Microbial signature present in thrombotic material of acute ischemic stroke patients retrieved by mechanical thrombectomy

doi:10.21203/rs.3.rs-1715062/v1

This study is done to examine the microbiota in thrombi retrieved from the middle cerebral artery after mechanical thrombectomy for symptomatic carotid plaque within 6 hours of an acute ischemic stroke. To evaluate their role in atherosclerosis, thrombi were submitted to next-generation sequencing for a bacterial signature. In these study we are describing allots of bacteria found in thrombus samples, which are associated with stroke. Enrollment of 14 Acute ischemic stroke (AIS) patients was done who underwent mechanical thrombectomy in which 72% (n = 10) male and 28% (n = 4) female. The patients' median age was 16.75 ± 3.68 year old, and they were all female. During the window of acute stroke with symptomatic carotid stenosis and blockage of the ipsilateral middle cerebral artery, all were submitted to mechanical thrombectomy. After mechanical thrombectomy we extracted the DNA from thrombus sample. The microbial composition of all samples was compared using 16S rRNA gene amplicon next-generation sequencing.All of the thrombi retrieved for bacterial DNA in qPCR from the fourteen patients who had thrombectomy for ischemic stroke were positive. The 14 thrombus samples included more than 30 microorganisms. large bacterial diversity was found in thrombus in ischemic stroke patients in this study, which also indicated that microbes present in thrombus also played role in formation of plaque and Bacteria have direct mechanisms such as acidification and local inflammation of the plaque milieu with lactobacillus, biofilm dispersion leading to inflammation with pseudomonas fluorescence, or enterococci, or indirect mechanisms such as Toll 2 like signalling by the gut microbiota, could all lead to thromboembolic and cause stroke. It justifies more research into the involvement of bacteria in thrombus formation in stroke pathophysiology and prognosis.

Stroke

Mechanical Thrombectomy

Thrombus

Microbiome

Metagenomics

Stroke, as the second leading cause of death and disability, has established causes of disparities and changes in trends in stroke burden across countries (1). In 2008, the yearly economic burden of informal care giving linked with stroke among the elderly in the United States was projected to be $14.2 billion (2). As previously reported, atherosclerotic plaque may have an infectious aetiology (3).The majority of previous stroke research focused on extracranial carotid atherosclerotic plaque on carotid endarterectomy patients, for example, detecting Helicobacter pylori in human carotid atherosclerotic plaques (4).

The gut microbiota appears to play a role in the aetiology and outcome of stroke, according to an increasing body of research. After an ischemic stroke, the composition of the gut flora changes. The gut microbiota, on the other hand, can affect stroke outcome and play a role in stroke causation. The gut microbiota has been associated to stroke risk factors such as hypertension, diabetes, and obesity in clinical and experimental investigations (5).

Infectious agents have been linked to persistent inflammation, which plays a role in atherosclerosis etiopathogenesis (6). Since the 1970s, researchers have been looking into the relationship between infection and atherosclerosis.

Pathogens have been linked to atherosclerosis in several studies, including (a) pathogens found in human atherosclerotic vessels (8), (b) infection linked to atherosclerosis (9) in animal studies, and (c) seroepidemiological studies showing an association between pathogen-specific antibodies and atherosclerosis (10).

Multiple pathogens, including bacteria such as Chlamydia pneumoniae, Helicobacter pylori, and periodontal pathogens, have been implicated in these associations (6).

Chlamydia pneumoniae is the pathogen whose association with atherosclerosis has received the most attention, with varying results. Chlamydia pneumoniae was the first infectious organism discovered in human atherosclerotic plaque cells, but only rarely in normal arterial cells. It is in addition one of the few agents found in plaques that have yielded viable organisms (3, 11).

2.1 Subjects

A total of 14 patients with acute ischemic stroke, middle cerebral occlusion, and symptomatic carotid stenosis were admitted to Pacific Medical University & Hospital's Department of Neurology between February 2019 and March 2022.

Acute ischemic stroke with symptomatic carotid stenosis and occlusion of the middle cerebral artery, as diagnosed by neurologists, was the main criterion for inclusion. To assess ischemia lesions, all patients had a brain magnetic resonance imaging (MRI) or computed tomography (CT) scan within a 6-hour timeframe. They all had symptomatic carotid stenosis plaque, as well as a thromboembolic event that culminated in a middle cerebral artery blockage. Magnetic resonance angiograms of the neck vessel and circle of Willis vessels were conducted in addition to normal magnetic resonance imaging of the brain.

All of these carotid plaques were confirmed to be fragile or unstable plaques by morphological inspection using carotid Doppler and MRI plaque imaging. The following were among the excluding criteria: prior intravenous thrombolysis treatment; cardio embolic stroke; dissection-associated stroke; patent foramen ovale; vasculitis; history of malignancy, autoimmune disease, chronic hemodialysis kidney disease, parkinson's disease; prior intravenous thrombolysis treatment; cardio embolic stroke; hypercoagulant or genetic underlying cause of stroke; faecal thrombosis. A total of fourteen patients were enrolled in the study cohort. The National Institutes of Health Stroke Scale (NIHSS) admission rate was 16.75 ± 3.68 percent (range, 11 to 22). This study was authorized according to ethical standards by the Pacific Medical University & Hospital Regional Review Board in Rajasthan, India. The use of human participants in this study has been approved by the regional review board. All of the participants had given their informed consent in writing.

2.2 Mechanical Thrombectomy and Thrombus Sample Collection

All procedures were carried out in a biplane angiography cath lab (Phillips, Allura xper FD 20 − 15). A conventional adapted seldinger approach was used to place an introducer sheath on a femoral artery, and a guiding catheter with a tip balloon up to 9F was navigated into the carotid artery proximal to the occluded location. The stent retriever was inserted using a micro catheter with a guide wire (0.021 inches) that was manoeuvred through the blocked location and over the thrombus (solitaire; Medtronic). Multiple retrieval runs were possible with Solitaire revascularization devices, with up to three retrievals in the same vessel and two retrievals per unit. The stent retrieval device was deployed by pulling the micro catheter back at the desired point after it was positioned across the visible clot. The Solitaire resheathing was done as needed, but not more than thrice. The thrombus was removed using negative aspiration using a 20 cc Syringe. The final angiography was performed to determine whether or not the flow had been restored (TICI flow). Thrombectomy was repeated until the angiographic outcome was satisfactory. The thrombus was split into a 1.5 mL eppendorf micro centrifugal tube for DNA extraction and qPCR analysis.

2.3 DNA extraction

The DNA Extraction QIAamp DNA Mini Kit (Qiagen, Hilden, Germany) was used to extract DNA from frozen thrombotic material, with minor modifications to the protocol. The technique of mechanical tissue disaggregation was developed expressly for preparing thrombotic material for DNA extraction. A NanoDrop spectrophotometer was used to determine the purity and concentration of DNA. For qPCR analysis, the minimal quality (ng/L) and purity (ratio 260/280) values were 5 and 1.5, respectively.

2.4 16S sequencing (metagenomics analysis)

For the sequencing, 16S Amplicon-Seq hypervariable regions V1–V9 were amplified using primers, including the forward 16S primer 5’-ATCGCCTACCGTGAC-barcode-AGAGTTTGATCMTGGCTCAG-3’ and Reverse 16S primer 5’-ATCGCCTACCGTGAC-barcode-CGGTTACCTTGTTACGACTT-3’ with gene-specific sequences, MinION, Oxford Nanopore, and molecular barcodes. The PCR mixture for the full-length 16S rRNA gene (50 µL total volume) contained 10 ng of DNA template (10 µL), 25 µL long Amp Taq 2X master mix (NEB M0287), 1 µL 16S barcode (Barcode 01 to Barcode 04 each for a single sample separately), and 14 µL nuclease-free water as indicated in the 16S barcoding kit (SQK-RAB204). The thermal PCR profile consisted of an initial denaturation of 60 seconds at 95°C followed by 25 cycles of 20 seconds at 95°C, 30 seconds at 55°C, 2 minutes at 65°C, and a final stage of 5 minutes at 65°C. The amplicons were washed with the Beckman Coulter (AMPure XP beads) using a ratio of 0.5X. Usage of the Nanodrop spectrophotometer was evaluated for each sample quantity. The various bar-coded samples were pooled in equimolar ratio to get a final pool (100–150 ng in 10 µL) to do the sequencing library by adding 1 µL of RAP (Rapid Annealing Primer) to the bar-coded DNA. The prepared sample (11 µL DNA sample) was combined with Library Loading Beads (25.5 µL) and Fuel Mix Running Buffer (35.5 µL) and loaded onto SpotON Flow Cells Mk I (R9.4.1) (FLO-MIN106) for 12 hours using MinKNOWTM 19.06.8.

2.5 Data availability

Data that support the findings of this study are available upon reasonable request from the Pacific Medical University study investigators.

3.1 Patient characteristics:

We recruited 14 AIS patients who underwent mechanical thrombectomy 72% (n = 10) male and 28% (n = 4) female. The median age for the patients was 52 ± 13.6 years. All were subjected to mechanical thrombectomy during the window time of acute stroke with symptomatic carotid stenosis, with occlusion of the ipsilateral middle cerebral artery. The median delay time between the onset of ischemic stroke and hospital arrival was 6 hours.

Seven patients had a history of hypertension and 3 had diabetes mellitus. Before mechanical thrombectomy and thrombus sample collection, no patient had a history of serious infection, septicemia, or recent antibiotic treatment

3.2 Presence of bacterial DNA microbial features in thrombus aspirates;

Thrombus samples were collected from all participants who are going to mechanicalthrombectomy for acute stroke. Notably 100%( 14) of aspirated thrombi were positive for bacterial DNA in qPCR. More than 30 bacteria were present in 14 thrombus samples. The majority of bacteria were Staphylococcus aureus lactobacillus heliveticus, Prevotella amni, Prevotella clorans, Lactobacillus jensenni, Lactobacillus gasseri, Lactobacillus crispatus, Lactobacillus indneri, Lactobacillus, Staphylococcus, and Stenotrophomonas in all 14 thrombi. Other abundant bacteria were Staphylococcus aureus, and Bacillus cereus, Bacillus subtilis, Streptococcus pneumonia, Streptococcus pyogenes, streptococcus thermophilus and Planococcus sps. Enterococci faececium, as well as many others shown in figure. In reality, the majority of bacterial species found in abundance in (figure − 1) could be commensal or low-grade pathogens that live in close proximity to the human body. During temporary bacteremia without overt sepsis, their entry into the bloodstream from the mouth cavity, gut, or skin may cause them to become trapped in carotid plaque.

Our research found that bacterial DNA, 16 rRNA, was present in thrombi samples taken from stroke patients.

To the best of our knowledge, this is a large-cohort prospective study in which the thrombus features of acute ischemic stroke patients were fully characterized using 16S Sequencing technology, and it investigates the bacteria that are related to stroke.

There are, majority of bacteria found in the thrombus, the higher abundance of bacteria Stenotrophomonas maltophilia, Peptostreptococcus anaerobius Acinetobacter schindleri(5.77%) and Acinetobacter indicus (5.77%) while lactobacillus crispatus (20%), Lactobacillus gasseri (8%), and Lactobacillus iners (6.4%). It's possible that the majority of bacterial species in thrombus are commensal or low-grade pathogens found in the human body, which can enter the bloodstream via the mouth, gut, or skin and cause atherosclerotic plaque. Our findings reveal that next-generation sequencing can predict the relative abundance of bacteria.

Bacterial binding to platelets causes aggregation, as demonstrated by Streptococcus sanguinis, Staphylococcus epidermidis, and Chlamydia pneumoniae (12). Many bacteria, such as the gram-positive bacteria Staphylococcus aureus, can activate the blood clotting system and alter specific coagulation factors, however they do not cause the coagulation cascade to begin.

These bacteria use the coagulation system to shield themselves from being targeted by the host's immune system (13). Extracellular matrix (ECM) proteins such as fibrinogen, fibronectin, and collagen, for example, help Staphylococcus aureus form clumps (14).

Platelet aggregation development aids in the colonization of host tissues (15). Staphylococcus aureus is one of the germs that can use blood clotting to their advantage (13). Platelets, on the other hand, work along with neutrophils to create microbial traps, but this increases the risk of thrombosis (12).

Bacterial adherence to the surface of the midblock plays a key function in clot formation. Exotoxins released by Staphylococcus aureus bacteria interact with cell membranes, causing platelet activation and aggregation as well as smooth muscle contraction. Both of these causes have the potential to cause thrombosis.

As a result of the coagulase enzyme interacting with fibrinogen, plasma coagulation occurs. The most prevalent pathogen associated in venous thrombosis with osteomyelitis is Methicillin-resistant Staphylococcus aureus (MRSA). Deep vein thrombosis in children has been linked to generalized soft tissue infection, septic arthritis, osteomyelitis, and myositis (16).

MRSA infections caused higher and more severe vascular consequences than methicillin-resistant Staphylococcus aureus (MSSA) infections, especially in individuals with lung involvement. MRSA infections were also associated with higher levels of inflammation and needed longer hospital stays (17).

Many common bacteria cause infections and raise the risk of thrombotic consequences such as ischemic stroke and myocardial infarction (18, 19). Microbes also trigger an inflammatory response that involves the fibrinolytic system (20). The bacterial infection causes significant alterations in the coagulation and fibrinolysis balance (21)

Thrombosis might occur as a result of an infection that was not properly managed. However, the processes of infection-induced thrombosis, as well as the mechanisms of infection control by the host in pathogen transmission, remain poorly known.

Pathogen-induced thrombosis is related with the production of inflammatory chemicals that activate platelets, as well as damage to the endothelium, which leads to fibrin deposition and thrombus formation (22).

During the research, it was discovered that bacteria or bacterial lipopolysaccharide play a role in platelet activation and thrombus formation. Platelets can interact with gram-negative and gram-positive bacteria via direct binding via platelet membrane surface receptors and bacterial surface protein (23).

Surprisingly, changes in the distribution of bacteria patterns in thrombus from various origins were discovered. The majority of bacteria in cardioebolic stroke patients are clustered or found on the surface of red blood cells, while bacteria in LLA stroke patients are clustered or found on the surface of red blood cells (24).

However, a study found that even in the absence of atherosclerotic plaque, the presence of bacteria promotes thrombogenesis. When other bacteria congregate in a cluster, the bacillus' unique spatial localization may alter the coagulation process, which is immediately activated in blood within minutes (25).

Furthermore, some bacteria species, such as Staphylococcus aureus, release protocogulant factor, which promotes plasma coagulation to prevent lethal immune system attacks (26).

Our study showed that there are so many bacteria like bacillus and Staphylococcus aureus Pseudomonas putida, Staphylococcus epidermidis, Staphylococcus hominis, and Finegoldia magna Etc present in thrombus aspirated of stroke patients and their relative abundance that is. 3% respectively. The ability of some gram negative to build biofilms in other body locations was greatly improved (24).The Bactria in the thrombus or clot with acute ischemic stroke may play vital role in promoting blood coagulation.

Early adverse events (haemorrhagic transfusion, etc.) after mechanical thrombectomy are associated with a poor prognosis in stroke patients. The level of opportunistic pathogens such as acenatobacter and enterobacteriacy was significantly higher in patients with adverse events within 48 hours of admission than in those without.

Acinetobacter species are ubiquitous, widely distributed organisms that contribute to a variety of nosocomial illnesses (27, 28). Enterobacteriaceae enrichment in the gut is linked to an increased risk of stroke (29) and is an independent predictor of poor outcomes in stroke patients (30).

Surprisingly, when the amount of nutrients available is rapidly reduced, bacteria biofilm can be employed to respond to the resulting dispersion (31). Bacteria that form biofilms, such as Pseudomonas fluorescens and enterococcus, have been shown to raise factor VIII levels, are nonpathogenic motile, or are low-grade pathogens that increase the risk of stroke (32).

High norepinephrine enhances biofilm dispersion events, especially in reaction to stress, when a critical ingredient is liberated from its bound condition, such as free iron generation (31).

Sternotrophomonas Maltophilia is a low-virulent bacterium that forms biofilms (33). The StmPr1 gene codes for a S. maltophilia protease that can break down collagen, fibronectin, and fibrinogen protein components, potentially causing local tissue injury and bleeding (34) In periodontal disease, F. nucleatum is linked to the development of Lemierre's syndrome, a pulmonary infection accompanied by septic thrombophelebitis of the inner and outer jugular veins, arthritis, and aneurysms (35–37).

Furthermore, a larger abundance of Acinetobacter was linked to a higher 3-month mortality rate in stroke patients, according to our findings. In addition to three major clinical indications, univariate and multivariate Cox regression analysis revealed that the quantity of Acinetobacter in the thrombus was an important risk factor for 90-day death. However, the exact mechanism by which Acinetobacter has a role in poor clinical outcomes in stroke patients is unknown. Targeting the thrombus microbiota for precise regulation could help prevent thrombosis, reduce the risk of stroke, and improve the clinical outcome of stroke patients.

The study showed evidence for significant of diversity of bacteria in thrombus in ischemic stroke patients. Our study indicated that microbes present in thrombus which is formation of plaque as well as associated with stroke .It is among the first fully characterize the thrombus microbial feature observe which is closely associated with pathogens and risk of death receiving mechanical thrombectomy. It is further investigation on the role of bacteria in thrombus in the pathogenesis and prognosis of stroke

Ethical Approval and Consent to participate

Not applicable.

Consent for publication

Not applicable.

Availability of data and materials

Not applicable.

Competing interests

The authors declare that they have no competing interests.

Funding

The study has been funded by the Department of Science and Technology Govt. of Rajasthan India.

Authors' contributions

LBY, and AV, ST designed and wrote the manuscript. ST, LBY, and MV, revised the article. All authors have read and approved the manuscript. LBY, AV,, ST, and, contributed equally.

Acknowledgement

The Pacific Medical University and Hospital provided the authors with facilities and resources during the research. The writers gratefully acknowledge financial support from the Rajasthan Government's Department of Science and Technology.

Feigin VL, Lawes CM, Bennett DA, Barker‑Collo SL, Parag V (2009) Worldwide stroke incidence and early case fatality reported in 56 population‑based studies: A systematic review. Lancet Neurol;8:355‑69.
Joo H, Dunet DO, Fang J, Wang G (2014) Cost of informal care giving associated with stroke among the elderly in the United States. Neurology. Nov 11;83(20):1831
Calandrini CA, Ribeiro AC, Gonnelli AC, Ota‐Tsuzuki C, Rangel LP, Saba‐Chujfi E, Mayer MP(2014). Microbial composition of atherosclerotic plaques. Oral diseases. Apr;20(3):e128-34.
Dempsey RJ, Vemuganti R, Varghese T, Hermann BP(2010). A review of carotid atherosclerosis and vascular cognitive decline: a new understanding of the keys to symptomology. Neurosurgery. Aug 1;67(2):484-94.
Winek K, Dirnagl U, Meisel A(2017) Role of the gut microbiota in ischemic stroke. Neurol Int Open.;1:E287–E293.
Epstein SE, Zhu J, Najafi AH, Burnett MS (2009) Insights into the role of infection in atherogenesis and in plaque rupture. Circulation. Jun 23;119(24):3133-41.
Leinonen M, Saikku P(2002)Evidence for infectious agents in cardiovascular disease and atherosclerosis. The Lancet infectious diseases. Jan 1;2(1):11-7.
Ramirez JA(1996) Isolation of Chlamydia pneumoniae from the coronary artery of a patient with coronary atherosclerosis. Annals of internal medicine. Dec 15; 125(12):979-82.
Muhlestein JB, Anderson JL, Hammond EH, Zhao L, Trehan S, Schwobe EP, Carlquist JF(1998) Infection with Chlamydia pneumoniae accelerates the development of atherosclerosis and treatment with azithromycin prevents it in a rabbit model. Circulation. Feb 24;97(7):633-6.
Saikku PM, Mattila K, Nieminen MS, Huttunen JK, Leinonen M, Ekman MR, Mäkelä PH, Valtonen V(1988). Serological evidence of an association of a novel Chlamydia, TWAR, with chronic coronary heart disease and acute myocardial infarction. The Lancet. Oct 29;332(8618):983-6.
LaBiche R, Koziol D, Quinn TC, Gaydos C, Azhar S, Ketron G, Sood S, DeGraba TJ(2001) Presence of Chlamydia pneumoniae in human symptomatic and asymptomatic carotid atherosclerotic plaque. Stroke. Apr;32(4):855-60.
Hamzeh-Cognasse H, Damien P, Chabert A, Pozzetto B, Cognasse F, Garraud O(2015). Platelets and infections - Complex interactions with bacteria. Front Immunol.; 6:1–18. doi:10.3389/fimmu.2015.00082.
McAdow M, Missiakas DM, Schneewind O (2012)Staphylococcus aureus secretes coagulase and von willebrand factor binding protein to modify the coagulation cascade and establish host infections. J Innate Immun.;4(2):141–148. doi:10.1159/000333447.
Crosby HA, Kwiecinski J, Horswill AR(2016) Staphylococcus aureus aggregation and coagulation mechanisms, and their function in host–pathogen interactions In: Advances in Applied Microbiology;:1–41. doi:10.1016/bs.aambs.2016.07.018.
Thomer L, Schneewind O, Missiakas D(2016) Pathogenesis of Staphylococcus aureus bloodstream infections. Annu Rev Pathol.;11(1):343–364. doi:10.1146/annurev-pathol-012615-044351.
Crary SE, Buchanan GR, Drake CE, Journeycake JM. (2006) venous thrombosis and thromboembolic in children with osteomyelitis. J Pediatr.;149(4):537–541. doi:10.1016/j.jpeds.2006.06.067.
Gonzalez BE, Teruya J, Mahoney DH, Hulten KG, Edwards R, Lamberth LB, Hammerman WA, Mason EO, Kaplan SL(2006)Venous thrombosis associated with staphylococcal osteomyelitis in children. Pediatrics. May 1;117(5):1673-9.
Lee CY, Lee YS, Tsao PC, Jeng MJ, Soong WJ(2016)Musculoskeletal sepsis associated with deep vein thrombosis in a child. Pediatr Neonatol.;57(3):244–247.
Kastrup CJ, Boedicker JQ, Pomerantsev AP, Moayeri M, Bian Y, Pompano RR, Kline TR, Sylvestre P, Shen F, Leppla SH, Tang WJ.( 2008) Spatial localization of bacteria controls coagulation of human blood by'quorum acting'. Nature chemical biology. Dec;4(12):742-50.
Antoniak S. (2018) the coagulation system in host defense. Res Pract Thromb Haemost.; 2(3):549–557.
Delano MJ, Ward PA (2016) the immune system’s role in sepsis progression, resolution, and long-term outcome. Immunol Rev.; 274(1):330–353.
Beristain-Covarrubias N, Perez-Toledo M, Thomas MR, Henderson IR, Watson SP, Cunningham AF(2019) Understanding infection-induced thrombosis: lessons learned from animal models. Front Immunol.;10.
Violi F, Cangemi R, Calvieri C( 2014) Pneumonia, thrombosis and vascular disease. J Thromb Haemost.;12(9):1391–1400.
Liao Y, Zeng XL, Xie XM, Liang D, Qiao HY, Wang WC, Guan M, Huang SM, Jing Z, Leng XY. (2021) Overexpansion of Proteobacteria in the Thrombus of Stroke Patients Treated with Mechanical Thrombectomy and the Risk for Death.
Kastrup CJ, Boedicker JQ, Pomerantsev AP, Moayeri M, Bian Y, Pompano RR, Kline TR, Sylvestre P, Shen F, Leppla SH, Tang WJ( 2008) Spatial localization of bacteria controls coagulation of human blood by'quorum acting'. Nature chemical biology. Dec;4(12):742-50.
Loof TG, Goldmann O, Naudin C, Mörgelin M, Neumann Y, Pils MC, Foster SJ, Medina E, Herwald H(2015)Staphylococcus aureus-induced clotting of plasma is an immune evasion mechanism for persistence within the fibrin network. Microbiology. Mar 1;161(3):621-7.
Dijkshoorn L, Nemec A, Seifert H(2007) An increasing threat in hospitals: multidrug-resistant Acinetobacterbaumannii. Nat Rev Microbiol.; 5(12):939-51.4.
Gales AC, Jones RN, Forward KR, Linares J, Sader HS, Verhoef J(2001)Emerging importance of multidrug-resistant Acinetobacter species and Stenotrophomonas maltophilia as pathogens in seriously ill patients: geographic patterns, epidemiological features, and trends in the SENTRY Antimicrobial Surveillance Program(1997–1999).Clinical Infectious Diseases. May 15;32(Supplement_2):S104-13.
Zeng X, Gao X, Peng Y, Wu Q, Zhu J, Tan C, Xia G, You C, Xu R, Pan S, Zhou H(2019). Higher risk of stroke is correlated with increased opportunistic pathogen load and reduced levels of butyrate-producing bacteria in the gut. Frontiers in cellular and infection microbiology. Feb 4;9:4.
Xu K, Gao X, Xia G, Chen M, Zeng N, Wang S, You C, Tian X, Di H, Tang W, Li P(2021) Rapid gut dysbiosis induced by stroke exacerbates brain infarction in turn. Gut. Aug 1;70(8):1486-94.
McDougald D, Rice SA, Barraud N, Steinberg PD, Kjelleberg S(2011) Should we stay or should we go: mechanisms and ecological consequences for biofilm dispersal. Nat Rev Microbiol.; 10:39–50.
Shaban A, Hymel B, Chavez-Keatts M, Karlitz JJ, Martin-Schild S (2015)Recurrent posterior strokes in inflammatory bowel disease patients. Gastroenterol Res Pract.; :672460.
Flores-Treviño S, Bocanegra-Ibarias P, Camacho-Ortiz A, Morfín-Otero R, Salazar-Sesatty HA, Garza-González E(2019) Stenotrophomonas maltophilia biofilm: its role in infectious diseases. Expert Rev Anti Infect Ther.;17:877–893
Looney WJ, Narita M, Mühlemann K(2009)Stenotrophomonas maltophilia: an emerging opportunist human pathogen. Lancet Infect Dis.; 9:312–323.
Ford PJ, Gemmell E, Chan A, Carter CL, Walker PJ, Bird PS, West MJ, Cullinan MP, Seymour GJ(2006)Inflammation, heat shock proteins and periodontal pathogens in atherosclerosis: an immunohistologic study. Oral microbiology and immunology. Aug;21(4):206-11.
Pyysalo MJ, Pyysalo LM, Pessi T, Karhunen PJ, Öhman JE(2013). The connection between ruptured cerebral aneurysms and odontogenic bacteria. J Neurol Neurosurg Psychiatry.;84:1214–1218.
Elkaim R, Dahan M, Kocgozlu L, Werner S, Kanter D, Kretz JG, Tenenbaum H(2008). Prevalence of periodontal pathogens in subgingival lesions, atherosclerotic plaques and healthy blood vessels: a preliminary study. Journal of periodontal research. Apr;43(2):224-31.

Table-1

Patient characteristics

PT1

PT 2

PT 3

PT 4

PT 5

PT6

PT 7

PT 8

PT 9

PT10

PT11

PT12

PT13

PT14

Age /Gender

46 M

53M

70M

33M

50F

30F

55F

40M

51M

43F

51M

49M

43M

62M

IV Thrombolysis

YES

NO

YES

NO

YES

NO

NIHSS/ admission

15

13

17

12

10

19

2

11

14

13

18

21

22

19

MRS/ admission

4

2

4

3

2

4

3

2

4

5

6

9

Duration of samples from onset /hour

6

4

3

4

3

5

2

4

5.5

3.6

2

5

6

3

Diabetes

NO

YES

NO

YES

NO

YES

NO

Hypertension

NO

YES

NO

YES

NO

YES

NO

YES

NO

YES

No competing interests reported.

Microbial signature present in thrombotic material of acute ischemic stroke patients retrieved by mechanical thrombectomy

Status:

Version 1

Abstract

Figures

1. Introduction

2. Material Methods

2.1 Subjects

2.2 Mechanical Thrombectomy and Thrombus Sample Collection

2.3 DNA extraction

2.4 16S sequencing (metagenomics analysis)

2.5 Data availability

3. Results

3.1 Patient characteristics:

3.2 Presence of bacterial DNA microbial features in thrombus aspirates;

4. Discussion

Conclusion

Declarations

References

Tables

Additional Declarations

Status:

Version 1