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The 2020 Sustainability Index (SI), a benchmark system, shows that 2% of the 184 municipal water and wastewater utilities (WWS) in Sweden have a good performance level (green), i.e., they meet all benchmark requirements for the “climate adaptation and flood safety” (“CA and FS”). In this study, ten Swedish WWS organizations were selected and studied in depth. The goal was to present them as good learning examples to inspire other utilities seeking to improve their results and to clarify and concretize the driving factors, strategies, and important explanations for their success, and the challenges they face. A total of 52 SI annual documents from ten utilities were analyzed. Six of their managers were interviewed in depth. One of the ten utilities studied was green on the parameter “CA and FS”. Flooding events in two utilities drove climate adaptation. The formation of an interdisciplinary group in two organizations had a major impact on their success. Two utilities focused on low-lying sites and enclosed spaces. Three believed capacity building increased their chances of success. The biggest challenge was sharing responsibility. That only 2% of municipal water and wastewater utilities are green is not the whole truth. Therefore, there should be other factors in SI that measure performance in “CA and FS”. To improve the results, new laws are needed to solve the problem of shared responsibility.

The book Resilient Water Services and Systems: The Foundation of Well-Being was published 2019. It is edited by Petri Juuti, Harri Mattila, Riikka Rajala, Klaas Schwartz, Chad Staddon, and published by the International Water Association (IWA) Publishers, 2019. The book is reviewed and recommended for both scholars and practitioners to study.

Background: The proportion of Mycoplasma pneumoniae isolates carrying point mutations in the 23S region of the genome associated with macrolide resistance has increased. We evaluated the probability of developing M. pneumoniae macrolide resistance mutations during macrolide treatment. Methods: M. pneumoniae strains from initial and follow-up oropharyngeal samples from 38 Swedish patients were tested for 23S rRNA mutations by amplifying and pyrosequencing the gene target region. The duration of symptoms and of M. pneumoniae carriage were recorded in cases with and without the macrolide resistance mutation. Results: Macrolide resistance mutations were absent in all baseline strains. Twenty-two patients were prescribed macrolides; the longest M. pneumoniae carriage time in these patients was 7 months and the longest symptomatic period was 7.5 months. Macrolide resistance was detected after macrolide treatment in 1 patient (carriage time 3.5 months and symptomatic period 7.5 months). Conclusions: M. pneumoniae may develop macrolide resistance during macrolide treatment. A long duration of M. pneumoniae carriage and symptoms is common in patients regardless of the presence of resistance. (Less)

The development of a plasmid-based genetic transformation protocol for Chlamydia trachomatis provides the basis for the detailed investigation of the function of the chlamydial plasmid and its individual genes or coding sequences (CDS). In this study we constructed a plasmid vector with CDS6 deleted (pCDS6KO) from the original Escherichia coli/C. trachomatis shuttle vector pGFP::SW2. pCDS6KO was transformed into a clinical isolate of C. trachomatis from Sweden that is plasmid-free (C. trachomatis SWFP-). Penicillin-resistant transformants expressing the green fluorescent protein were selected. These transformants did not stain with iodine, indicating that this property is regulated by CDS6 or its gene product. In addition, mature inclusions of C. trachomatis SWFP- transformed by pCDS6KO displayed an identical morphological phenotype to the untransformed plasmid-free recipient host. In this phenotype the morphology of inclusions was altered with the chlamydiae lining the periphery of the inclusion leaving a 'hole' in the centre. These green fluorescent inclusions appear 'doughnut-shaped' with an empty centre when examined under blue light, giving rise to a characteristic 'black hole' phenotype. Our study demonstrates the power of the new genetic system for investigating chlamydial gene function using gene deletion technology.

The synergism of infection with conventional cardiovascular risk factors in atherosclerosis is much debated. We hypothesized that coronary arterial injury correlates with infection recurrence and pathogen burden and is further aggravated by hypercholesterolemia. Forty-two Göttingen minipigs were assigned to repeated intratracheal inoculation of PBS, Chlamydia pneumoniae (Cpn), or both Cpn and influenza virus at 8, 11, and 14 wk of age. Animals were fed either standard or 2% cholesterol diet (chol-diet). At 19 wk of age coronary vasomotor responses to acetylcholine (ACh) and adenosine were assessed in vivo and blood and tissue samples were collected. Nonparametric tests were used to compare the groups. In cholesterol-fed animals, total cholesterol/HDL was significantly increased in infected animals compared with noninfected animals [3.13 (2.17–3.38) vs. 2.03 (1.53–2.41), respectively; P = 0.01]. C-reactive protein (CRP) rose in infected animals [10.60 (4.96–18.00) vs. 2.47 (1.44–3.01) μg/ml in noninfected; P < 0.01] without significant difference between the mono- and coinfected groups. Among coinfected animals, both CRP and haptoglobin were lower in those fed chol-diet than in those fed standard diet ( P < 0.05). The vasoconstricting response to ACh was most prominent in coinfected animals {769.3 (594–1,129) cm; P = 0.03 vs. noninfected [342 (309–455) cm] and P = 0.07 vs. monoinfected [415 (252.5–971.8) cm]}. Among monoinfected animals, similar to CRP, a trend for less vasoconstriction was observed in those fed chol-diet ( P = 0.08). Coinfection of piglets appears to be associated with more pronounced coronary muscarinic vasomotor dysfunction. In monoinfected animals, use of chol-diet seems to dampen both coronary dysfunction and systemic inflammation induced by infection.

Objectives: Endothelial cells of coronary arteries undergo apoptosis in patients with coronary artery disease. Carotid artery ultrasound studies show that after acute infections intima media thickness is increased. The possible effects of infections on endothelial cell apoptosis and the presence of apoptosis in early life remain unknown. Methods: Chlamydia pneumonia was inoculated three times at three weeks intervals to half of 32 piglets starting at the age of 8 weeks. Half of the piglets received cholesterol feeding. Morphological studies were done at the age of 19 weeks. Right coronary arteries were carefully excised and trimmed for excess tissue. The vessels were cut perpendicular to the long axis and sectioned at 5-μm thickness. TUNEL-assay was based on nick end labelling and staining of internucleosomal DNA fragments that are the biochemical hallmark of apoptosis. The percentage of apoptotic endothelial cells from intact endothe-lial cells was calculated using light microscopy. Results. Endothelial apoptosis appeared in 0.13% (SD 0.30%) of endothelial cells of non-infected piglets but in 0.75% (SD 0.62) of infected piglets (p

The Occurrence of plasma CRP during the febrile response after the first injection of penicillin was followed in 10 patients with early syphilis. An increase in CRP was noted after 12 hours, with a maximum after 24 hours. The appearance of this acute phase protein could not be correlated to cutaneous reaction, increased body temperature or leukocytosis, nor were baseline values of CRP correlated to clinical or serological activity of the infection. Patients with high levels of CRP prior to treatment suffered the most intense systemic reactions. No activation of the complement system during the Jarisch-Herxheimer reaction was detected from sequential determinations of C3, C4 and properdin.

Background. Mannose-binding lectin (MBL) is a multifunctional protein involved in innate immunity. We tested whether MBL and elevated viral and bacterial antibodies were risk factors for acute coronary events. Design. Controlled cohort study. Methods. A total of 354 patients with unstable angina pectoris (UA) or acute myocardial infarction (AMI) were compared with 334 paired controls. Results. Enterovirus titres were associated with increased risk of UA (odds ratio 10.04, P

The clinical performance of two nucleic acid amplification assays targeting the cryptic plasmid and two assays targeting rRNA molecules in Chlamydia trachomatis was examined. First-catch urine samples from Malmoe, Sweden, were tested for C. trachomatis with the Abbott real-time PCR assay m2000 and an in-house PCR for the new variant strain of C. trachomatis with a deletion in the cryptic plasmid. Aliquots of the urine samples were sent to Aarhus, Denmark, and further examined with the Roche COBAS Amplicor CT (RCA) PCR, the Gen-Probe Aptima Combo 2 assay (AC2) targeting the C. trachomatis 23S rRNA, and the Aptima C. trachomatis assay (ACT) targeting the 16S rRNA molecule. A positive prevalence of 9% (163/1,808 urine samples examined) was detected according to the combined reference standard. The clinical sensitivity and specificity of the four assays were as follows: for ACT, 100% (163/163) and 99.9% (1,643/1,645), respectively; for AC2, 100% (163/163) and 99.6% (1,640/1,645); for m2000, 68.7% (112/163) and 99.9% (1,644/1,645); for RCA, 63.8% (104/163) and 99.9% (1,643/1,645). The two Gen-Probe assays detected all mutant strains characterized by the in-house PCR as having the deletion in the cryptic plasmid, whereas the Roche and the Abbott PCRs targeting the plasmid were both unable to detect the plasmid mutant. The difference in clinical sensitivity between the plasmid PCR assays m2000 and RCA, on the one hand, and the rRNA assays AC2 and ACT, on the other, could be attributed almost exclusively to the presence of the plasmid mutant in about one-quarter of the Chlamydia -positive samples examined.

Objectives: The role of persistent chlamydial infection of the Fallopian tubes in ectopic pregnancy is still unresolved. Therefore, we examined tissue of the Fallopian tubes from patients with ectopic pregnancy for the presence of Chlamydia trachomatis. In addition, other markers of C. trachomatis infection implicated in the pathogenesis of tubal damage were studied including antibodies to heat shock protein 60 of chlamydial and human origin. Study design: Fresh frozen tubal tissue from 55 patients with ectopic pregnancy in a hospital setting were examined for the presence of C. trachomatis DNA by polymerase chain reaction (PCR) and blood sample were analysed for antibodies to C. trachomatis including heat shock protein 60 (hsp60). Results: Chlamydial DNA was not detected in any of the 55 tubal specimens using a commercial test, Cobas Amplicor, Roche, and an in-house real time PCR able to detect a few copies of the organism. Logistic regression showed that chlamydial IgG antibodies were more common in a subgroup of patients with previous PID than in controls (OR = 7.84, CI 1.78-34.6). Specific antibodies to hsp60 of chlamydial (OR = 7.00, CI 1.50-32.6) but not of human origin (OR = 2.13, CI 0.14-31.6) were associated with ectopic pregnancy in this group. Conclusions: No evidence of persistent infection of C. trachomatis in the fallopian tubes at the time of ectopic pregnancy was found in this study. (C) 2006 Elsevier Ireland Ltd. All rights reserved. (Less)

A nested case–control study of invasive and in situ cervical cancer was performed within a community-based cohort of 13 595 Taiwanese women assembled in 1991, with a follow-up period of 9 years. Baseline serum or plasma samples were analysed for antibodies against human papillomavirus (HPV) types 6, 16 and 18 and Chlamydia trachomatis. In total, 114 cases (42 incident cases identified during follow-up and 72 prevalent cases identified at baseline) and 519 matched controls were included in the study. HPV-16 seropositivity was strongly associated with cervical cancer (OR=6.33; 95 % CI 3.45–11.62). Overall, C. trachomatis was not associated with cervical cancer, but was associated with cervical cancer in analyses restricted to incident cases of cancer (OR=2.94; 95 % CI 1.17–7.42) or to cases in which serum samples were analysed (OR=3.13; 95 % CI 1.16–8.47). An antagonistic interaction between HPV-6 and -16 was found in a multiplicative model. These results suggest that different HPV types might interfere in cervical carcinogenesis and that C. trachomatis is associated with cervical cancer in prospective studies, and support the notion that HPV-16 seropositivity is strongly associated with cervical cancer.

The risk for coronary events may rise during acute infection. Perturbation in coronary endothelial function emerges as one important link. We investigated whether simvastatin could protect the coronary arterial function from the adverse effects of acute infection in swine.Coronary endothelium-dependent and -independent vasomotor responses were assessed by Doppler velocimetry in 12 Chlamydia pneumoniae-infected and 6 sham-infected swine 2 weeks after intratracheal inoculation. Half of animals from the infection group were pre-treated with simvastatin (80 mg daily), while the remaining animals received placebo. The treatment was started 2 weeks prior to inoculation and continued until the end of the study. ANOVA was used for statistical calculations. Data are mean+/-S.D.All animals inoculated with C. pneumoniae developed IgM antibodies against this organism. As compared to noninfected animals, peak-to-baseline coronary flow velocity (CFV) ratio after bradykinin was significantly decreased in infected animals regardless of statin treatment (p=0.01). Intracoronary 10(-6) M acetylcholine caused slight dilatory responses in both noninfected and infected-treated animals (CFV ratio: 1.6+/-0.2 and 1.4+/-0.2, respectively; p0.1), while a velocity drop (CFV ratio: 0.7+/-0.1; p0.01 versus noninfected-infected and treated), indicating constriction, was observed in infected-nontreated animals; 10(-5) M acetylcholine caused vasoconstriction in all animals, with a significantly more prolonged response in the infected-nontreated group (p0.01). Intracoronary adenosine and SNP induced similar dilatory responses in all groups (p0.5). There were no differences in markers of systemic inflammation (fibrinogen, amyloid, and CRP) and lipid profile (HDL, LDL and total cholesterol) between the groups (p0.2).Acute infection is associated with impairment of the muscarinic and kinin-related reactivity of coronary circulation. These functional abnormalities are in part prevented by simvastatin through mechanisms unrelated to lipid lowering.

Introduction Chronic rejection (CHR) of organ allografts, one of the most significant problems in modern transplantation, is not fully understood. This study sought to evaluate the influence of selected parameters on late kidney transplant function. Patients and Method The studied group consisted of eighty-six patients who received allogeneic transplants between 1988 and 1999 for leukocyte Chlamydia pneumoniae–DNA, immunoglobulin (Ig)A/IgG anti–C pneumoniae, blood lipids, ischemic damage in the donor and during organ preservation, HLA mismatch, and acute rejection episodes. Results Eighty-six patients were segregated as 26 patients (30%) with histologically proven chronic graft rejection (CHR[+]) and 59 patients (70%) without (CHR[−]). The presence of C pneumoniae–DNA in peripheral blood leukocytes was significantly more frequent in CHR(+) than CHR(−) group (46% vs 20%). Patients with leukocytes positive for C pneumoniae–DNA more frequently (50%) had CHR than patients negative for C pneumoniae–DNA (22%). CHR(+) patients showed significantly lower HDL levels (47 mg/dL vs 58 mg/dL) and higher triglyceride levels (193 mg/dL vs 148 mg/dL). To study the cumulative effect of differences between the CHR(+) and CHR(−) groups, we applied a multiple binary logistic regression analysis. An econometric model enabled us to calculate the probability of CHR for a given patient taking into account covariates chosen by means of stepwise selection: the presence of C pneumoniae–DNA in blood leukocytes, the use of continuous pulsatile perfusion in hypothermia, myocardial infarction occurrence, and triglyceride concentrations. Conclusion The presence of C pneumoniae–DNA in peripheral blood leukocytes increased the risk of CHR, which may be predicted by a multifactor analysis of chosen parameters. (Less)

Background: Upregulation of proinflammatory endothelial cell adhesion molecules and decreased bioactivity of endothelial nitric oxide (NO) are important in the pathogenesis of atherosclerosis. We investigated the effects of co-infection with Chlamydia pneumoniae and Helicobacter pylori on these two events in apoE-KO mice. Methods: Thirty-two apoE-KO mice, 8 weeks old, were equally divided into 4 groups. The first 2 groups were infected with either C. pneumoniae or H. pylori, while the 3rd group was infected with both C. pneumoniae and H. pylori. Mice from the 4th group and 4 wild-type mice served as controls. Thoracic and abdominal aortas were harvested after 10 weeks, and staining for vascular cell adhesion molecule-1 (VCAM-1) and intracellular adhesion molecule-1 was analyzed by immunocytochemistry. The endothelial vasomotor responses of thoracic aortas to methacholine were studied in organ chambers in the absence and presence of L-NAME. The plasma levels of nitrate/nitrite were measured. Results: Staining for VCAM-1 was more intense at the branching sites of aortas from mice with co-infection than in mono-infected or noninfected apoE-KO mice. The relaxation responses to methacholine and the plasma levels of nitrate/nitrite were significantly less in the co-infected group than in the other groups (p < 0.05). Conclusion: Co-infection of apoE-KO mice with C. pneumoniae and H. pylori seems to be associated with impaired bioactivity of endothelial NO and increased expression of VCAM-1 at branching sites. The findings may suggest an additive interaction of these pathogens in atherogenesis.

and 39%, respectively, for IgG and 7, 7, 3, and 7%, respectively, for IgA. The IgG seroprevalence rates for the PCR-positive women were two to three times higher than those for the PCR-negative women, i.e., 72 versus 29%, 72 versus 29%, 47 versus 26%, and 74 versus 25% for CT-EIA, SeroCT, CT pELISA, and the MIF assay, respectively. After discrepancy analysis, the sensitivity, specificity, positive predictive value, and negative predictive value were calculated for the IgG assays; for CT-EIA they were 84.7, 98.6, 98.4, and 86.7%, respectively; for CT pELISA they were 71.4, 97.3, 96.2, and 78.3%, respectively; for SeroCT they were 84.7, 98.6, 98.4, and 86.3%, respectively; and for the MIF assay they were 79.2, 83.1, 98.3, and 83.1%, respectively. In conclusion, these peptide-based ELISA systems for the serological detection of C. trachomatis infection performed as well as the MIF assay. Since these tests are less time-consuming, less expensive, and easier to perform than the MIF assay, they might be useful in the serodiagnosis of chlamydial infection. Chlamydia trachomatis infection is the most prevalent sexually transmitted disease in Europe and the United States. In women, this infection can lead to severe sequelae like ectopic pregnancy and tubal infertility. C. trachomatis serology has been used for both diagnostic purposes and large epidemiological studies. However, widespread introduction of C. trachomatis-specific serology has not gained wide acceptance. The microimmunofluorescence (MIF) assay (10) is still regarded as the “gold standard” in the serological diagnosis of C. trachomatis infections. However, the MIF assay is not suited for use by routine laboratories since reading of the specific fluorescence requires a high degree of expertise. Also, current serological tests employ either group-specific lipopolysaccharide (LPS) or reticulate bodies as antigen and thus show crossreactivity with C. pneumoniae (4). Serological cross-reactivity leads to high rates of false-positive results, especially in a population with a low prevalence of C. trachomatis infection, since the rates of C. pneumoniae seroprevalence are up to 60%. In addition, there have been reports on serological cross-reactivity due to proteins from other bacteria, e.g., Acinetobacter (2). Several user-friendly enzyme immunoassays with increased specificity have been developed by using LPS-stripped C. trachomatis particles (8). Recently, three commercially available assays have been developed by using specific synthetic peptides based on the major outer membrane of C. trachomatis. These assays have the potential to be both specific and sensitive. Proper comparison of these serological assays to the MIF assay is strongly needed since these new tests are well standardized, less expensive, and less laborious than MIF. Therefore, we analyzed these new commercially available, peptide-based enzyme-linked immunosorbent assay (ELISA) systems for the detection of specific serum immunoglobulin G (IgG) and IgA antibodies to C. trachomatis in relation to the detection of C. trachomatis infections in the corresponding cervical scrapings by PCR. The performances of these new assays were compared to that of the MIF assay.

A method for detection and genotyping of genital Chlamydia trachomatis infections based on omp1 gene amplification and sequencing was developed. DNA was extracted from urogenital or urine samples using a Chelex-based method, and an approximately 1,100-bp-long fragment from the omp1 gene was directly amplified and sequenced. Genotyping was performed by BLAST similarity search, and phylogenetic tree analysis was used to illustrate the evolutionary relationships between clinical isolates and reference strains. The method was used to determine the genotypes of C. trachomatis in 237 positive urogenital and/or urine specimens collected at a Swedish sexually transmitted disease clinic during 1 year. The most common genotypes corresponded to serotypes E (47%) and F (17%). The omp1 gene was highly conserved for genotype E (106 of 112 samples without any mutation) and F (41 of 42 samples without any mutation) strains but appear slightly less conserved for genotypes G ( n = 6) and H ( n = 6), where the sequences displayed one to four nucleotide substitutions relative to the reference sequence. Genotyping of samples collected at the follow-up visit indicated that two patients had become reinfected, while three other patients suffered treatment failure or reinfection. One woman appeared to have a mixed infection with two different C. trachomatis strains. This omp1 genotyping method had a high reproducibility and could be used for epidemiological characterization of sexually transmitted Chlamydia infections.

Serology is often used to diagnose acute infections by Chlamydia pneumoniae . In this study paired sera from patients with acute respiratory tract infection during an epidemic of C. pneumoniae infections were examined by five different antibody tests. These tests were the complement fixation (CF) test, the microimmunofluorescence (MIF) test, a recombinant enzyme immunoassay (rEIA) (Medac) based on a recombinant lipopolysaccharide of chlamydia and measuring antibodies to a common chlamydial antigen, and two tests that utilize preparations of C. pneumoniae organisms, the SeroCp-EIA (Savyon) (with preserved lipopolysaccharide) and the LOY-EIA (Labsystems) (without this antigen). Both of the last two tests should measure specific antibodies to C. pneumoniae , although cross-reacting antibodies may also be detected by the SeroCp-EIA. Acute infection of C. pneumoniae was serologically confirmed in 44% of the cases by at least two different tests. Using an expanded “gold standard,” i.e., the presence of significant reactions in at least two tests, the sensitivity of the CF test was 69%, that of the MIF test was 88%, that of the rEIA was 89%, that of the LOY-EIA was 96%, and that of the SeroCp-EIA was 92%. Specificity was high for all methods, but adjustments of diagnostic criteria were made to several of the tests. The basis for these adjustments and supportive data are presented. Infections of C. pneumoniae were detected in patients from 8 to 83 years of age. Two peaks in the incidence of such infections were observed: one among young teenagers and a second in adults 30 to 45 years of age, corresponding to parents of young teen-agers. The tests were equally sensitive in different age groups. Reinfections seemed to be rare.

Background —Arterial relaxation is largely regulated by endothelial nitric oxide (NO). Its diminished activity has been associated with incipient atherosclerosis. We investigated the endothelium-dependent relaxation of aorta in apolipoprotein E–knockout (apoE-KO) mice exposed to single or repeated Chlamydia pneumoniae inoculation. Methods and Results —Forty-eight apoE-KO mice, 8 weeks old, were inoculated intranasally with C pneumoniae (n=24) or saline (n=24) every 2 weeks over a 6-week period. Twenty mice (10 infected and 10 controls) were killed at 2 weeks and 6 weeks, respectively, after the first inoculation. The smooth muscle tone of aortic rings was measured in vitro at both time points. The norepinephrine-precontracted thoracic aortic rings were successively exposed to methacholine in the absence and presence of N G -nitro- l -arginine methyl ester (L-NAME) and diclofenac. The methacholine-induced relaxation was attenuated in the infected mice at 6 weeks in both the absence and presence of L-NAME ( P P P P Conclusions — C pneumoniae impairs arterial endothelial function, and the NO pathway is principally involved. Cyclooxygenase-dependent vasoconstricting products may also account for the infection-induced impaired relaxation. These findings further support the role of C pneumoniae infection in atherosclerosis development.

Background. To study the incidence of non-gonococcal salpingitis, gonococcal salpingitis and ectopic pregnancy in a defined population over a 28-year period on the assumption that the frequency of salpingitis and ectopic pregnancy may indirectly illustrate the epidemiological pattern of Chlamydia trachomatis. Design. A retrospective epidemiological study. Setting. University hospital with an urban catchment area. Patients. Five thousand two hundred and thirty-three patients admitted to the hospital between 1969 and 1996 with a diagnosis of ectopic pregnancy, non-gonococcal salpingitis, or gonococcal salpingitis. Results. The frequencies of both non-gonococcal and gonococcal salpingitis increased steeply early in the period under study, rising to a peak in the early 1970s, then decreasing throughout the period except for the last 3 years when a slight increase was seen again. The frequency of ectopic pregnancy showed a steady increase, peaking in the late 1980s and early 1990s and then declining at the end of the study period. While the introduction of more sensitive pregnancy tests and programs for assisted fertility would increase the rate of ectopic frequency the decline during the ʼnineties cannot be accounted for in this way. The peak of salpingitis cases in the early 'seventies seems to be mirrored exactly by the peak of ectopic pregnancies fifteen years later in the late 'eighties. Conclusion. The frequencies of salpingitis and of ectopic pregnancy can probably be used to estimate the incidence of preceding Chlamydia trachomatis. Thus the incidence of C. trachomatis has probably declined since the early 'seventies like that of N. gonorrheae.

Objectives: To investigate the presence of Chlamydia pneumoniae DNA in the wall of infrarenal abdominal aortic aneurysms, and in the wall of non-aneurysmal infrarenal abdominal aortas. Design: Case-control study. Materials and methods: The study group consisted of 40 patients operated transperitoneally for an infrarenal abdominal aortic aneurysm (IAAA) (eight females, 32 males; mean age 69 years, median age 68 years). Specimens from the aneurysm wall were taken peroperatively under sterile conditions. The control group consisted of 40 deceased persons without aortic aneurysms (14 females, 26 males; mean age 71 years, median age 70 years). Specimens from the non-aneurysmal infrarenal aortas (NIAA) were collected within 48 h after death. The specimens from both groups were frozen at −70 °C immediately after collection. A nested polymerase chain reaction (PCR) method, using two sets of primers designed to detect a fragment of the major outer membrane protein gene of C. pneumoniae , was used. Results: The detection of C. pneumoniae -specific DNA was significantly higher in the study group (14/40 = 35%) than in the control group (2/40=5%); ( p =0.001). No clinical factor predicting the presence of C. pneumoniae in the aneurysm wall, could be found. Conclusion: Chlamydia pneumoniae was detected at a significantly higher frequency in the wall of IAAAs than in the wall of NIAAs. Although this finding does not prove that C. pneumoniae causes IAAAs, further studies on the possible role of C. pneumoniae in the pathogenesis of aneurysms should be performed.

Chlamydia pneumoniae is a common cause of respiratory tract infection and community-acquired pneumonia. During an extensive outbreak of C. pneumoniae in northern Sweden, 319 respiratory samples from 129 persons were collected. Sputum, throat, and nasopharyngeal samples were obtained and analyzed by nested touchdown polymerase chain reaction (PCR), EIA, and culture in Hep-2 and McCoy cells. Serology was performed by complement fixation and microimmunofluorescence tests. By PCR, 30 patients were diagnosed with C. pneumoniae compared with 26 positive by EIA and 23 by culture. The finding of C. pneumoniae in the respiratory samples was accompanied by serology indicating acute infection in 26 (96%) of 27 patients for whom adequate sera were available. Nested PCR was sensitive and reliable for diagnosing acute respiratory C. pneumoniae infection. Sputum samples had the highest diagnostic efficacy, and the nested type of PCR was superior to one-step PCR. EIA and culture were less sensitive than nested PCR.

Our study had three objectives: to extend the plasmid-based transformation protocol to a clinical isolate of C. trachomatis belonging to the trachoma biovar, to provide "proof of principle'' that it is possible to "knock out'' selected plasmid genes (retaining a replication competent plasmid) and to investigate the plasticity of the plasmid. A recently developed, plasmid-based transformation protocol for LGV isolates of C. trachomatis was modified and a plasmid-free, genital tract C. trachomatis isolate from Sweden (SWFP-) was genetically transformed. Transformation of this non-LGV C. trachomatis host required a centrifugation step, but the absence of the natural plasmid removed the need for plaque purification of transformants. Transformants expressed GFP, were penicillin resistant and iodine stain positive for accumulated glycogen. The transforming plasmid did not recombine with the host chromosome. A derivative of pGFP::SW2 carrying a deletion of the plasmid CDS5 gene was engineered. CDS5 encodes pgp3, a protein secreted from the inclusion into the cell cytoplasm. This plasmid (pCDS5KO) was used to transform C. trachomatis SWFP-, and established that pgp3 is dispensable for plasmid function. The work shows it is possible to selectively delete segments of the chlamydial plasmid, and this is the first step towards a detailed molecular dissection of the role of the plasmid. The 3.6 kb beta-galactosidase cassette was inserted into the deletion site of CDS5 to produce plasmid placZ-CDS5KO. Transformants were penicillin resistant, expressed GFP and stained for glycogen. In addition, they expressed b-galactosidase showing that the lacZ cassette was functional in C. trachomatis. An assay was developed that allowed the visualisation of individual inclusions by X-gal staining. The ability to express active beta-galactosidase within chlamydial inclusions is an important advance as it allows simple, rapid assays to measure directly chlamydial infectivity without the need for plaquing, fluorescence or antibody staining.

OBJECTIVE: To estimate the prevalence of Chlamydia trachomatis, C. psittaci and C. pneumoniae antibodies in sera from altogether 931 blood donors, patients with symptoms of urethritis, assumed salpingitis and sexually acquired reactive arthritis (SARA), and women with fertility problems. METHODS: IgG antibodies to C. trachomatis, C. psittaci and C. pneumoniae were determined by microimmunofluorescence (MIF) tests. All patients were also tested for genital C. trachomatis infection using direct immunofluorescence (DIF) tests. RESULTS: The DIF-positive cases had a significantly (p < 0.0001) higher prevalence of C. trachomatis antibodies than the DIF negatives, i.e. 88.5% versus 14% in men with urethritis, 94.3% versus 36.4% in women with salpingitis, 66.7% versus 16.7% in SARA patients and 90.6% versus 20.8% in women with fertility problems. Antibody reactivity to all three chlamydial species was found significantly (p < 0.0001) more often in the patient groups and in those with a DIF-confirmed genital C. trachomatis infection than in blood donors. CONCLUSIONS: Presence of serum antibodies to C. trachomatis is tightly associated with the presence of chlamydiae in the genital tract, which also influences the cross-reactivities occurring in the MIF tests between chlamydial species.

Objective The longitudinal epidemiological development of the new variant of Chlamydia trachomatis was studied after appropriate testing procedures had been introduced when the strain was detected in 2006. Methods The number of cases of the new variant of C trachomatis was followed from 2007 through 2011 from the laboratory records. Testing for C trachomatis is centralised to one laboratory with around 80–85 000 persons being tested annually in a population of 1.1 million. Results During the 5-year period, 410 973 patients were tested of which 25 723 cases were positive. The proportion of the new variant of all positive cases declined from 30% in 2007 to 6% in 2011. While the number of the new variant of C trachomatis declined, the ordinary wild-type strains remained largely unchanged. Conclusions A selective decline of the new variant of C trachomatis has occurred after appropriate laboratory testing was introduced. A new balance point between 5% and 10% for the new variant seems to be gradually approached.

We aimed to evaluate the stability of the Chlamydia trachomatis multi locus VNTR analysis (MLVA-ompA) and multi sequence typing (MST) systems through multiple passages in tissue culture. Firstly, we analyzed the stability of these markers through adaptation of C. trachomatis to tissue culture and secondly, we examined the stability of a four-locus MLVA-ompA and a five-locus MST system after multiple passages in tissue culture. Marker sequences were monitored through successive chlamydial developmental cycles to evaluate the stability of the individual DNA markers through many bacterial divisions and this, in turn, informed us of the usefulness of using such typing systems for short and long-term molecular epidemiology. Southampton genitourinary medicine (GUM) clinic isolates from endocervical swabs collected from C. trachomatis positive women were passaged through tissue culture. MLVA-ompA typing was applied to primary swab samples and to the same samples after C. trachomatis had been passaged through cell culture (eight passages). Sequence data from time-zero and passage-eight isolates were aligned with reference sequences to determine the stability of the markers. The Swedish new variant (nvCT) underwent 72 passages in cell culture and the markers of the two schemes were similarly analyzed. Analysis of genetic markers of the MLVA-ompA typing system before and after the isolates were introduced to tissue culture showed no change in the dominant sequence. The nvCT that had been passaged 72 times over the duration of a year also showed no variation in the dominant sequence for both the genotyping schemes. MLVA-ompA and MST markers are stable upon adaptation of C. trachomatis to tissue culture following isolation of strains from primary endocervical swab samples. These markers remain stable throughout multiple rounds of cell-division in tissue culture, concomitant with the incubation period and appearance of symptoms normally associated with host-infection. Both genotyping schemes are, therefore, suitable for epidemiology of C. trachomatis.

Background: Respiratory agents may be detected in the oropharynx of healthy individuals. The extent of this condition and the reasons behind it are largely unknown. The objective of this study was to determine the factors associated with the presence of respiratory agents in the oropharynx of adolescents healthy enough to attend school activities. Methods: On a single day in December, samples from the posterior wall of the oropharynx of adolescents aged 10-15 y were obtained using cotton-tipped swabs. The samples were analyzed by real-time polymerase chain reaction (PCR) for the presence of 13 respiratory viruses and 2 bacteria (Mycoplasma pneumoniae and Chlamydophila pneumoniae). Results: Out of the 232 adolescents sampled, 67 (29%) had any respiratory symptom. A positive PCR result was found in 50 individuals (22%). Human rhinovirus was the most commonly found agent. Respiratory agents were significantly more frequent in the younger age group (10-13 y) than in the older age group (14-15 y): 26% (38/148) vs 14% (12/84), respectively; p = 0.04. Cough was the only symptom that was more common among individuals with a positive PCR test than among those with a negative PCR test: 8/50 (16%) vs 11/182 (6%); p = 0.02. Family size and class size were not associated with the likelihood of a positive PCR test. Conclusions: The presence of respiratory agents in the oropharynx is a frequent finding among adolescents healthy enough to attend school activities. The high prevalence was found to be associated with young age, but not with the size of the family or class. (Less)

Objective We sought to analyze the prevalence and clinical manifestations of Mycoplasma genitalium infection in a heterogeneous population of women. Study Design The study was designed as a cross-sectional case-control study. Women attending a gynecological outpatient service from 2003 through 2008 were invited to participate. Results The prevalence of M genitalium was 2.1% and of Chlamydia trachomatis was 2.8% among 5519 tested women. A total of 679 women were included. Both pelvic inflammatory disease (PID) and cervicitis were independently associated with M genitalium (odds ratio, 9.00; 95% confidence interval, 1.62–49.89 and odds ratio, 3.80; 95% confidence interval, 2.06–7.03, respectively). Women with C trachomatis had a higher frequency of both PID (18.3% vs 4.9%, P P M genitalium . Conclusion M genitalium was an independent and strong risk factor for both cervicitis and PID although, compared to C trachomatis , clinical manifestations were less frequent.

International audience; Chlamydia trachomatis is responsible for both trachoma and sexually transmitted infections, causing substantial morbidity and economic cost globally. Despite this, our knowledge of its population and evolutionary genetics is limited. Here we present a detailed phylogeny based on whole-genome sequencing of representative strains of C. trachomatis from both trachoma and lymphogranuloma venereum (LGV) biovars from temporally and geographically diverse sources. Our analysis shows that predicting phylogenetic structure using ompA, which is traditionally used to classify Chlamydia, is misleading because extensive recombination in this region masks any true relationships present. We show that in many instances, ompA is a chimera that can be exchanged in part or as a whole both within and between biovars. We also provide evidence for exchange of, and recombination within, the cryptic plasmid, which is another key diagnostic target. We used our phylogenetic framework to show how genetic exchange has manifested itself in ocular, urogenital and LGV C. trachomatis strains, including the epidemic LGV serotype L2b.

To study influenza virus shedding during acute infection, viral load was longitudinally measured by quantitative PCR in nasal flocked swabs from patients with seasonal H3N2 influenza at a Swedish emergency department, including both hospitalized patients and outpatients. Influenza A was detected in 65/184 patients. Sampling was repeated every 3-4 days in 45 patients, with the aim of continuing sampling until day 12 after disease onset. Home visits were offered. Antibodies were measured on paired sera in 95/184 patients. Fifty percent of the patients remained polymerase chain reaction (PCR)-positive 8 days after disease onset in a Kaplan-Meier survival curve. The longest observed duration of viral shedding was 12 days. The average viral load was initially low, peaked on days 2-3 of disease and then declined. Viral decline results remained similar when all 15 (25%) oseltamivir-treated patients were excluded. Significant antibody titre changes were seen in all the 35 PCR verified cases with available paired sera and in 8 of the 58 patients with negative PCR tests on acute phase nasal samples. In conclusion, quantitative PCR testing indicated the presence of influenza virus for up to 12 days, which could have implications for disease transmission and infection control.

Background Disease severity in Mycoplasma pneumoniae (MP) infection could potentially be related to bacterial factors such as MP genotype (MP1 or MP2; distinguished by different adhesions proteins) or bacterial load in airway secretions. We have compared these parameters in patients who were hospitalized for MP pneumonia, with outpatients with mild MP disease. Methods MP bacterial load was measured by real-time PCR in 45 in- and outpatients ("clinical study group") in whom MP DNA had been detected in oropharyngeal secretions by PCR. In addition, genotype and phylogenetic relationships were determined. The phylogenetical assessment was done by partial DNA sequencing of the P1 gene on isolates from 33 patients in the clinical study-group where sufficient DNA was available. The assessment was further extended to isolates from 13 MP-positive family members and 37 unselected MP positive patients from the two subsequent years and two different geographical locations. In total 83 strains were molecular characterized. Results Mean MP loads were significantly higher in 24 hospitalized patients than in 21 outpatients (1600 vs. 170 genomic equivalents/μL, p = 0.009). This difference remained significant after adjustment for age and days between disease onset and sampling. Hospitalized patients also had higher C-reactive protein levels. Mean levels were 188 vs 20 mg/L (p = 0,001). The genotype assessment showed MP genotype 1 in 17 of the 33 sequenced strains from the clinical study-group, and type 2 in 16 of these patients. Within each genotype, sequence differences were minimal. No association between disease severity and MP genotype was observed. In the extended genotype assessment, MP1 was found in similar proportions. In family contacts it was found in 53% and among patients from the two subsequent years 53% and 40%. Conclusions A higher MP bacterial load in throat secretions at diagnosis was associated with more advanced respiratory disease in patients, but MP genotype did not influence disease severity. Both MP genotypes co-circulated during recent outbreaks in Sweden.

In an analytical-method comparison study of clinical samples, the Abbott RealTi m e CT new formulation assay ( m 2000 real-time PCR), consisting of a duplex PCR targeting different parts of the cryptic plasmid in Chlamydia trachomatis , was compared both with version 2 of the Roche Cobas TaqMan CT assay, comprising a duplex PCR for a target in the cryptic plasmid and the omp1 gene, and with the Gen-Probe Aptima Combo 2 assay (AC2) targeting the C. trachomatis 23S rRNA molecule. First-catch urine samples from Sweden were tested in Malmö, Sweden, for C. trachomatis with the m 2000 real-time PCR assay and with an in-house PCR for the new variant C. trachomatis strain with a deletion in the cryptic plasmid. Aliquots of the urine samples were sent to Aarhus, Denmark, where they were further examined with the TaqMan CT and AC2 assays. A positive prevalence of 9.1% (148/1,632 urine samples examined) was detected according to the combined reference standard. The sensitivities and specificities of the three assays were as follows: for the Abbott m 2000 assay, 95.3% (141/148) and 99.9% (1,483/1,485), respectively; for the Roche TaqMan assay, 82.4% (122/148) and 100.0% (1,485/1,485); and for the Gen-Probe AC2 assay, 99.3% (147/148) and 99.9% (1,484/1,485). The plasmid mutant strain was detected in 24% (36/148) of the C. trachomatis -positive samples. There is a difference in sensitivity between the new formulations of the Abbott and the Roche assays, but both assays detected the wild-type and new variant C. trachomatis strains equally well.

Chlamydia trachomatisis a major cause of bacterial sexually transmitted infections worldwide. In 2006, a new variant ofC. trachomatis(nvCT), carrying a 377 bp deletion within the plasmid, was reported in Sweden. This deletion included the targets used by the commercial diagnostic systems from Roche and Abbott. The nvCT is clonal (serovar/genovar E) and it spread rapidly in Sweden, undiagnosed by these systems. The degree of spread may also indicate an increased biological fitness of nvCT. The aims of this study were to describe the genome of nvCT, to compare the nvCT genome to all availableC. trachomatisgenome sequences and to investigate the biological properties of nvCT. An early nvCT isolate (Sweden2) was analysed by genome sequencing, growth kinetics, microscopy, cell tropism assay and antimicrobial susceptibility testing. It was compared with relevantC. trachomatisisolates, including a similar serovar EC. trachomatiswild-type strain that circulated in Sweden prior to the initially undetected expansion of nvCT. The nvCT genome does not contain any major genetic polymorphisms – the genes for central metabolism, development cycle and virulence are conserved – or phenotypic characteristics that indicate any altered biological fitness. This is supported by the observations that the nvCT and wild-typeC. trachomatisinfections are very similar in terms of epidemiological distribution, and that differences in clinical signs are only described, in one study, in women. In conclusion, the nvCT does not appear to have any altered biological fitness. Therefore, the rapid transmission of nvCT in Sweden was due to the strong diagnostic selective advantage and its introduction into a high-frequency transmitting population.

Background: Atherosclerosis is considered to be an inflammatory disease. Infections are a significant cause of inflammation. Acute infections might precipitate acute coronary syndromes (ACS) whereas chronic infections might be stimuli for the development of atherosclerosis. Methods: Coronary angiograms were done on 211 of 335 patients with ACS and the percentage of coronary obstruction was determined. Serum antibody levels to Chlamydia pneumoniae, C. pneumoniae heat shock protein 60 (CpnHSP60), human heat shock protein 60 (hHSP60), enterovirus (EV), herpes simplex virus (HSV), cytomegalovirus (CMV), and two major periodontal pathogens, Aggregatibacter actinomycetemcomitans and Porphyromonas gingivalis, were measured in healthy controls (n = 355) and all patients. Results: Serum antibody levels to periodontal pathogens did not correlate with ACS. However, IgA-class antibody levels to Aggregatibacter actinomycetemcomitans ( p = 0.021), CpnHSP60 ( p = 0.048) an hHSP60 ( p = 0.038) were higher in patients with coronary occlusion or obstruction compared to those without any obstruction. Odds ratios for coronary changes in the highest quartile as compared to the lower quartiles were for A. actinomycetemcomitans IgA 7.84 (95% CI 1.02—60.39, p = 0.048), for CpnHSP60 IgA 8.61 (1.12—65.89, p = 0.038), and for human HSP60 IgA 3.51 (0.79—15.69, p = 0.100). Conclusions: We have previously reported that EV and HSV titres correlated significantly to acute coronary events. They do not correlate to the degree of coronary obstruction as shown here. However, infection by A. actinomycetemcomitans or C. pneumoniae or host response against them associated with coronary obstruction. Clinical coronary events may arise by the effect of acute infections and obstructing lesions by a chronic inflammatory stimulus.

BACKGROUND:: In 2006, a new genetic variant of Chlamydia trachomatis (nvCT) was discovered in Sweden. Clinical manifestations of this infection were studied in a high-risk population. METHODS:: During 2007, a prospective case-control study on sexual lifestyle and urogenital infections was performed at the Centre for Sexual Health (CSH), affiliated to Malmo University Hospital. A total of 629 C. trachomatis positive cases and 1252 negative controls were included. At Malmo University Hospital, Department of Obstetrics and Gynecology, all cases of pelvic inflammatory disease (PID) were assessed and correlated to the prevalence of nvCT. RESULTS:: Patients with nvCT or wild type C. trachomatis (wtCT) infection did not differ regarding their sexual lifestyle. Men with nvCT or wtCT infection did not differ in uro-genital symptoms or clinical findings. Women with nvCT infection reported painful urination (12.2% vs. 25.8%, P = 0.02) and were diagnosed with urethritis (11.1% vs. 40.0%, P = 0.04) less often than women with wtCT infection. The ratio of lower abdominal pain in women with nvCT infection was only half of that in women with wtCT infection (13.4% vs. 27.8%, P = 0.02). PID was detected in 0.8% of women with C. trachomatis infection in Malmo. All these cases were due to wtCT infection. CONCLUSIONS:: Symptomatic urethral infection and lower abdominal pain was less common in women with nvCT as compared to wtCT. Infection with nvCT was more frequently asymptomatic suggesting a possible difference in virulence between the nvCT strain and the wtCT strain. (Less)

Background Diagnosis of Mycoplasma pneumoniae (MP) infection is traditionally based on serology, which may require more than two weeks for diagnostic antibodies to develop. PCR-based methods offer earlier diagnosis. During a community outbreak of MP infection, we compared semi-nested and real-time PCR of oropharyngeal swabs with serology for diagnosis of MP infection at different time points after disease onset. PCR-positive individuals were followed longitudinally to assess the persistence of MP DNA in throat secretions. We also studied carriage of MP among household contacts and school children. Results MP infection was diagnosed in 48 of 164 patients with respiratory tract infection. Forty-five (29%) had detectable MP DNA in oropharynx. A significant increase in MP IgG IgG titre or MP IgM antibodies was detected in 44/154 (27%) subjects. Two MP PCR-positive patients lacked antibody responses. Sera were missing from another two patients. The agreement between serology and PCR was good, κ = 0.90. During the first three weeks after disease onset the performance of PCR was excellent and all patients but one were detected. In contrast, only 21% of the patients with confirmed MP infection were positive by serum 1 during the first symptomatic week (56% during the second and 100% during the third week). Only 1/237 (0.4%) school children was positive by PCR. This child had respiratory symptoms. Eighteen of 22 (75%) symptomatic household contacts were MP PCR positive. Persistence of MP DNA in the throat was common. Median time for carriage of MP DNA was 7 weeks after disease onset (range 2 days – 7 months). Adequate antibiotic treatment did not shorten the period of persistence. Bacterial load, measured by quantitative real-time PCR declined gradually, and all followed patients eventually became PCR-negative. Conclusion PCR is superior to serology for diagnosis of MP infection during the early phases of infection. Persistent, sometimes long-term, carriage of MP DNA in the throat is common following acute infection, and is not affected by antibiotic therapy. Asymptomatic carriage of MP even during an outbreak is uncommon.

BACKGROUND: Infections caused by Chlamydia pneumoniae are considered to participate in inflammatory processes leading to coronary artery disease. After a primary infection, the bacteria remain dormant intracellularly causing a chronic inflammatory stimulus. MATERIALS AND METHODS: Blood samples were obtained from 235 patients with acute myocardial infarction (AMI) and 108 patients with unstable angina pectoris (UA). We evaluated the prognostic significance of bacterial and viral antibody titers, serum troponin T, C-reactive protein, and chlamydial lipopolysaccharide (cLPS) concentrations during acute coronary syndrome of patients with AMI and UA for cardiovascular death and new UA and AMI that required hospital care during a 6-year follow-up. RESULTS: Serum cLPS levels correlated with C-reactive protein and serum troponin T concentrations during acute coronary events. Patients with AMI had significantly higher serum concentration of cLPS compared with patients with UA. Enterovirus antibody titers and cholesterol-lowering therapy at admission of the index event were negatively correlated with cLPS concentration (r = -.198, P = .0003 and r = -.26, P = .019, respectively). The presence of circulating cLPS was associated with a hazard ratio of 2.04 for a new cardiovascular event during the follow-up period (P = .006). The area under the curve in the receiver operating graph was .572. CONCLUSION: cLPS is evidently liberated from the infected atherosclerotic tissue during an acute coronary event. Our study supports the view that inflammation caused by C. pneumoniae infection is an important but as yet poorly understood factor in the development of atherosclerosis and may play a role in acute vascular events. (Less)

Background Impaired muscarinic receptor-mediated vasodilation is an important feature of early atherosclerosis. Earlier studies on apolipoprotein E-knockout mice (apoE-KO) mice suggested adverse effects of Chlamydia pneumoniae infection on the endothelial vasomotor responses of aortas to the muscarinic agonist methacholine. Using additional aorta samples the present study investigated the responses to bradykinin. Methods and Results ApoE-KO mice were repeatedly inoculated with either Chlamydia pneumoniae (C. pneumoniae) or saline. At 2, 6, and 10 weeks after the first inoculation, precontracted aorta rings from both groups were exposed to bradykinin in the absence and presence of L-NAME and diclofenac. In noninfected animals, the vasomotor responses to bradykinin were similar at all timepoints (p > 0.5). Compared with noninfected animals, the responses in infected animals tended to increase through the study period (p < 0.05 at 10 weeks). Although diclofenac and L-NAME had no effect in noninfected mice, they inhibited the responses to bradykinin in infected mice at 6 and, more markedly, 10 weeks (p < 0.05 for both). Conclusion Bradykinin stimulation of aorta endothelium from C. pneumoniae-infected apoE-KO animals appears to activate compensatory kinin receptor-related mechanisms that could involve nitric oxide and vasorelaxing prostanoids. Although the precise molecular mechanisms require further investigation, one could speculate that strategies increasing bradykinin availability might reverse the arterial dysfunction during chronic infectious disease.

The human polymnaviruses JC virus (JCV) and BK virus (BKV) are oncogenic in experimental systems and commonly infect humans. JCV DNA has been reported to be present in human colon mucosa and in colorectal cancers. To investigate whether the risk for colorectal cancer is associated with JCV or BKV infection, we performed a case-control study nested in the Janus biobank, a cohort of 330,000 healthy Norwegian subjects. A 30-year prospective follow-up using registry linkages identified 386 men with colorectal cancer who had baseline serum samples taken >3 months before diagnosis. Control subjects were matched for sex, age and date of blood sampling and county of residence. Seropositivity for JCV or BKV had high (97-100%) sensitivity for detection of viral DNA-positive subjects and discriminated the different polyomaviruses. Seropositivity was mostly stable over time in serial samples. The relative risk for colorectal cancer among JCV seropositive subjects was 0.9 (95% CI: 0.7-1.3) and the BKV-associated relative risk was 1.1 (95% CI: 0.8-1.5). Determining seropositivity using alternative cutoffs also found no evidence of excess risk. In summary, this prospective study found no association between JCV or BKV infections and excess risk for colorectal cancer.

Aims: The aim of the study was to explore whether exposure to microbial agents determines the prevalence of acute coronary events. Methods and results: Patients with unstable angina pectoris and myocardial infarction (N = 335) and their paired controls were investigated. The subjects answered a questionnaire about their childhood contagious diseases: varicella, scarlet fever, measles, rubella, mononucleosis and mumps. Blood samples were taken for bacterial and viral serology. The odds ratio for CHD was highest in the upper quartile of the enterovirus (EV), herpes simplex virus (HSV) and Chlamydia pneumoniae HSP60 IgG antibody titers (1.86, p = 0.001, 1.57, p < 0.048 and 1.70, p = 0.016, respectively). The antibody titers increased cumulatively the risk for CHD (odds ratios 1.89, 2.24, 3.92 and p-values < 0.001, 0.001 and 0.047). Childhood contagious diseases (n = 6) had a protecting effect against CHD (odds ratio 0.86, p = 0.013). The risk for acute coronary events decreased significantly with increasing number of childhood contagious diseases (p = 0.007). Conclusions: Infections have a dual role in the genesis of CHD. EV, HSV and C. pneumoniae heat shock protein 60 IgG antibodies are associated with increased risk for CHID. Protection from infections usually suffered during the childhood before the era of MMR vaccination may predispose the individual to CHD. (Less)

Human papillomavirus (HPV) persistence is the major cause of cervical cancer, but most HPV infections will not persist and risk factors for HPV persistence are not well known. Chlamydia (C.) trachomatis infection seems to also be associated with cervical cancer. We investigated whether C. trachomatis infection is a risk factor for HPV persistence. In a cohort of 12,527 women participating in a population-based HPV screening trial in Sweden, 6,418 women completed testing for HPV DNA by general primer PCR and typing by reverse dot blot hybridization. On average 19 months later, 303 women that had been HPV-positive and had normal cytology at enrollment completed a new HPV test. Environmental exposures were assessed by an 87-item questionnaire. Previous sexually transmitted infections were also investigated by serology. At follow-up, 44% of the women were positive for the same type of HPV DNA as at enrollment. Persistence correlated with length of follow-up (p < 0.01) and condom use seemed to protect against HPV persistence (p < 0.05). The most significant risk factor for persistent presence of HPV DNA was self-reported history of previous C. trachomatis infection (relative risk in multivariate model = 2.09; 95% confidence interval = 1.05-4.18). We conclude that persistence of oncogenic HPV infections is more likely among women with a previous C. trachomatis infection.

Aim: The purpose of the study was to assess the association between seropositivity to various infectious agents and stable coronary artery disease (CAD), controlling simultaneously for a variety of potential confounders. We also investigated whether the choice of a larger reference group might affect the results, and whether or not seropositivity to multiple agents was associated with a systemic inflammatory response. Methods: We assessed the simultaneous prevalence of antibodies against Helicobacter pylori, Chlamydia, cytomegalovirus, and herpes simplex virus in 312 patients with angiographically proven coronary artery disease (CAD) and in 479 age and sex matched controls. C-reactive protein, interleukin-6, fibrinogen, PAI- I -activity, D-dimer, von Willebrand Factor, plasma viscosity, and a complete blood cell count were determined in all subjects. Results: Seropositivity to all of the four agents was 21.8% in cases and 13.6% in controls (P = 0.0003). We found a dose-response relationship between combined IgG-seropositivity to H. pylori, Chlamydia, cytomegalovirus, and herpes simplex virus and odds for the presence of angiographically confirmed stable CAD which, however, was strongly reduced after controlling for a variety of potential confounders. The dose-response pattern was no longer evident if a more stable reference group (subjects seropositive for two agents) was used instead of the relatively small reference group with zero or one seropositivity. We found no consistent pattern between IgG-seropositivity to several pathogens and inflammatory markers. Conclusions: Based on serological evidence of various infectious agents, this study suggests that the aggregate number of persistent infections is not independently associated with an increased risk for CAD if control for confounding and use of a stable reference group are guaranteed. (C) 2003 Published by Elsevier Ireland Ltd. (Less)

Background: Coronary endothelial dysfunction contributes to the pathogenesis of acute coronary syndromes (ACSs). Acute Chlamydia pneumoniae infection has been epidemiologically associated with ACS. In this study, we investigated whether acute C. pneumoniae infection could alter the endothelial vasomotor function of porcine coronary vessels. Methods and results: Twenty pigs, 7–9 kg in weight, were inoculated intratracheally with C. pneumoniae (n=12) or saline (n=8), and investigated at 3 days (five infected/four non-infected) and 2 weeks (5+2 infected/four non-infected) after inoculation. The endothelium-dependent reactivity of coronary microcirculation was assessed at both time points by measuring peak coronary flow velocity (CFV) in response to bradykinin, before and after infusions with glutathione, an antioxidant, and Image-arginine, a substrate for nitric oxide synthase (NOS). CFV after bradykinin was significantly decreased in infected animals at both time points. At 2 weeks, both glutathione and Image-arginine significantly improved CFV after bradykinin. CFV after sodium nitroprusside (SNP) was similar in both groups. At 3 days, the relaxation responses of bradykinin-induced pre-contracted left anterior descending (LAD) coronary rings to bradykinin were significantly less in infected animals. NG-nitro-Image-arginine-methyl-ester, an NOS inhibitor, had significantly greater inhibitory effect on bradykinin-induced relaxation in infected animals. Plasma nitrate–nitrite and fibrinogen, and NOS activity from LAD coronary samples were significantly increased in infected animals. Conclusion: Acute C. pneumoniae infection causes endothelial dysfunction of both resistance and epicardial coronary vessels, and favours a pro-coagulant status. These effects could in part account for the epidemiologically suggested association between acute infection and ACS. (Less)

Our objectives were to compare the prevalence of sexually transmitted infections (STIs) in 103 women undergoing induced legal abortion (LA) and in 101 women with confirmed, recent illegal abortion (IA), in Maputo, Mozambique. For the purpose of this study, LA was considered the abortion provided in the Maputo Central Hospital with the approval of the Ministry of Health, and IA the one not provided through the approved facility, mentioned above. Women with IA were recruited in the outpatient gynaecology ward and women with LA in the emergency gynaecology ward in the Maputo Central Hospital, during the same time period. Serological tests for syphilis (rapid plasma reagin, ELISA-IgG and fluorescent treponemal antibody absorption), gonorrhoea (indirect haemagglutination) and chlamydia (microimmunofluorescence) were carried out. Direct immunofluorescence for detection of Chlamydia trachomatis antigen was done on endocervical smears from all the women. The prevalence of syphilis seropositivity in IA women is twice that of LA women (odds ratio [OR] 2.13; 95% confidence interval [CI] 0.55-9.95), with 10.9% and 4.9%, respectively. Exposure to gonorrhoea in these 2 groups is similar (OR 1.18; 95% CI 0.63-2.20), with seroprevalence of 31.1% in the LA and 34.7% in the IA group. The high titres are also similar in both groups. Serology findings for C. trachomatis indicate prevalence of seropositivity of 40.6% in the LA and 44.4% in the IA group with no significant difference (OR 1.17; 95% CI 0.64-2.13). In conclusion, STIs are highly prevalent in both IA and LA groups in Maputo. Urgent interventions are needed to reduce their prevalence and consequently their adverse consequences.

Abstract —Infectious agents may be involved in atherothrombogenesis. The potential pathogenic pathway, however, remains unclear. We investigated the association between various infectious agents and lipoproteins known to have an atherogenic effect. We recruited 470 healthy blood donors and 238 patients with angiographically proven coronary heart disease (CHD), aged 40 to 68 years. Seropositivity to Chlamydia pneumoniae (CP), chlamydial lipopolysaccharide, and cytomegalovirus (CMV) was determined; infection with Helicobacter pylori (HP) was assessed by using the [ 13 C]urea breath test. In all subjects, total cholesterol, high density lipoprotein (HDL) cholesterol, lipoprotein(a), and various apolipoproteins (apos) were determined. In unadjusted analysis, mean HDL cholesterol concentration was significantly decreased in HP-positive healthy subjects (1.36 vs 1.44 mmol/L, P =0.006) compared with HP-negative subjects. The HDL cholesterol to total cholesterol ratio was significantly decreased in HP-positive (0.259 vs 0.276, P =0.01) and CP-seropositive (0.266 vs 0.280, P =0.04) healthy subjects compared with (sero)negatives. Mean apoAI levels were significantly lower in HP-positive healthy subjects (1.46 vs 1.51 g/L, P =0.03) and in CMV-positive healthy subjects (1.47 vs 1.52 g/L, P =0.01) compared with (sero)negative subjects. After multivariable adjustment by means of linear regression analysis, only the association between HP infection and decreased HDL cholesterol ( P =0.002), decreased HDL cholesterol to total cholesterol ratio ( P =0.005), decreased apoAI ( P =0.02), and increased apoB ( P =0.02) persisted and remained significant. There was no independent association between other lipoproteins and serological markers of CP or CMV infection. Current infection with HP, but not seropositivity to CP or CMV, was associated with an atherogenic, modified lipid profile. These lipid alterations could explain, at least in part, the reported weak association between chronic HP infection and atherosclerotic diseases.

The reported rate of detection of Chlamydia pneumoniae DNA within atherosclerotic lesions by PCR varies between 0 and 100%. In this study, identical sets of coded experimental atheroma samples ( n = 15) and spiked controls ( n = 5) were analyzed by 16 test methods in nine centers by means of PCR. The positive controls were correctly identified to levels of 1, 0.1, and 0.01 inclusion bodies of C. pneumoniae /ml of tissue homogenate by 16 (100%), 11 (69%), and 3 (19%) of the test methods, respectively. Three out of 16 negative controls (19%) were rated positive. Positivity rates for atheroma samples varied between 0 and 60% for the different test methods, with the maximum concordant result for positivity being only 25% for one carotid artery sample. There was no consistent pattern of positive results among the various laboratories, and there was no correlation between the detection rates and the sensitivity of the assay used.

The lack of standardization in chlamydia serology has made interpretation of published data difficult. This study was initiated to determine the extent of interlaboratory variation of microimmunofluorescence (MIF) test results for the serodiagnosis of Chlamydia pneumoniae infections. Identical panels of 22 sera were sent to 14 laboratories in eight countries for the determination of IgG and IgM antibodies by MIF. Although there was extensive variation in the numeric titer values, the overall percentage agreement with the reference standard titers from the University of Washington was 80%. For results by serodiagnostic category, the best agreement was for four-fold rise in IgG titers, while the lowest agreement was for negative or low IgG titers. Agreement for IgM titers was 50%-95%. Four laboratories failed to discern false-positive IgM titers possibly because of the presence of rheumatoid factor. Further studies are underway to determine the source of interlaboratory variation for the MIF test.

This study was performed to generate and evaluate recombinant antigens for use in a species-specific Chlamydia trachomatis immunoassay. In a molecular genetic approach, fragments of the C. trachomatis major outer-membrane protein (MOMP) were produced as fusion proteins to create three different constructs encompassing the variable domains I, II and IV of selected C. trachomatis serovars. The recombinant MOMP polyantigens were affinity-purified and used in an enzyme-linked immunosorbent assay. Antibody detection was evaluated with 103 patient sera and the results were compared with titres obtained in the micro-immunofluorescence test. The results showed that the generated MOMP polyantigens detected the presence of C. trachomatis-specific human antibodies with little cross-reaction to C. pneumoniae-specific antibodies. When compared to the micro-immunofluorescence assay the MOMP polyantigen detected the presence of anti-C. trachomatis IgG antibodies with a sensitivity of 80% and a specificity of 91%.

A commercial test (rELISA) based on a recombinant chlamydial lipopolysaccharide (LPS) antigen has been evaluated for the diagnosis of acute infections caused by Chlamydia pneumoniae (TWAR) and Chlamydia psittaci. This test and a microimmunofluorescence test (MIF) were compared in 160 patients with community-acquired pneumonia. Seventeen of nineteen cases with significant titre changes detected by rELISA were confirmed by MIF. The two remaining cases not confirmed by MIF were considered false-positive reactions. One case positive by MIF only was judged not to be a true-positive reaction. All three cases occurred in patients with Mycoplasma pneumoniae infection and may be the result of a mitogenic effect. High antibody titres have been used to indicate acute C. pneumoniae infection. We found high MIF or rELISA titres to be equally common in patients and controls; no association between the two tests was detected. An unexpected cross-reactivity between the rELISA antigen and parvovirus was observed, which might have diagnostic implications. Both MIF and rELISA detected acute C. pneumoniae and C. psittaci infection, and there was good agreement between the tests. Single serum diagnosis was generally not feasible with either MIF or rELISA.