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- Active Invasion of Porphyromonas gingivalis and Infection-Induced Complement Activation in ApoE-/- Mice Brains. [JOURNAL ARTICLE]
- J Alzheimers Dis 2014 Jul 24.
Periodontal disease is a polymicrobial inflammatory disease that leads to chronic systemic inflammation and direct infiltration of bacteria/bacterial components, which may contribute to the development of Alzheimer's disease. ApoE-/- mice were orally infected (n = 12) with Porphyromonas gingivalis, Treponema denticola, Tannerella forsythia, and Fusobacterium nucleatum as mono- and polymicrobial infections. ApoE-/- mice were sacrificed following 12 and 24 weeks of chronic infection. Bacterial genomic DNA was isolated from all brain tissues except for the F. nucleatum mono-infected group. Polymerase chain reaction was performed using universal 16 s rDNA primers and species-specific primer sets for each organism to determine whether the infecting pathogens accessed the brain. Sequencing amplification products confirmed the invasion of bacteria into the brain during infection. The innate immune responses were detected using antibodies against complement activation products of C3 convertase stage and the membrane attack complex. Molecular methods demonstrated that 6 out of 12 ApoE-/- mice brains contained P. gingivalis genomic DNA at 12 weeks (p = 0.006), and 9 out of 12 at 24 weeks of infection (p = 0.0001). Microglia in both infected and control groups demonstrated strong intracellular labeling with C3 and C9, due to on-going biosynthesis. The pyramidal neurons of the hippocampus in 4 out of 12 infected mice brains demonstrated characteristic opsonization with C3 activation fragments (p = 0.032). These results show that the oral pathogen P. gingivalis was able to access the ApoE-/- mice brain and thereby contributed to complement activation with bystander neuronal injury.
- Effect of repeated adjunctive antimicrobial photodynamic therapy on subgingival periodontal pathogens in the treatment of chronic periodontitis. [JOURNAL ARTICLE]
- Lasers Med Sci 2014 Jul 24.
The aim of this study was to compare the effect of subgingival ultrasonic scaling followed by repeated (three times) antimicrobial photodynamic therapy (PDT), ultrasonic scaling alone (US), and scaling and root planing with hand instruments (SRP) for initial periodontal treatment. Twenty-seven non-smoking systemically healthy chronic periodontitis patients were included. Residual pockets ≥4 mm deep and bleeding on probing were debrided either with SRP, US alone, or US followed by a single episode of PDT during supportive periodontal treatment. Probing pocket depth (PPD), bleeding on probing (BOP), and clinical attachment level (CAL) were monitored over 12 months. The presence of five periodontal pathogens in the pockets was determined by a commercially available micro-IDent test. Intergroup and intragroup statistical analysis was performed. All three treatments resulted in a significant clinical improvement. Additional application of PDT to US failed to result in further improvement in terms of PPD reduction and CAL gain. However, it resulted in a higher reduction of BOP at 3 and 12 months comparing to US alone or SRP (PDT from 25 to 13 and to 9 %, US from 23 to 16 and to 12 %, and SRP from 17 to 10 and to 9 %, respectively). PDT reduced the proportion of positive sites after 6 months for Treponema denticola (TD) significantly more effectively than US or SRP (p < 0.0001). Additionally, PDT resulted in a greater reduction of Aggregatibacter actinomycetemcomitans (AA), Tannerella forsythia (TF), and TD in medium pockets (4-6 mm) (p < 0.02) and of TD in deep pockets (>6 mm) compared to mechanical debridement alone (p < 0.05).
- Bioinformatic investigation of the cost management strategies of five oral microbes. [JOURNAL ARTICLE]
- Mol Oral Microbiol 2014 Jul 22.
Some amino acids are more energetically costly to synthesise de novo, therefore many microbes have evolved to regulate the metabolic expenditure of the cell and reduce the energy burden of extracellular unrecyclable proteins. Several oral bacterial species take up amino acids and peptides obtained from proteolysis of host proteins and hence do not rely only on de novo synthesis. The aim of this study was to investigate if five oral bacterial species implement cost management strategies to reduce the energy burden of extracellular unrecyclable proteins. Since the relative de novo amino acid synthetic costs are proportional to the masses of the amino acids, the energy costs of producing proteins were assessed by calculating the mean amino acid mass for each protein. For Porphyromonas gingivalis, Treponema denticola, Tannerella forsythia, Prevotella intermedia, and Streptococcus sanguinis, the outer membrane/extracellular proteins are made up of a much larger percentage of lower average mass amino acids whereas cytoplasmic proteins are made up of a larger proportion of higher average mass amino acid residues. These results are consistent with the five oral bacterial species employing energy-saving mechanisms in the production of extracellular unrecyclable proteins. Interestingly, the P. gingivalis and S. sanguinis genomes exhibited significantly lower predicted mean amino acid masses compared with that of the genomes of the other three species suggesting this may provide them with an energy advantage with respect to protein biosynthetic cost. This article is protected by copyright. All rights reserved.
- Quantitative analysis of classical and new putative periodontal pathogens in subgingival biofilm: a case-control study. [JOURNAL ARTICLE]
- J Periodontal Res 2014 Jul 14.
A number of species/phylotypes have been newly implicated as putative periopathogens. The objective of this study was to explore associations among classical and new pathogens in subgingival biofilm and to assess their relative importance to chronic periodontitis.Pooled subgingival biofilm samples were obtained from 40 patients with chronic periodontitis and 40 healthy controls. Taqman q-PCR assays were used to determine the absolute and relative counts of Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola, Parvimonas micra, Filifactor alocis, oral Synergistetes and oral TM7s. Microbial associations were assessed using cluster analysis. Different statistical models were used to explore associations between microbial parameters and periodontitis.The median log and relative counts were lowest for TM7s (4.4 and 0.0016%, respectively) and highest for oral Synergistetes (7.2 and 1.4%, respectively). Oral Synergistetes clustered strongly with the red complex, particularly T. forsythia (100% rescaled similarity). All species/phylotypes except TM7s were significantly associated with periodontitis (Mann-Whitney test; p ≤ 0.005). However, P. gingivalis and F. alocis lost association after adjusting for confounders (ordinal regression). In receiving operator characteristic curve analysis, the log counts of oral Synergistetes were the best markers of periodontitis (82.5% sensitivity and specificity), followed by those of T. forsythia, P. micra and T. denticola. In prediction analysis, however, P. micra was the only microbial predictor of periodontal parameters.Oral Synergistetes are presented here as new members of the red complex, with relative importance to periodontitis exceeding that of the classical members. P. micra is shown as an important periodontal pathogen warranting more attention.
- [An increase in the prevalence of syphilis in women in Eastern Bohemia - 30 years of surveillance.] [JOURNAL ARTICLE]
- Epidemiol Mikrobiol Imunol 2014; 63(2):95-98.
Epidemiological data on sexually transmitted infections in the Czech Republic has been carefully reported for many years. Here we present an analysis of regional data on syphilis spanning more than 30 years in eastern Bohemia. The epidemiological data were derived from the mandatory reporting of sexually transmitted diseases covering the period 1981-2011. The data showed a minimal incidence of cases in 1990 and an increasing trend thereafter. Two peaks in the incidence are apparent - within years 1995-1999 and 2003-2007. Interestingly, while before 1990 the numbers of positive men always exceeded those of women, their numbers equalized or even reversed within the two peaks. The results may also reflect trends in social change in the country after 1989. The analysis showed that the regional prevalence of syphilis in eastern Bohemia still remains low compared to some other regions of the Czech Republic. Keywords: syphilis - Treponema pallidum - Czech Republic.
- Epidemiology of Curable Sexually Transmitted Infections among Women at Increased Risk for HIV in Northwestern Tanzania: Inadequacy of Syndromic Management. [Journal Article]
- PLoS One 2014; 9(7):e101221.
Curable, non-viral pathogens account for a significant burden of sexually transmitted infections (STIs), and there is established evidence that STIs increase both HIV acquisition and transmission. We investigated the prevalence, trends, and factors associated with Chlamydia trachomatis, Neisseria gonorrhoeae, Trichomonas vaginalis and Treponema pallidum, and the performance of syndromic management, among a cohort of women working in bars, hotels, and other food and recreational facilities near large-scale mines in northwestern Tanzania.HIV-negative women aged 18-44 years (N = 966) were enrolled and followed for 12 months in a microbicides feasibility study. We collected sociodemographic and behavioural data, performed clinical examinations, and tested for STIs, at enrolment and 3-monthly. Risk factors for STIs were investigated using logistic regression models with random effects. Sensitivity, specificity and predictive values of syndromic management were calculated.At enrolment, the prevalences of C. trachomatis, N. gonorrhoeae, T. vaginalis, and high-titre active syphilis were 111/956 (12%), 42/955 (4%), 184/945 (19%) and 46/965 (5%), respectively. There were significant decreases over time for C. trachomatis and T. vaginalis (OR trend per month: 0.94 [95% CI 0.91, 0.97]; and 0.95 [0.93, 0.98], respectively; both p<0.001). The majority of these infections were not diagnosed by the corresponding syndrome; therefore, most participants were not treated at the diagnosis visit. Syndromic management was poorly predictive of laboratory-diagnosed infections. We identified a number of risk factors for STIs, including low educational level, some sexual behaviours, and ever having been pregnant.This analysis demonstrates that the prevalences of curable STIs are high among women who work in food and recreational facilities in northwestern Tanzania. Most of these infections are missed by syndromic management. Accurate and affordable rapid-point-of-care tests and innovative interventions are needed to reduce the burden of STIs in this population which is at increased risk for HIV.
- Advances and applications of molecular cloning in clinical microbiology. [Journal Article]
- Biotechnol Genet Eng Rev 2014.:65-78.
Molecular cloning is based on isolation of a DNA sequence of interest to obtain multiple copies of it in vitro. Application of this technique has become an increasingly important tool in clinical microbiology due to its simplicity, cost effectiveness, rapidity, and reliability. This review entails the recent advances in molecular cloning and its application in the clinical microbiology in the context of polymicrobial infections, recombinant antigens, recombinant vaccines, diagnostic probes, antimicrobial peptides, and recombinant cytokines. Culture-based methods in polymicrobial infection have many limitation, which has been overcome by cloning techniques and provide gold standard technique. Recombinant antigens produced by cloning technique are now being used for screening of HIV, HCV, HBV, CMV, Treponema pallidum, and other clinical infectious agents. Recombinant vaccines for hepatitis B, cholera, influenza A, and other diseases also use recombinant antigens which have replaced the use of live vaccines and thus reduce the risk for adverse effects. Gene probes developed by gene cloning have many applications including in early diagnosis of hereditary diseases, forensic investigations, and routine diagnosis. Industrial application of this technology produces new antibiotics in the form of antimicrobial peptides and recombinant cytokines that can be used as therapeutic agents.
- Dried blood spot testing for the antenatal screening of HTLV, HIV, syphilis, toxoplasmosis and hepatitis B and C: prevalence, accuracy and operational aspects. [JOURNAL ARTICLE]
- Braz J Infect Dis 2014 Jul 9.
Screening for vertically transmitted infection is mandatory and must be conducted at the first prenatal consultation. The most vulnerable women's groups are those at the lowest socio-economic level. Dried blood spot testing on filter paper could represent a secure way to screen pregnant women in the prenatal period.A cross-sectional study was conducted between November 2009 and March 2010, in the Metropolitan Region of Salvador, Bahia, Brazil, to compare the accuracy of the dried blood spot in filter paper and venipuncture serological as screening methods for HIV, HTLV, VHB, VHC, Treponema pallidum, and Toxoplasma gondii during prenatal period. Results of the venous blood sample collected in tubes were considered the gold standard.Serum samples and dried blood spot were obtained from 692 pregnant women aged between 14 and 42 years, with a median age of 26. Thirteen women were seropositive for T. gondii (1.88%; 95% CI: 0.60-2.71%), five for T. pallidum (0.72%; 95% CI: 0.15-1.61%), two for HBV (0.29%; 95% CI: 0.050.95%) and one for HTLV-1 (0.14%; 95% CI: 0.01-0.71%). No one was positive for HCV and HIV. The dried blood spot accuracy for syphilis and HTLV were 100% (95% CI: 99.25-100) and 100% (95% CI: 99.45-100%), respectively. The average time between blood collection and recording of the sample in the reference laboratory was 4.93 (3.82) days and between dried blood spot processing and active search for pregnant women was 3.44 (4.27) days.The use of dried blood spot may represent a secure way to expedite access to results of vertically transmitted diseases in the prenatal period, particularly in regions with scarce healthcare resources.
- Differential inflammatory responses of bovine foot skin fibroblasts and keratinocytes to digital dermatitis treponemes. [JOURNAL ARTICLE]
- Vet Immunol Immunopathol 2014 May 21.
Bovine digital dermatitis (BDD) is a serious infectious inflammatory lameness causing pain and suffering to many cattle worldwide and which has severe economic implications. This study set out to investigate relationships between the treponemes considered causal of BDD and the local inflammatory response of the bovine host. Here we describe, for the first time, the isolation of bovine foot skin keratinocytes and fibroblasts as separate cell lineages. These cell lines were then exposed to treponeme whole-cell sonicates, and the gene expression of selected host inflammatory mediators investigated using quantitative reverse transcriptase PCR. Several genes, including those encoding RANTES/CCL5, MMP12, TNFα, TGFβ and TIMP3 were significantly upregulated in fibroblasts exposed to whole-cell sonicates derived from BDD treponeme phylotypes. For each of the above genes there were similar fibroblast expression increases for all three BDD treponeme phylotypes tested, suggesting common virulence mechanisms. With bovine foot skin keratinocytes, we were unable to detect expression of RANTES/CCL5 and after incubation with BDD treponeme constituents we were unable to observe any significant changes in expression of inflammatory mediators tested. These contrasting results suggest fibroblasts rather than keratinocytes may be an important shared target of pathogenesis for BDD treponemes.
- Malignant Lues in an immunocompetent patient. [JOURNAL ARTICLE]
- Int J STD AIDS 2014 Jul 11.
Malignant Lues is a rare form of secondary syphilis mostly associated with HIV infection. It is an uncommon presentation of syphilis even rarer in immunocompetent patients. We present the case of a 57-year-old homosexual man referred to our department due to a 4-month history of a disseminated, slightly painful, nodular-ulcerative cutaneous eruption associated with low-grade fever, malaise and asthenia. Regarding the clinical features and serological and histopathological findings, the diagnosis of syphilis maligna was assumed. Serology for HIV was repeatedly negative. This case is interesting, not only because a very uncommon form of secondary syphilis was identified but also for being diagnosed in an immunocompetent patient. Lack of awareness of this type of presentation delays the diagnosis and treatment, leading to an increase in morbidity and spread of infection.