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Background: Acute ischemic stroke may be due to embolism from ruptured atherosclerotic carotid arteries. DNA of oral bacteria, mainly the viridans streptococci group, has been detected in thrombus aspirates of patients with ischemic stroke as well as in carotid endarterectomy samples. Because viridans streptococci are known to possess thrombogenic properties, we studied whether their presence in thrombus aspirates and in carotid artery specimens can be confirmed using bacterial immunohistochemistry., Methods and Results: Thrombus aspirates from 61 patients with ischemic stroke (70.5% men; mean age, 66.8 years) treated with mechanical thrombectomy, as well as carotid endarterectomy samples from 20 symptomatic patients (65.0% men; mean age, 66.2 years) and 48 carotid artery samples from nonstroke autopsy cases (62.5% men; mean age, 66.4 years), were immunostained with an antibody cocktail against 3 species ( Streptococcus sanguinis , Streptococcus mitis, and Streptococcus gordonii ) of viridans streptococci. Of the thrombus aspirates, 84.8% were immunopositive for viridans streptococci group bacteria, as were 80.0% of the carotid endarterectomy samples, whereas immunopositivity was observed in 31.3% of the carotid artery samples from nonstroke autopsies. Most streptococci were detected inside neutrophil granulocytes, but there were also remnants of bacterial biofilm as well as free bacterial infiltrates in some samples., Conclusions: Oral streptococci were found in aspirated thrombi of patients with acute ischemic stroke as well as in carotid artery samples. Our results suggest that viridans streptococci group bacteria may play a role in the pathophysiology of ischemic stroke.

Methods: Thrombus aspirates and control arterial blood were taken from 71 patients (70.4% male; mean age, 67.4 years) with acute ischemic stroke. Tooth pathology was registered using CT scans. Carotid stenosis was estimated with CTA and ultrasonography. The presence of bacterial DNA from aspirated thrombi was determined using quantitative PCR. We also analyzed the presence of these bacterial DNAs in carotid endarterectomies from patients with peripheral arterial disease., Results: Bacterial DNA was found in 59 (83.1%) of the thrombus aspirates (median, 8.6-fold). Oral streptococcal DNA was found in 56 (78.9%) of the thrombus aspirates (median, 5.1-fold). DNA from A. actinomycetemcomitans and P. gingivalis was not found. Most patients suffered from poor oral health and had in median 19.0 teeth left. Paradoxically, patients with better oral health had more oral streptococcal DNA in their thrombus than the group with the worst pathology ( p = 0.028). There was a trend (OR 7.122; p = 0.083) in the association of ≥50% carotid artery stenosis with more severe dental pathology. Oral streptococcal DNA was detected in 2/6 of carotid endarterectomies., Conclusions: Stroke patients had poor oral health which tended to associate with their carotid artery stenosis. Although oral streptococcal DNA was found in thrombus aspirates and carotid endarterectomy samples, the amount of oral streptococcal DNA in thrombus aspirates was the lowest among those with the most severe oral pathology. These results suggest that the association between poor oral health and acute ischemic stroke is linked to carotid artery atherosclerosis., Competing Interests: The authors state no disclosures or conflict of interest., (Copyright © 2021 Olli Patrakka et al.)

Background: The gut microbiome is thought to remain stable into old age. Gut bacteria and their translocation may play a role in the development of coronary heart disease (CHD) by modulating cholesterol levels and immune responses, as well as by producing toxic metabolites and bacterial endotoxins. The association of changes in the gut microbiome with the severity of coronary atherosclerosis and the ability of gut bacteria themselves to translocate into coronary plaques has not been studied., Materials and Methods: As a part of the Tampere Sudden Death Study, we measured age-dependent changes in the relative ratios of major intestinal bacterial communities (Bacteroides species [spp.], the Clostridium leptum group, the Clostridium coccoides group, Bifidobacterium spp., Enterobactericeae, Lactobacillus spp.) and Streptococcus spp. in both feces and coronary plaques of the same male autopsy cases (n = 67, age range 44-95) using real-time quantitative PCR (qPCR). The area of coronary atherosclerotic lesions were measured by computer-assisted morphometry. Fecal bacterial DNA measurements from healthy volunteers served as a control for gut bacterial analyses of autopsy cases. The relative amount of bacterial DNA in a sample was determined with the comparative Cq method., Results: The relative ratios of fecal Lactobacillus spp., Bifidobacterium spp., the Clostridium coccoides group, and Bacteroides spp. did not differ between controls and autopsy cases and showed no age-dependence. In contrast, the ratios of the Clostridium leptum group, Enterobactericeae, and Streptococcus spp. increased with age. Elevated relative ratios of fecal Enterobactericeae associated with a larger coronary plaque fibrotic area (p = 0.001), and the Clostridium leptum group with a larger calcification area (p = 0.015). Intestinal bacterial DNA could be amplified in 67.6% of the coronary plaques, the most common being Streptococcus spp. (41.0%), followed by Enterobactericeae (12.1%), Clostridium leptum (2.4%), and Lactobacillus spp. (2.4%). The percentages of Streptococcus spp. DNA decreased, and those of Enterobactericeae increased in coronary plaques along with age., Conclusions: DNA of the Clostridium leptum group and pathogenic Enterobactericeae increase in the gut microbiome with age and can be detected in the same individual's coronary plaques along with pathogenic Streptococcus spp., associating with more severe coronary atherosclerosis., Competing Interests: We have the following interests: Sari Tuomisto, Terho Lehtimäki and Pekka J. Karhunen have been employed by Fimlab Laboratories Ltd. There are no patents, products in development or marketed products to declare. This does not alter our adherence to all the PLOS ONE policies on sharing data and materials.

Background Chronic infections have been reported to be risk factors for both coronary heart disease and ischemic stroke. DNA of oral bacteria, mainly from the viridans streptococci group, has been detected in coronary thrombus aspirates of myocardial infarction and cerebral aneurysms. Viridans streptococci are known to cause infective endocarditis and possess thrombogenic properties. We studied the presence of oral bacterial DNA in thrombus aspirates of patients with acute ischemic stroke treated with mechanical thrombectomy. Methods and Results Thrombus aspirates and arterial blood were taken from 75 patients (69% men; mean age, 67 years) with acute ischemic stroke. The presence of Streptococcus species, mainly the Streptococcus mitis group, belonging to viridans streptococci as well as Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans in samples were determined using a quantitative polymerase chain reaction with specific primers and probes. The relative amount of bacterial DNA in a sample was determined with the comparative threshold cycle method. Bacterial DNA was detected in 84% (n=63) of aspired thrombi, and 16% (n=12) of samples were considered bacterial DNA negative. DNA of Streptococcus species, mainly the S mitis group, was found in 79% (n=59) of samples. The median relative amount of Streptococcus species DNA was 5.10-fold higher compared with the control blood samples from the same patients. All thrombi were negative for both P gingivalis and A actinomycetemcomitans. Conclusions This is the first study showing the common presence of bacterial DNA from viridans streptococci in aspired thrombi of patients with acute ischemic stroke. Streptococcal bacteria, mostly of oral origin, may contribute to the progression and thrombotic events of cerebrovascular diseases.

Death does not occur instantaneously and organs do not decompose at the same rate or in the same way. Nulligravid human uteri and prostate glands are the last internal organs to deteriorate during decomposition; however, the reason for this very important observation is still enigmatic. Recent studies have elucidated that the composition and abundance of microbes in the human thanatomicrobiome (microbiome of death) varies by organ and changes as a function of time and temperature. The ileocecal area has the largest absolute postmortem burden that spreads to the liver and spleen and continues to the heart and brain depending on the cause of death. To truly understand the mechanisms of microbial assembly during decomposition, a thorough examination of different strategies utilized by the trillions of microbes that colonize decaying tissues is needed from a multi-organ and multidisciplinary approach. In this review, we highlight interdisciplinary research and provide an overview of human decomposition investigations of thanatomicrobiomic changes in internal organs.

Objective: Increasing data supports the role of bacterial inflammation in adverse events of cardiovascular and cerebrovascular diseases. In our previous research, DNA of bacterial species found in coronary artery thrombus aspirates and ruptured cerebral aneurysms were mostly of endodontic and periodontal origin, where Streptococcus mitis group DNA was the most common. We hypothesized that the genomes of S mitis group could be identified in thrombus aspirates of patients with lower limb arterial and deep venous thrombosis., Methods: Thrombus aspirates and control blood samples taken from 42 patients with acute or acute-on-chronic lower limb ischemia (Rutherford I-IIb) owing to arterial or graft thrombosis (n = 31) or lower limb deep venous thrombosis (n = 11) were examined using a quantitative real-time polymerase chain reaction to detect all possible bacterial DNA and DNA of S mitis group in particular. The samples were considered positive, if the amount of bacterial DNA in the thrombus aspirates was 2-fold or greater in comparison with control blood samples., Results: In the positive samples the mean difference for the total bacterial DNA was 12.1-fold (median, 7.1), whereas the differences for S mitis group DNA were a mean of 29.1 and a median of 5.2-fold. Of the arterial thrombus aspirates, 57.9% were positive for bacterial DNA, whereas bacterial genomes were found in 75% of bypass graft thrombosis with 77.8% of the prosthetic grafts being positive. Of the deep vein thrombus aspirates, 45.5% contained bacterial genomes. Most (80%) of bacterial DNA-positive cases contained DNA from the S mitis group. Previous arterial interventions were significantly associated with the occurrence of S mitis group DNA (P = .049, Fisher's exact test)., Conclusions: This is the first study to report the presence of bacterial DNA, predominantly of S mitis group origin, in the thrombus aspirates of surgical patients with lower limb arterial and deep venous thrombosis, suggesting their possible role in the pathogenesis of thrombotic events. Additional studies will, however, be needed to reach a final conclusion., (Copyright © 2017 Society for Vascular Surgery. Published by Elsevier Inc. All rights reserved.)

The purpose of the prospective research was to evaluate the benefit of urine specimen as a collection technique for biological forensic evidence in adult volunteers following consensual intercourse. For detecting Y-chromosomal material Buccal Swab Spin Protocol(®) was used in DNA extraction and purification and samples were analysed with Quantifiler Y Human Male DNA Quantification Kit(®). The time frame for positive Y-DNA was evaluated. Immediate microscopy for detection of spermatozoa was performed. Y-DNA was detected in 173/205 (84.4%) urine samples. Of the 86 first post-coital void urine samples available, Y-DNA was detected in 83 (96.5%) specimens. Of the 119 urine samples from volunteers with post-coital activities Y-DNA was still measurable in 70 (58.8%) urine specimens. The male DNA amount was below 0.023 ng/μl in 28/153 (18.3%) urine samples. Of the 22 urine samples obtained after 24 post-coital hours, 9 (40.9%) were still Y-DNA positive. No associations were found between coital durance, coital frequency during the past two weeks prior to the study intercourse, post-coital activities, and the urine sample Y-DNA positivity. Of the 111 urine samples where the immediate microscopy was performed, in 66 (59.5%) samples spermatozoa were verified and one sample even contained motile spermatozoa. Microscopy detected 66 (67.3%) and failed to detect spermatozoa in 32 (32.7%) of Y-DNA positive samples. In addition to conventional invasive swab techniques, urine samples seem to be an effective biological trace collection method for Y-DNA and spermatozoa within 24 h following penile-vaginal penetration. Furthermore, it may be considered as a non-invasive collection method in suspected acute child sexual abuse cases to diminish time delay in forensic evidence collection and to improve patients' positive attitudes towards evidence collection., (Copyright © 2015 Elsevier Ltd and Faculty of Forensic and Legal Medicine. All rights reserved.)

Introduction: Hantavirus infections are characterized by both activation and dysfunction of the endothelial cells. The underlying mechanisms of the disease pathogenesis are not fully understood. Here we tested the hypothesis whether the polymorphisms of endothelial nitric oxide synthase, eNOS G894T, and inducible nitric oxide synthase, iNOS G2087A, are associated with the severity of acute Puumala hantavirus (PUUV) infection., Patients and Methods: Hospitalized patients (n = 172) with serologically verified PUUV infection were examined. Clinical and laboratory variables reflecting disease severity were determined. The polymorphisms of eNOS G894T (Glu298Asp, rs1799983) and iNOS G2087A (Ser608Leu, rs2297518) were genotyped., Results: The rare eNOS G894T genotype was associated with the severity of acute kidney injury (AKI). The non-carriers of G-allele (TT-homozygotes) had higher maximum level of serum creatinine than the carriers of G-allele (GT-heterozygotes and GG-homozygotes; median 326, range 102-1041 vs. median 175, range 51-1499 μmol/l; p = 0.018, respectively). The length of hospital stay was longer in the non-carriers of G-allele than in G-allele carriers (median 8, range 3-14 vs. median 6, range 2-15 days; p = 0.032). The rare A-allele carriers (i.e. AA-homozygotes and GA-heterozygotes) of iNOS G2087A had lower minimum systolic and diastolic blood pressure than the non-carriers of A-allele (median 110, range 74-170 vs.116, range 86-162 mmHg, p = 0.019, and median 68, range 40-90 vs. 72, range 48-100 mmHg; p = 0.003, respectively)., Conclusions: Patients with the TT-homozygous genotype of eNOS G894T had more severe PUUV-induced AKI than the other genotypes. The eNOS G894T polymorphism may play role in the endothelial dysfunction observed during acute PUUV infection.

Background: We recently reported that large amounts of oral bacterial DNA can be found in thrombus aspirates of myocardial infarction patients. Some case reports describe bacterial findings in pericardial fluid, mostly done with conventional culturing and a few with PCR; in purulent pericarditis, nevertheless, bacterial PCR has not been used as a diagnostic method before., Objective: To find out whether bacterial DNA can be measured in the pericardial fluid and if it correlates with pathologic-anatomic findings linked to cardiovascular diseases., Methods: Twenty-two pericardial aspirates were collected aseptically prior to forensic autopsy at Tampere University Hospital during 2009-2010. Of the autopsies, 10 (45.5%) were free of coronary artery disease (CAD), 7 (31.8%) had mild and 5 (22.7%) had severe CAD. Bacterial DNA amounts were determined using real-time quantitative PCR with specific primers and probes for all bacterial strains associated with endodontic disease (Streptococcus mitis group, Streptococcus anginosus group, Staphylococcus aureus/Staphylococcus epidermidis, Prevotella intermedia, Parvimonas micra) and periodontal disease (Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Treponema denticola, Fusobacterium nucleatus, and Dialister pneumosintes)., Results: Of 22 cases, 14 (63.6%) were positive for endodontic and 8 (36.4%) for periodontal-disease-associated bacteria. Only one case was positive for bacterial culturing. There was a statistically significant association between the relative amount of bacterial DNA in the pericardial fluid and the severity of CAD (p=0.035)., Conclusions: Oral bacterial DNA was detectable in pericardial fluid and an association between the severity of CAD and the total amount of bacterial DNA in pericardial fluid was found, suggesting that this kind of measurement might be useful for clinical purposes.

The purpose of the research was to evaluate gynecological evidence collection techniques; the benefit of cervical canal brush sample compared to vaginal fornix and cervical swab samples and the time frame for detecting Y-chromosomal material QiAmp DNA Mini Kit(®) and Quantifiler Y Human Male DNA Quantification Kit(®) in adult volunteers following consensual intercourse. Eighty-four adult female volunteers following consensual intercourse were recruited for the study. By combining all sample collecting techniques, 81.0% of the volunteers were Y-DNA positive. Up to 60 h the conventional swab sampling techniques detected more Y-DNA positive samples when compared to the brush technique. However, after 60 h, the cervical canal brush sample technique showed its benefit by detecting 27.3% (6/22) of Y-DNA positive samples, which were Y-DNA negative in both conventional swab sampling techniques. By combining swab and brush techniques, 75% of the volunteers were still Y-DNA positive in 72-144 post-coital hours. The rate of measurable Y-DNA decreased approximately 3% per hour. Despite reported consensual intercourse, 6.8% (3/44) of volunteers were Y-DNA negative within 48 h. Y-DNA was not detected after 144 post-coital hours (6 days). In conclusion, the brush as a forensic evidence collection method may provide additional biological trace evidence from the cervical canal, although the best biological trace evidence collection can be obtained by combining all three sampling techniques. The time frame for gynecological forensic evidence sample collection should be considered to be at least a week if sexual violence is suspected., (Copyright © 2014 Elsevier Ltd and Faculty of Forensic and Legal Medicine. All rights reserved.)

Background: The liver is the first line of defence against continuously occurring influx of microbial-derived products and bacteria from the gut. Intestinal bacteria have been implicated in the pathogenesis of alcoholic liver cirrhosis. Escape of intestinal bacteria into the ascites is involved in the pathogenesis of spontaneous bacterial peritonitis, which is a common complication of liver cirrhosis. The association between faecal bacterial populations and alcoholic liver cirrhosis has not been resolved., Methods: Relative ratios of major commensal bacterial communities (Bacteroides spp., Bifidobacterium spp., Clostridium leptum group, Enterobactericaea and Lactobacillus spp.) were determined in faecal samples from post mortem examinations performed on 42 males, including cirrhotic alcoholics (n = 13), non-cirrhotic alcoholics (n = 15), non-alcoholic controls (n = 14) and in 7 healthy male volunteers using real-time quantitative PCR (RT-qPCR). Translocation of bacteria into liver in the autopsy cases and into the ascites of 12 volunteers with liver cirrhosis was also studied with RT-qPCR. CD14 immunostaining was performed for the autopsy liver samples., Results: Relative ratios of faecal bacteria in autopsy controls were comparable to those of healthy volunteers. Cirrhotics had in median 27 times more bacterial DNA of Enterobactericaea in faeces compared to the healthy volunteers (p = 0.011). Enterobactericaea were also the most common bacteria translocated into cirrhotic liver, although there were no statistically significant differences between the study groups. Of the ascites samples from the volunteers with liver cirrhosis, 50% contained bacterial DNA from Enterobactericaea, Clostridium leptum group or Lactobacillus spp.. The total bacterial DNA in autopsy liver was associated with the percentage of CD14 expression (p = 0.045). CD14 expression percentage in cirrhotics was significantly higher than in the autopsy controls (p = 0.004)., Conclusions: Our results suggest that translocation of intestinal bacteria into liver may be involved as a one factor in the pathogenesis of alcoholic liver cirrhosis.

Post mortem or even normal changes during life occurring in major gut bacterial populations are not known. We investigated Bacteroides sp., Bifidobacterium sp., Clostridium leptum, Clostridium coccoides, Streptococcus sp., Lactobacillus sp. and Enterobacteriacaea ratios in 7 fecal samples from healthy volunteers and in 61 autopsies rectum and cecum samples and studied the effect of post mortem time using quantitative real-time PCR. Bacterial ratios in stool samples from volunteers and rectum samples from autopsy cases were similar and did not change significantly up to 5 days post mortem. In cecum, significant post mortem time-dependent differences were observed in ratios of Bacteroides sp. (p = 0.014) and Lactobacillus sp. (p = 0.024). Our results showed that ratios of Bacteroides sp., Bifidobacterium sp., Clostridium leptum, Clostridium coccoides, Streptococcus sp., Lactobacillus sp. and Enterobacteriacaea can be investigated in autopsy rectum samples up to 5 days after death.

Postmortem bacteriology can be a valuable tool for evaluating deaths due to bacterial infection or for researching the involvement of bacteria in various diseases. In this study, time-dependent postmortem bacterial migration into liver, mesenteric lymph node, pericardial fluid, portal, and peripheral vein was analyzed in 33 autopsy cases by bacterial culturing and real-time quantitative polymerase chain reaction (RT-qPCR). None suffered or died from bacterial infection. According to culturing, pericardial fluid and liver were the most sterile samples up to 5 days postmortem. In these samples, multigrowth and staphylococci were not or rarely detected. RT-qPCR was more sensitive and showed higher bacterial positivity in all samples. Relative amounts of intestinal bacterial DNA (bifidobacteria, bacteroides, enterobacter, clostridia) increased with time. Sterility of blood samples was low during the studied time periods (1-7 days). The best postmortem microbiological sampling sites were pericardial fluid and liver up to 5 days after death., (© 2013 American Academy of Forensic Sciences.)

Introduction: Puumala virus (PUUV) infection is a viral hemorrhagic fever with renal syndrome (HFRS) characterized by thrombocytopenia and acute impairment of renal function. We aimed to assess whether genetic polymorphisms of platelet antigens together with those of von Willebrand factor (VWF) and plasminogen activator inhibitor (PAI-1) correlate with disease severity. Patients and methods 172 consecutive hospital-treated patients with serologically confirmed acute PUUV infection were included. Platelet glycoprotein (GP) IIIa T>C (rs5918), GP Ia T>C (rs1126643), GP Ib C>T (rs6065), GP VI T>C (rs1613662), VWF A>G (rs1063856) and PAI-1 A>G (rs2227631) were genotyped. The associations of the rarer alleles with variables reflecting the severity of the disease were analyzed., Results: PAI-1G-carriers had higher maximum creatinine level compared with the non-carriers (median 213 μmol/l, range 60-1499 μmol/l vs. median 122 μmol/l, range 51-1156 μmol/l, p = 0.01). The GG-genotypes had higher creatinine levels than GA- and AA-genotypes (medians 249 μmol/l, 204 μmol/l and 122 μmol/l, respectively, p = 0.03). Polymorphisms of GP VI and VWF associated with lower creatinine levels during PUUV infection. The minor C-allele of GP Ia associated with lower platelet counts (median 44 × 10(9)/l, range 20-90 × 10(9)/l vs median 64 × 10(9)/l, range 3-238 × 10(9)/l; p = 0.02)., Conclusions: Polymorphism of PAI-1, a major regulator of fibrinolysis, has an adverse impact on the outcome of kidney function in PUUV-HFRS. Platelet collagen receptor GP Ia polymorphism associates with lower platelet count., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Objectives: The frequency of adult-type hypolactasia (lactase non-persistence) varies widely among different ethnic groups. The cultural historical hypothesis assumes a link between the occurrence of hypolactasia and the distribution of dairy farming. The nomadic Nenets have been reindeer herders for generations and have therefore not consumed any dairy products. The hypotheses here was that the prevalence of lactase non-persistence (-13910 C/C genotype) among Nenets people having four Nenets grandparents is high, while the prevalence among Nenets originating from ethnically mixed families is lower., Study Design: The material was collected in four typical Nenets settlements in the Nenets Autonomous Okrug in Russia. One-third of the adult Nenets population were invited to answer a questionnaire and to donate buccal samples for genotyping by a doctor from the team of medical professionals who make rounds in this area. The total number of available participants was 177., Methods: Genotyping was performed with the AbiPrism system. We used the method of concordance of grandparents' national origin to ascribe ethnicity., Results: The prevalence of adult-type hypolactasia (-13910 C/C) among Nenets who had four Nenets grandparents was found to be 90%. The figures among others reporting three, two and one grandparent of Nenets origin were 72, 60 and 28%, respectively., Conclusion: The findings are in accord with the cultural historical hypothesis.

Background: Milk is an important source of nutrients. The consumption of milk, however, may cause abdominal complaints in lactose intolerant individuals. The frequency of -13910C/C genotype is known to be high among Northern Russians, exceeding the prevalence in northern Europe. In our study we tested two hypotheses: 1) subjects with lactase non-persistent genotype (-13910C/C) have more gastrointestinal (GI) symptoms associated with milk 2) subjects with lactase non-persistence avoid using milk., Methods: In total, 518 students aged 17 to 26 years were randomly selected from different departments in the Northern State Medical University (NSMU) for genotyping the lactase activity-defining -13910C/T variant. All subjects filled in a questionnaire covering their personal data, self-reported GI symptoms and milk consumption habits., Results: Northern Russians consume very small amounts of milk daily. Among carriers of the lactase non-persistent (LNP) genotype there were 10 percentage units of milk-consumers fewer than among lactase-persistent (LP) subjects (p = 0.03). Complaints of GI disorders caused by milk were different between the genotypes (p = 0.02). Among all types of food analyzed only milk was associated with increased GI symptoms among subjects with the LNP genotype (OR = 1.95, CI 1.03-3.69), Conclusions: Subjects with -13910C/C have more GI symptoms from milk. Subjects with lactase non-persistent genotype avoid using milk. In the case of increasing milk consumption symptoms may increase the need for medical consultation. It is thus important either for people themselves or for health care staff to be aware of lactase persistence/non-persistence.