOGEV, D. and NAOT, Y., 1998. Molecular biology and pathogenicity of mycoplasmas. Microbiology and Molecular Biology Reviews, vol. 62, no. 4, pp. 1094-1156. http://dx.doi.org/10.1128/MMBR.62.4.1094-1156.1998. PMid:9841667.
http://dx.doi.org/10.1128/MMBR.62.4.1094...
). They may also favor the proliferation of other infectious organisms by altering the host immune system (Higuchi et al., 2003HIGUCHI, M.L., REIS, M.M., SAMBIASE, N.V., PALOMINO, S.A., CASTELLI, J.B., GUTIERREZ, P.S., AIELLO, V.D., RAMIRES, J.A., 2003. Coinfection with Mycoplasma Pneumoniae and Chlamydia Pneumoniae in ruptured plaques associated with acute myocardial infarction. Arquivos Brasileiros de Cardiologia, vol. 81, no. 1, pp. 12-22.). The association of C. pneumonia with atherosclerosis has been reported earlier (Danesh et al., 1997DANESH, J., COLLINS, R. and PETO, R., 1997. Chronic infections and coronary heart disease: is there a link? Lancet, vol. 350, no. 9075, pp. 430-436. http://dx.doi.org/10.1016/S0140-6736(97)03079-1. PMid:9259669.
http://dx.doi.org/10.1016/S0140-6736(97)...
). Since there is a similarity in epidemiological behavior and antibiotic resistance pattern between M. pneumoniae and C. pneumoniae, it has been suggested that M. pneumoniae may also play a role in the development of atherosclerosis (Taylor-Robinson and Thomas, 1998TAYLOR-ROBINSON, D. and THOMAS, B.J., 1998. Chlamydia pneumoniae in arteries: the facts, their interpretation, and future studies. Journal of Clinical Pathology, vol. 51, no. 11, pp. 793-797. http://dx.doi.org/10.1136/jcp.51.11.793. PMid:10193317.
http://dx.doi.org/10.1136/jcp.51.11.793...
). Higuchi et al. (2000)HIGUCHI, M.L., SAMBIASE, N., PALOMINO, S., GUTIERREZ, P., DEMARCHI, L.M., AIELLO, V.D. and RAMIRES, J.A., 2000. Detection of mycoplasma pneumoniae and chlamydia pneumoniae in ruptured atherosclerotic plaques. Brazilian Journal of Biology = Revista Brasileira de Biologia, vol. 33, no. 9, pp. 1023-1026. PMid:10973132., during the detection of C. pneumonia from necropsy specimens, demonstrated the presence of M. pneumoniae for the first time (Higuchi et al., 2000HIGUCHI, M.L., SAMBIASE, N., PALOMINO, S., GUTIERREZ, P., DEMARCHI, L.M., AIELLO, V.D. and RAMIRES, J.A., 2000. Detection of mycoplasma pneumoniae and chlamydia pneumoniae in ruptured atherosclerotic plaques. Brazilian Journal of Biology = Revista Brasileira de Biologia, vol. 33, no. 9, pp. 1023-1026. PMid:10973132.). Thereafter few other studies have also demonstrated the association of M. pneumoniae infections with atherosclerosis (Momiyama et al., 2004MOMIYAMA, Y., OHMORI, R., TANIGUCHI, H., NAKAMURA, H. and OHSUZU, F., 2004. Association of mycoplasma pneumoniae infection with coronary artery disease and its interaction with chlamydial infection. Atherosclerosis, vol. 176, no. 1, pp. 139-144. http://dx.doi.org/10.1016/j.atherosclerosis.2004.04.019. PMid:15306186.
http://dx.doi.org/10.1016/j.atherosclero...
; Reunanen et al., 2005REUNANEN, A., ROIVAINEN, M. and KLEEMOLA, M., 2005. Increased titer of antibodies to mycoplasma pneumoniae may be associated with coronary heart disease. Atherosclerosis, vol. 180, no. 1, pp. 209-210. http://dx.doi.org/10.1016/j.atherosclerosis.2004.12.032. PMid:15823295.
http://dx.doi.org/10.1016/j.atherosclero...
). In contrast, other studies reported discordant results regarding the role of M. pneumoniae in atherosclerosis pathogenesis. Thus, the association of M. pneumoniae is controversial and remains as unknown. The present study aims to evaluate the possible association of mycoplasma infections in patients diagnosed with atherosclerosis by ELISA and PCR methods.

2. Materials and Methods

2.1. Patients

A prospective study was conducted in 97 patients with coronary artery disease (CAD) admitted to the Department of Cardiology and underwent coronary artery bypass surgery (CABG) between February 2016 and November 2019. Fifty-eight (59.8%) patients were male and 39 (40.2%) were female. The age ranged from 38 to 73 years (mean (SD) 56.3 ± 7.8 years). An intraoperative endarterectomy was performed to obtain the atherosclerotic tissue sample during CABG. Blood samples were collected for the detection of mycoplasma antibodies (IgA) by ELISA from the CAD patients. Blood samples collected from 10 normal individuals whose baseline characteristics were similar to the patient group serve as a control for the ELISA test. Clinical characteristics of patients were obtained from the medical records. Tissue samples and serum separated from the collected blood samples were stored at -200C until further analysis. A written informed consent was obtained from each patient before surgery and the institutional ethics committee approved the study.

2.2. Serological analysis

Mycoplasma pneumoniae specific IgA, IgG and IgM were measured by using the Anti-Mycoplasma pneumoniae IgA/IgG/IgM ELISA Kit (Verion/Serion, Germany) as per the manufacturer instructions. Absorbance within 30 min of the addition of the stop solution was measured at 450 nm. As indicated by the manufacturer, a titer of IgA >14 U/ml, a titer of IgG >30 U/ml and a titer of IgG >1.1RU were considered as positive.

2.3. DNA extraction

DNA from the atherosclerotic tissues was extracted using the DNeasy® Tissue kit (Qiagen, Germany) according to the manufacturer’s instructions. For the PCR amplification, 100 ng of DNA was used. The purity and the quantification of DNA were checked using the NanoDropTM spectrophotometer (Thermo Fisher Scientific, USA). Extraction of DNA, preparation of PCR mix, PCR amplification, and analysis of PCR products were performed in separate laboratories.

2.4. Polymerase chain reaction

Detection of M. pneumoniae targeting the P1 adhesion gene was performed by PCR (Chaudhry et al., 2013CHAUDHRY, R., SHARMA, S., JAVED, S., PASSI, K., DEY, A.B. and MALHOTRA, P., 2013. Molecular detection of mycoplasma pneumoniae by quantitative real-time PCR in patients with community acquired pneumonia. The Indian Journal of Medical Research, vol. 138, pp. 244-251. PMid:24056602.). PCR was performed using a 25μl master mix containing 2.5μl of template DNA, 0.2µM (2μl) of each primer (Forward: 5’- CAAGCCAAACACGAGCTCCGGCC-3’ and Reverse: 5’-CAGTGTCAGCTGTTTGTCCTTCCCC-3’), 12.5 μl of TEMPase Hot Start Master Mix (Ampliqon, Denmark) and 6μl of DNAse/ RNAse free water. The following PCR cycling conditions were used: initial denaturation at 94°C for 2 min followed by 35 cycles at 94°C for 1 min, 55°C for 1 min and 72°C for 2 min and a final extension step at 72°C for 10 min. An expected amplicon size of 543 bp was resolved in 1.2% agarose gel electrophoresis and visualized under UV transilluminator.

2.5. Statistical analysis

Data with continuous values were represented as medians and ranges and categorical values as numbers and percentages. A Chi-Square test was performed to determine statistical significance and a Pearson correlation coefficient analysis was performed to determine the relationship of two variables using SPSS statistical software (IBM® SPSS®, USA). A P value < 0.05 was considered to be statistically significant.

3. Results

A total of 97 blood samples were collected from patients with CAD who underwent CABG. The mean age of the patients was 56.3 ± 7.8 years. Twelve patients were in <40 years of age range, 36 patients were in 41-50 years of age range, 28 patients were in 51-60 years of age range, 17 patients were in 61-70 years of age range and 4 patients were in >70 years of age range (Figure 1). The predominant age group affected by CAD ranged from 41-50 years, followed by the 51-60 years. The distribution of CAD patients based on sex and other demographic details like family history and history of myocardial infarction is shown in Table 1. The clinical data of all the patients are presented in Table 2.

Figure 1
Distribution of age among the included patients.
Table 1
Demographic detail of CAD patients.
Table 2
Clinical characteristics of CAD patients.

Acute infection of M. pneumoniae was diagnosed in 43.3% (42) of DNA samples using molecular detection of P1 adhesion gene. The anti-M. pneumoniae were positive in 36.1% of cases and negative in 63.9% of cases. There was no significant difference between the PCR and the serological results (p=0.358) (Table 3). Similarly, there were no significant differences in the detection of M. pneumoniae by PCR and serology between men and women (p>0.05). Twenty-five (25.8%) cases had IgG antibodies, 15 (15.5%) cases had IgM antibodies, 3 (3.1%) cases had IgA antibodies, 10 (10.3%) cases had both IgM + IgG antibodies and 1 (1%) case of each had IgM + IgA and IgG + IgA antibodies. None of the cases was positive for all three antibodies (Figure 2). A sub-group analysis of the 35 (36.1%) anti-M. pneumoniae seropositive patients revealed that 30 (35.7%) patients had hypertension, 29 (82.9%) patients had myocardial infarction, 27 (77.1%) patients had the total cholesterol of >250 mg/dl, 24 (68.6%) patients had LDL of >60 mg/dl, 22 (62.9%) patients had triglycerides of >150 mg/dl, 18 (51.4%) patients had HDL of <100 mg/dl and 15 (42.9%) patients had an urea level of >40 mg/dl.

Table 3
Molecular and serology results of M. pneumoniae.
Figure 2
Serological test of M. pneumoniae.

Of the 35 (36.1%) samples positive for any of the tested antibodies, 31 (32.0%) samples were positive for M. pneumoniae by PCR. A Pearson correlation coefficient analysis revealed an excellent correlation between the PCR and the serological results (r=0.921, p<0.001). A sub-group analysis of the 42 (43.3%) patients positive for M. pneumoniae by PCR revealed that 37 (88.1%) patients had hypertension, 33 (78.6%) patients had myocardial infarction, 29 (69%) patients had the total cholesterol of >250 mg/dl, 26 (61.9%) patients had triglycerides of >150 mg/dl, 25 (59.5%) patients had LDL of >60 mg/dl, 21 (50%) patients had HDL of <100 mg/dl and 17 (40.5%) patients had an urea level of >40 mg/dl. There was no significant statistical difference in the sex and age proportions between the test and control groups (p<0.05). A majority (17, 40.5%) of the M. pneumoniae positive patients are within the 41-50 years of age group, followed by 10 (23.8%) patients in the age group of 61-70 years and 2 (4.8%) patients were >70 years of age.

4. Discussion

M. pneumoniae could be a possible candidate to play a role in the pathogenesis of atherosclerosis, as it has been related to cardiovascular disease and its ability to induce chronic inflammation (Higuchi et al., 2000HIGUCHI, M.L., SAMBIASE, N., PALOMINO, S., GUTIERREZ, P., DEMARCHI, L.M., AIELLO, V.D. and RAMIRES, J.A., 2000. Detection of mycoplasma pneumoniae and chlamydia pneumoniae in ruptured atherosclerotic plaques. Brazilian Journal of Biology = Revista Brasileira de Biologia, vol. 33, no. 9, pp. 1023-1026. PMid:10973132.). Even though M. pneumoniae accounts for 10%-30% of commonly acquired pneumonia, some studies have identified that M. pneumoniae is also present in atherosclerotic plaque (Higuchi et al., 2000HIGUCHI, M.L., SAMBIASE, N., PALOMINO, S., GUTIERREZ, P., DEMARCHI, L.M., AIELLO, V.D. and RAMIRES, J.A., 2000. Detection of mycoplasma pneumoniae and chlamydia pneumoniae in ruptured atherosclerotic plaques. Brazilian Journal of Biology = Revista Brasileira de Biologia, vol. 33, no. 9, pp. 1023-1026. PMid:10973132.). Furthermore, this bacterium is identified in the serological antibody test in patients with CAD (Momiyama et al., 2004MOMIYAMA, Y., OHMORI, R., TANIGUCHI, H., NAKAMURA, H. and OHSUZU, F., 2004. Association of mycoplasma pneumoniae infection with coronary artery disease and its interaction with chlamydial infection. Atherosclerosis, vol. 176, no. 1, pp. 139-144. http://dx.doi.org/10.1016/j.atherosclerosis.2004.04.019. PMid:15306186.
http://dx.doi.org/10.1016/j.atherosclero...
). The association between M. pneumoniae infection and cardiovascular events remains undetermined. Therefore the present study was attempted to determine the prevalence of M. pneumoniae in adult patients with CAD using PCR and ELISA tests. The interpretation of serological results should be based on a combination of IgA, IgG and IgM antibodies’ profiles. The presence of M. pneumoniae IgA, IgG or IgM in the serum generally reflects current/recurrent, past or current infection, respectively (Ngeh et al., 2004NGEH, J., GUPTA, S., GOODBOURN, C. and MCELLIGOTT, G., 2004. Mycoplasma pneumoniae in elderly patients with stroke. A case-control study on the seroprevalence of M. Pneumoniae in elderly patients with acute cerebrovascular disease - the m-peps study. Cerebrovascular Diseases (Basel, Switzerland), vol. 17, no. 4, pp. 314-319. http://dx.doi.org/10.1159/000077342. PMid:15026614.
http://dx.doi.org/10.1159/000077342...
). In our study, M. pneumoniae antibodies were present in 36.1% of patients. Of these, 25.8% of patients had IgG antibodies, 15.5% of patients had IgM antibodies, 3.1% of patients had IgA antibodies, 10.3% of patients had both IgM + IgG antibodies and 1% of patient each had IgM + IgA and IgG + IgA antibodies. The overall seroprevalence of M. pneumoniae infection in our study was found to be higher than reported in other studies (Hauksdottir et al., 1998HAUKSDÓTTIR, G.S., JÓNSSON, T., SIGURDARDÓTTIR, V. and LÖVE, A., 1998. Seroepidemiology of mycoplasma pneumoniae infections in iceland 1987-96. Scandinavian Journal of Infectious Diseases, vol. 30, no. 2, pp. 177-180. http://dx.doi.org/10.1080/003655498750003591. PMid:9730307.
http://dx.doi.org/10.1080/00365549875000...
; Rastawicki et al., 1998RASTAWICKI, W., KALUZEWSKI, S. and JAGIELSKI, M., 1998. Occurrence of serologically verified mycoplasma pneumoniae infections in poland in 1970-1995. European Journal of Epidemiology, vol. 14, no. 1, pp. 37-40. http://dx.doi.org/10.1023/A:1007431932087. PMid:9517871.
http://dx.doi.org/10.1023/A:100743193208...
; Tuuminen et al., 2000TUUMINEN, T., VARJO, S., INGMAN, H., WEBER, T., OKSI, J. and VILJANEN, M., 2000. Prevalence of chlamydia pneumoniae and mycoplasma pneumoniae immunoglobulin g and a antibodies in a healthy finnish population as analyzed by quantitative enzyme immunoassays. Clinical and Diagnostic Laboratory Immunology, vol. 7, no. 5, pp. 734-738. http://dx.doi.org/10.1128/CDLI.7.5.734-738.2000. PMid:10973446.
http://dx.doi.org/10.1128/CDLI.7.5.734-7...
; Daxboeck et al., 2002DAXBOECK, F., KIRCHER, K., KRAUSE, R., HEINZL, H., WENISCH, C. and STANEK, G., 2002. Effect of age on antibody titer to mycoplasma pneumoniae. Scandinavian Journal of Infectious Diseases, vol. 34, no. 8, pp. 577-579. http://dx.doi.org/10.1080/00365540110089836. PMid:12238572.
http://dx.doi.org/10.1080/00365540110089...
). However, a study from Austria which investigated 91 patients with internal carotid artery (ICA) stenosis for specific IgA antibodies to M. pneumoniae, reported a higher seroprevalence rate (18.7%) than that reported (3.1%) in our study (Daxbock et al., 2011DAXBÖCK, F., ASSADIAN, A., WATKINS-RIEDEL, T. and ASSADIAN, O., 2011. Persistently elevated iga antibodies to mycoplasma pneumoniae in patients with internal carotid artery stenosis. GMS Krankenhaushygiene Interdisziplinär, vol. 6, no. 1, pp. Doc04. PMid:22242085.). Another study from Saudi Arabia, which demonstrated the association of M. pneumoniae with ischemic heart disease (IHD), reported a higher rate of M pneumonia IgG antibodies (31.3%) in IHD patients than that reported (25.8%) in our study (Yiallouros et al., 2013YIALLOUROS, P., MOUSTAKI, M., VOUTSIOTI, A., SHARIFI, F. and KARPATHIOS, T., 2013. Association of mycoplasma pneumoniae infection with henoch-schonlein purpura. Prague Medical Report, vol. 114, no. 3, pp. 177-179. http://dx.doi.org/10.14712/23362936.2014.20. PMid:24093818.
http://dx.doi.org/10.14712/23362936.2014...
). A study from Japan evaluated the seropositivity for M. pneumoniae and C. pneumoniae in 396 patients with CAD, and 153 patients without CAD. The study reported that M. pneumoniae seropositivity (antibody titer ≥1/8 and ≥1/16) were significantly higher in patients with CAD than in patients without CAD, especially in those with myocardial infarction (Momiyama et al., 2004MOMIYAMA, Y., OHMORI, R., TANIGUCHI, H., NAKAMURA, H. and OHSUZU, F., 2004. Association of mycoplasma pneumoniae infection with coronary artery disease and its interaction with chlamydial infection. Atherosclerosis, vol. 176, no. 1, pp. 139-144. http://dx.doi.org/10.1016/j.atherosclerosis.2004.04.019. PMid:15306186.
http://dx.doi.org/10.1016/j.atherosclero...
). Similarly, in our study higher number of (29, 82.9%) patients who were positive for M. pneumoniae antibodies had a myocardial infarction, which is in agreement with that reported by Momiyama et al. (Momiyama et al., 2004). Horne et al., in a case-control study, reported that a significantly higher number of patients with elevated levels of IgA (not IgG) for M. pneumonia had coronary heart disease (Watkins-Riedel et al., 2001WATKINS-RIEDEL, T., STANEK, G. and DAXBOECK, F., 2001. Comparison of seromp iga with four other commercial assays for serodiagnosis of mycoplasma pneumoniae pneumonia. Diagnostic Microbiology and Infectious Disease, vol. 40, no. 1-2, pp. 21-25. http://dx.doi.org/10.1016/S0732-8893(01)00250-4. PMid:11448559.
http://dx.doi.org/10.1016/S0732-8893(01)...
). A recent study reported that a high level of anti-Mycoplasma pneumoniae IgM and IgG antibodies indicate a significant association of M. pneumoniae infection and history of this infection with increased risk for ischemic stroke (Roham et al., 2016ROHAM, M., ANBARI, K., MIRHABIBI, S. and GOUDARZI, G., 2016. The seroprevalence of mycoplasma pneumoniae IgM and IgG antibodies in patients with ischemic stroke. Iranian Journal of Microbiology, vol. 8, no. 6, pp. 383-388. PMid:28491249.). In our study, 10.3% of patients were sero-positive for both IgM and IgG antibodies, which indicates that these patients may be with an increased risk of ischemic stroke.

In our study, a total of 42 (43.3%) patients were positive for M. pneumonia by PCR. Maraha et al., investigated the presence of M. pneumoniae in 39 atherectomy specimens and in 64 degenerative heart valve (DHV) specimens. The study reported that only 2.5% of atherectomy and 3% of DHV specimens were positive for M. pneumoniae by PCR, which is much lower than that reported in our study (Maraha et al., 2000MARAHA, B., VAN DER ZEE, A., BERGMANS, A.M., PAN, M., PEETERS, M.F., BERG, H.F., SCHEFFER, G.J. and KLUYTMANS, J.A., 2000. Is mycoplasma pneumoniae associated with vascular disease? Journal of Clinical Microbiology, vol. 38, no. 2, pp. 935-936. http://dx.doi.org/10.1128/JCM.38.2.935-936.2000. PMid:10655422.
http://dx.doi.org/10.1128/JCM.38.2.935-9...
). Bayram et al., who investigated the presence of bacteria and viruses in atherosclerotic lesions of patients with CAD, reported that 6.7% atherosclerotic specimens and 3.3% on non-atherosclerotic vascular tissues samples were positive for M. pneumoniae by PCR (Bayram et al., 2011BAYRAM, A., ERDOGAN, M.B., EKSI, F., and YAMAK, B., 2011. Demonstration of chlamydophila pneumoniae, mycoplasma pneumoniae, cytomegalovirus, and epstein-barr virus in atherosclerotic coronary arteries, nonrheumatic calcific aortic and rheumatic stenotic mitral valves by polymerase chain reaction. Anadolu kardiyoloji dergisi : AKD = the Anatolian Journal of Cardiology, vol. 11, no. 3, pp. 237-243. http://dx.doi.org/10.5152/akd.2011.057.
http://dx.doi.org/10.5152/akd.2011.057...
). In our study, the result of PCR is coherent with serology, which is concordant with the findings in other studies (Welti et al., 2003WELTI, M., JATON, K., ALTWEGG, M., SAHLI, R., WENGER, A. and BILLE, J., 2003. Development of a multiplex real-time quantitative pcr assay to detect chlamydia pneumoniae, legionella pneumophila and mycoplasma pneumoniae in respiratory tract secretions. Diagnostic Microbiology and Infectious Disease, vol. 45, no. 2, pp. 85-95. http://dx.doi.org/10.1016/S0732-8893(02)00484-4. PMid:12614979.
http://dx.doi.org/10.1016/S0732-8893(02)...
; Kumar et al., 2016KUMAR, S., SAIGAL, S.R., SETHI, G.R. and KUMAR, S., 2016. Application of serology and nested polymerase chain reaction for identifying chlamydophila pneumoniae in community-acquired lower respiratory tract infections in children. Indian Journal of Pathology & Microbiology, vol. 59, no. 4, pp. 499-503. http://dx.doi.org/10.4103/0377-4929.191803. PMid:27721281.
http://dx.doi.org/10.4103/0377-4929.1918...
). As stated before, M. pneumoniae is one of the suspicious agents which may be associated with atherosclerosis alone or in coexistence with other conventional risk factors (Guan et al., 2008GUAN, X.R., JIANG, L.X. and MA, X.H., 2008. [Relationship between mycoplasma pneumoniae infection and acute myocardial infarction]. Zhongguo Wei Zhong Bing Ji Jiu Yi Xue, vol. 20, no. 4, pp. 236-237. [chinese]. PMid:18419961.). In this study, certain lipid risk factors (triglycerides, HDL and lipid tetrad index) were significantly higher (p<0.05) in the CAD patients who were positive for M. pneumoniae infection. The fascinating character of this bacterium is that they require cholesterol for survival because their membrane is constituted of cholesterol, a unique property among prokaryotes. Furthermore, this bacterium tends to be detected mainly in the lipid cores of coronary plaques in patients with MI (Higuchi et al., 2000HIGUCHI, M.L., SAMBIASE, N., PALOMINO, S., GUTIERREZ, P., DEMARCHI, L.M., AIELLO, V.D. and RAMIRES, J.A., 2000. Detection of mycoplasma pneumoniae and chlamydia pneumoniae in ruptured atherosclerotic plaques. Brazilian Journal of Biology = Revista Brasileira de Biologia, vol. 33, no. 9, pp. 1023-1026. PMid:10973132.; Higuchi and Ramires, 2002HIGUCHI, M.L. and RAMIRES, J.A., 2002. Infectious agents in coronary atheromas: a possible role in the pathogenesis of plaque rupture and acute myocardial infarction. Revista do Instituto de Medicina Tropical de São Paulo, vol. 44, no. 4, pp. 217-224. http://dx.doi.org/10.1590/S0036-46652002000400007. PMid:12219114.
http://dx.doi.org/10.1590/S0036-46652002...
). A study by Momiyama et al., hypothesized that the combination of M. pneumoniae infection and hyperlipidemia may be an important cofactor for CAD. Though the above study concluded that they could not find any interaction between M. pneumoniae infection and hyperlipidemia for CAD, in our study the CAD patients with M. pneumoniae infection had hyperlipidemia (Momiyama et al., 2004MOMIYAMA, Y., OHMORI, R., TANIGUCHI, H., NAKAMURA, H. and OHSUZU, F., 2004. Association of mycoplasma pneumoniae infection with coronary artery disease and its interaction with chlamydial infection. Atherosclerosis, vol. 176, no. 1, pp. 139-144. http://dx.doi.org/10.1016/j.atherosclerosis.2004.04.019. PMid:15306186.
http://dx.doi.org/10.1016/j.atherosclero...
). The non-inclusion of a parallel control group is one of the limitations, obtaining a tissue sample for non-atherosclerotic patients is practically difficult due to ethical considerations. Hence we could not include a parallel control group to compare the results obtained in this study. Secondly, this is a bi-center study and one should be cautious in interpreting this data to the general population.

5. Conclusion

Our study reported an unusually higher prevalence of M. pneumoniae by serological tests (36.1%) and PCR (43.3%). Although the hypothesis of the association of M. pneumoniae and CAD is yet to be proven, the unusually high prevalence of M. pneumoniae in CAD patients indicates an association, if not, in the development of atherosclerosis. Hence larger studies establishing the relationship of M. pneumoniae in the development of atherosclerosis are required.

References

  • ALVIAR, C.L., ECHEVERRI, J.G., JARAMILLO, N.I., FIGUEROA, C.J., CORDOVA, J.P., KORNIYENKO, A., SUH, J. and PANIZ-MONDOLFI, A., 2011. Infectious atherosclerosis: is the hypothesis still alive? A clinically based approach to the dilemma. Medical Hypotheses, vol. 76, no. 4, pp. 517-521. http://dx.doi.org/10.1016/j.mehy.2010.12.006 PMid:21216537.
    » http://dx.doi.org/10.1016/j.mehy.2010.12.006
  • BAYRAM, A., ERDOGAN, M.B., EKSI, F., and YAMAK, B., 2011. Demonstration of chlamydophila pneumoniae, mycoplasma pneumoniae, cytomegalovirus, and epstein-barr virus in atherosclerotic coronary arteries, nonrheumatic calcific aortic and rheumatic stenotic mitral valves by polymerase chain reaction. Anadolu kardiyoloji dergisi : AKD = the Anatolian Journal of Cardiology, vol. 11, no. 3, pp. 237-243. http://dx.doi.org/10.5152/akd.2011.057
    » http://dx.doi.org/10.5152/akd.2011.057
  • CHAUDHRY, R., SHARMA, S., JAVED, S., PASSI, K., DEY, A.B. and MALHOTRA, P., 2013. Molecular detection of mycoplasma pneumoniae by quantitative real-time PCR in patients with community acquired pneumonia. The Indian Journal of Medical Research, vol. 138, pp. 244-251. PMid:24056602.
  • CORRALES-MEDINA, V.F., MADJID, M. and MUSHER, D.M., 2010. Role of acute infection in triggering acute coronary syndromes. The Lancet. Infectious Diseases, vol. 10, no. 2, pp. 83-92. http://dx.doi.org/10.1016/S1473-3099(09)70331-7 PMid:20113977.
    » http://dx.doi.org/10.1016/S1473-3099(09)70331-7
  • DANESH, J., COLLINS, R. and PETO, R., 1997. Chronic infections and coronary heart disease: is there a link? Lancet, vol. 350, no. 9075, pp. 430-436. http://dx.doi.org/10.1016/S0140-6736(97)03079-1 PMid:9259669.
    » http://dx.doi.org/10.1016/S0140-6736(97)03079-1
  • DAXBÖCK, F., ASSADIAN, A., WATKINS-RIEDEL, T. and ASSADIAN, O., 2011. Persistently elevated iga antibodies to mycoplasma pneumoniae in patients with internal carotid artery stenosis. GMS Krankenhaushygiene Interdisziplinär, vol. 6, no. 1, pp. Doc04. PMid:22242085.
  • DAXBOECK, F., KIRCHER, K., KRAUSE, R., HEINZL, H., WENISCH, C. and STANEK, G., 2002. Effect of age on antibody titer to mycoplasma pneumoniae. Scandinavian Journal of Infectious Diseases, vol. 34, no. 8, pp. 577-579. http://dx.doi.org/10.1080/00365540110089836 PMid:12238572.
    » http://dx.doi.org/10.1080/00365540110089836
  • FABRICANT, C.G., FABRICANT, J., LITRENTA, M.M. and MINICK, C.R., 1978. Virus-induced atherosclerosis. The Journal of Experimental Medicine, vol. 148, no. 1, pp. 335-340. http://dx.doi.org/10.1084/jem.148.1.335 PMid:209124.
    » http://dx.doi.org/10.1084/jem.148.1.335
  • GÓIS, J., HIGUCHI, M., REIS, M., DIAMENT, J., SOUSA, J., RAMIRES, J. and OLIVEIRA, S., 2006. Infectious agents, inflammation, and growth factors: how do they interact in the progression or stabilization of mild human atherosclerotic lesions? Annals of Vascular Surgery, vol. 20, no. 5, pp. 638-645. http://dx.doi.org/10.1007/S10016-006-9076-1 PMid:16983590.
    » http://dx.doi.org/10.1007/S10016-006-9076-1
  • GUAN, X.R., JIANG, L.X. and MA, X.H., 2008. [Relationship between mycoplasma pneumoniae infection and acute myocardial infarction]. Zhongguo Wei Zhong Bing Ji Jiu Yi Xue, vol. 20, no. 4, pp. 236-237. [chinese]. PMid:18419961.
  • HAUER, A.D., DE VOS, P., PETERSE, N., TEN CATE, H., VAN BERKEL, T.J., STASSEN, F.R. and KUIPER, J., 2006. Delivery of chlamydia pneumoniae to the vessel wall aggravates atherosclerosis in LDLr-/- mice. Cardiovascular Research, vol. 69, no. 1, pp. 280-288. http://dx.doi.org/10.1016/j.cardiores.2005.07.011 PMid:16112098.
    » http://dx.doi.org/10.1016/j.cardiores.2005.07.011
  • HAUKSDÓTTIR, G.S., JÓNSSON, T., SIGURDARDÓTTIR, V. and LÖVE, A., 1998. Seroepidemiology of mycoplasma pneumoniae infections in iceland 1987-96. Scandinavian Journal of Infectious Diseases, vol. 30, no. 2, pp. 177-180. http://dx.doi.org/10.1080/003655498750003591 PMid:9730307.
    » http://dx.doi.org/10.1080/003655498750003591
  • HIGUCHI, M.L. and RAMIRES, J.A., 2002. Infectious agents in coronary atheromas: a possible role in the pathogenesis of plaque rupture and acute myocardial infarction. Revista do Instituto de Medicina Tropical de São Paulo, vol. 44, no. 4, pp. 217-224. http://dx.doi.org/10.1590/S0036-46652002000400007 PMid:12219114.
    » http://dx.doi.org/10.1590/S0036-46652002000400007
  • HIGUCHI, M.L., REIS, M.M., SAMBIASE, N.V., PALOMINO, S.A., CASTELLI, J.B., GUTIERREZ, P.S., AIELLO, V.D., RAMIRES, J.A., 2003. Coinfection with Mycoplasma Pneumoniae and Chlamydia Pneumoniae in ruptured plaques associated with acute myocardial infarction. Arquivos Brasileiros de Cardiologia, vol. 81, no. 1, pp. 12-22.
  • HIGUCHI, M.L., SAMBIASE, N., PALOMINO, S., GUTIERREZ, P., DEMARCHI, L.M., AIELLO, V.D. and RAMIRES, J.A., 2000. Detection of mycoplasma pneumoniae and chlamydia pneumoniae in ruptured atherosclerotic plaques. Brazilian Journal of Biology = Revista Brasileira de Biologia, vol. 33, no. 9, pp. 1023-1026. PMid:10973132.
  • KUMAR, S., SAIGAL, S.R., SETHI, G.R. and KUMAR, S., 2016. Application of serology and nested polymerase chain reaction for identifying chlamydophila pneumoniae in community-acquired lower respiratory tract infections in children. Indian Journal of Pathology & Microbiology, vol. 59, no. 4, pp. 499-503. http://dx.doi.org/10.4103/0377-4929.191803 PMid:27721281.
    » http://dx.doi.org/10.4103/0377-4929.191803
  • MARAHA, B., VAN DER ZEE, A., BERGMANS, A.M., PAN, M., PEETERS, M.F., BERG, H.F., SCHEFFER, G.J. and KLUYTMANS, J.A., 2000. Is mycoplasma pneumoniae associated with vascular disease? Journal of Clinical Microbiology, vol. 38, no. 2, pp. 935-936. http://dx.doi.org/10.1128/JCM.38.2.935-936.2000 PMid:10655422.
    » http://dx.doi.org/10.1128/JCM.38.2.935-936.2000
  • MOMIYAMA, Y., OHMORI, R., TANIGUCHI, H., NAKAMURA, H. and OHSUZU, F., 2004. Association of mycoplasma pneumoniae infection with coronary artery disease and its interaction with chlamydial infection. Atherosclerosis, vol. 176, no. 1, pp. 139-144. http://dx.doi.org/10.1016/j.atherosclerosis.2004.04.019 PMid:15306186.
    » http://dx.doi.org/10.1016/j.atherosclerosis.2004.04.019
  • NGEH, J., GUPTA, S., GOODBOURN, C. and MCELLIGOTT, G., 2004. Mycoplasma pneumoniae in elderly patients with stroke. A case-control study on the seroprevalence of M. Pneumoniae in elderly patients with acute cerebrovascular disease - the m-peps study. Cerebrovascular Diseases (Basel, Switzerland), vol. 17, no. 4, pp. 314-319. http://dx.doi.org/10.1159/000077342 PMid:15026614.
    » http://dx.doi.org/10.1159/000077342
  • OHAYON, J., GHARIB, A.M., GARCIA, A., HEROUX, J., YAZDANI, S.K., MALVE, M., TRACQUI, P., MARTINEZ, M.A., DOBLARE, M., FINET, G. and PETTIGREW, R.I., 2011. Is arterial wall-strain stiffening an additional process responsible for atherosclerosis in coronary bifurcations?: An in vivo study based on dynamic CT and MRI. American Journal of Physiology. Heart and Circulatory Physiology, vol. 301, no. 3, pp. H1097-H1106. http://dx.doi.org/10.1152/ajpheart.01120.2010 PMid:21685261.
    » http://dx.doi.org/10.1152/ajpheart.01120.2010
  • RASTAWICKI, W., KALUZEWSKI, S. and JAGIELSKI, M., 1998. Occurrence of serologically verified mycoplasma pneumoniae infections in poland in 1970-1995. European Journal of Epidemiology, vol. 14, no. 1, pp. 37-40. http://dx.doi.org/10.1023/A:1007431932087 PMid:9517871.
    » http://dx.doi.org/10.1023/A:1007431932087
  • RAZIN, S., YOGEV, D. and NAOT, Y., 1998. Molecular biology and pathogenicity of mycoplasmas. Microbiology and Molecular Biology Reviews, vol. 62, no. 4, pp. 1094-1156. http://dx.doi.org/10.1128/MMBR.62.4.1094-1156.1998 PMid:9841667.
    » http://dx.doi.org/10.1128/MMBR.62.4.1094-1156.1998
  • REUNANEN, A., ROIVAINEN, M. and KLEEMOLA, M., 2005. Increased titer of antibodies to mycoplasma pneumoniae may be associated with coronary heart disease. Atherosclerosis, vol. 180, no. 1, pp. 209-210. http://dx.doi.org/10.1016/j.atherosclerosis.2004.12.032 PMid:15823295.
    » http://dx.doi.org/10.1016/j.atherosclerosis.2004.12.032
  • ROHAM, M., ANBARI, K., MIRHABIBI, S. and GOUDARZI, G., 2016. The seroprevalence of mycoplasma pneumoniae IgM and IgG antibodies in patients with ischemic stroke. Iranian Journal of Microbiology, vol. 8, no. 6, pp. 383-388. PMid:28491249.
  • ROSS, R., 1999. Atherosclerosis--an inflammatory disease. The New England Journal of Medicine, vol. 340, no. 2, pp. 115-126. http://dx.doi.org/10.1056/NEJM199901143400207 PMid:9887164.
    » http://dx.doi.org/10.1056/NEJM199901143400207
  • TAYLOR-ROBINSON, D. and THOMAS, B.J., 1998. Chlamydia pneumoniae in arteries: the facts, their interpretation, and future studies. Journal of Clinical Pathology, vol. 51, no. 11, pp. 793-797. http://dx.doi.org/10.1136/jcp.51.11.793 PMid:10193317.
    » http://dx.doi.org/10.1136/jcp.51.11.793
  • TUUMINEN, T., VARJO, S., INGMAN, H., WEBER, T., OKSI, J. and VILJANEN, M., 2000. Prevalence of chlamydia pneumoniae and mycoplasma pneumoniae immunoglobulin g and a antibodies in a healthy finnish population as analyzed by quantitative enzyme immunoassays. Clinical and Diagnostic Laboratory Immunology, vol. 7, no. 5, pp. 734-738. http://dx.doi.org/10.1128/CDLI.7.5.734-738.2000 PMid:10973446.
    » http://dx.doi.org/10.1128/CDLI.7.5.734-738.2000
  • WATKINS-RIEDEL, T., STANEK, G. and DAXBOECK, F., 2001. Comparison of seromp iga with four other commercial assays for serodiagnosis of mycoplasma pneumoniae pneumonia. Diagnostic Microbiology and Infectious Disease, vol. 40, no. 1-2, pp. 21-25. http://dx.doi.org/10.1016/S0732-8893(01)00250-4 PMid:11448559.
    » http://dx.doi.org/10.1016/S0732-8893(01)00250-4
  • WELTI, M., JATON, K., ALTWEGG, M., SAHLI, R., WENGER, A. and BILLE, J., 2003. Development of a multiplex real-time quantitative pcr assay to detect chlamydia pneumoniae, legionella pneumophila and mycoplasma pneumoniae in respiratory tract secretions. Diagnostic Microbiology and Infectious Disease, vol. 45, no. 2, pp. 85-95. http://dx.doi.org/10.1016/S0732-8893(02)00484-4 PMid:12614979.
    » http://dx.doi.org/10.1016/S0732-8893(02)00484-4
  • YIALLOUROS, P., MOUSTAKI, M., VOUTSIOTI, A., SHARIFI, F. and KARPATHIOS, T., 2013. Association of mycoplasma pneumoniae infection with henoch-schonlein purpura. Prague Medical Report, vol. 114, no. 3, pp. 177-179. http://dx.doi.org/10.14712/23362936.2014.20 PMid:24093818.
    » http://dx.doi.org/10.14712/23362936.2014.20

Publication Dates

  • Publication in this collection
    10 Sept 2021
  • Date of issue
    2023

History

  • Received
    09 Dec 2020
  • Accepted
    16 Apr 2021
Instituto Internacional de Ecologia R. Bento Carlos, 750, 13560-660 São Carlos SP - Brasil, Tel. e Fax: (55 16) 3362-5400 - São Carlos - SP - Brazil
E-mail: bjb@bjb.com.br