Specific detection of Treponema pallidum in clinical samples: validation of a qPCR assay combining two genomic targets.

OBJECTIVES: We evaluated a real-time quantitative PCR (qPCR) for detection of the Treponema pallidum (TP) genome in clinical samples through simultaneous detection of two genomic targets. METHODS: We performed qPCR with TaqMan technology using two TP genes, polA and tpp47, as targets, with an internal positive control. The qPCR assay was compared with syphilis diagnosis based on a combination of clinical examination, serological results and inhouse nested PCR (nPCR). Samples were analysed at the National Reference Center for STIs at Cochin Hospital in Paris. RESULTS: In total, from October 2010 to December 2016, 320 documented clinical samples (mucosal and cutaneous swabs) were collected from patients with or without syphilis attending STI centres in France. The qPCR had an overall sensitivity of 89% (95% CI 85.1% to 92.1%), a specificity of 100%, a positive predictive value of 100% and a negative predictive value of 88% (95% CI 84.3% to 91.5%). The agreement between qPCR and nPCR results was 94% (κ=0.88, 95% CI 0.83 to 0.93). Calibration of the qPCR assay, by cloning both the polA and tpp47 genes, defined the detection threshold as 1 copy/µL of DNA elution. CONCLUSIONS: We validated a new qPCR for detecting the TP genome in clinical samples with excellent sensitivity and specificity. The cloning of polA and tpp47 genes for calibration would be interesting in the evaluation of bacterial loads in samples.

Characterization of a Cutibacterium acnes Camp Factor 1-Related Peptide as a New TLR-2 Modulator in In Vitro and Ex Vivo Models of Inflammation.

Cutibacterium acnes (C. acnes) has been implicated in inflammatory acne where highly mutated Christie-Atkins-Munch-Petersen factor (CAMP)1 displays strong toll like receptor (TLR)-2 binding activity. Using specific antibodies, we showed that CAMP1 production was independent of C. acnes phylotype and involved in the induction of inflammation. We confirmed that TLR-2 bound both mutated and non-mutated recombinant CAMP1, and peptide array analysis showed that seven peptides (A14, A15, B1, B2, B3, C1 and C3) were involved in TLR-2 binding, located on the same side of the three-dimensional structure of CAMP1. Both mutated and non-mutated recombinant CAMP1 proteins induced the production of C-X-C motif chemokine ligand interleukin (CXCL)8/(IL)-8 in vitro in keratinocytes and that of granulocyte macrophage-colony stimulating factor (GM-CSF), tumor necrosis factor (TNF)-α, IL-1ß and IL-10 in ex vivo human skin explants. Only A14, B1 and B2 inhibited the production of CXCL8/IL-8 by keratinocytes and that of (GM-CSF), TNF-α, IL-1ß and IL-10 in human skin explants stimulated with rCAMP1 and C. acnes. Following pretreatment with B2, RNA sequencing on skin explants identified the 10 genes displaying the strongest differential expression as IL6, TNF, CXCL1, CXCL2, CXCL3, CXCL8, IL-1ß, chemokine ligand (CCL)2, CCL4 and colony stimulating factor (CSF)2. We, thus, identified a new CAMP1-derived peptide as a TLR-2 modulator likely to be a good candidate for clinical evaluation.

Cutibacterium acnes as an Opportunistic Pathogen: An Update of Its Virulence-Associated Factors.

Cutibacterium acnes is a member of the skin microbiota found predominantly in regions rich in sebaceous glands. It is involved in maintaining healthy skin and has long been considered a commensal bacterium. Its involvement in various infections has led to its emergence as an opportunist pathogen. Interactions between C. acnes and the human host, including the human skin microbiota, promote the selection of C. acnes strains capable of producing several virulence factors that increase inflammatory capability. This pathogenic property may be related to many infectious mechanisms, such as an ability to form biofilms and the expression of putative virulence factors capable of triggering host immune responses or enabling C. acnes to adapt to its environment. During the past decade, many studies have identified and characterized several putative virulence factors potentially involved in the pathogenicity of this bacterium. These virulence factors are involved in bacterial attachment to target cells, polysaccharide-based biofilm synthesis, molecular structures mediating inflammation, and the enzymatic degradation of host tissues. C. acnes, like other skin-associated bacteria, can colonize various ecological niches other than skin. It produces several proteins or glycoproteins that could be considered to be active virulence factors, enabling the bacterium to adapt to the lipophilic environment of the pilosebaceous unit of the skin, but also to the various organs it colonizes. In this review, we summarize current knowledge concerning characterized C. acnes virulence factors and their possible implication in the pathogenicity of C. acnes.

Surveillance of Antibiotic Resistance Genes in Treponema Pallidum Subspecies Pallidum from Patients with Early Syphilis in France.

Benzathine penicillin G (BPG) is the reference treatment for early syphilis, but shortages have recently been reported, highlighting a need for the validation of alternative treatments. The aim of this study was to evaluate the genomic resistance of Treponema pallidum subspecies pallidum (TPA) to macrolides and doxycycline in France. Swabs from genital, anal, oral and cutaneous lesions were obtained from 146 patients with early syphilis in France. They were screened for mutations conferring resistance to macrolides and doxycycline by nested PCR and sequencing. Resistance to macrolides was detected in 85% of the isolates, but no point mutations conferring doxycycline resistance were detected. These findings confirm that, in France, resistance to macrolides is widespread. Moreover, we confirmed the absence of genomic resistance to doxycycline in the TPA strains. Therefore, doxycycline could be safely recommended as an alternative to BPG for the treatment of early syphilis.

Multi-locus sequence typing of Treponema pallidum subsp. pallidum present in clinical samples from France: Infecting treponemes are genetically diverse and belong to 18 allelic profiles.

Treponema pallidum subsp. pallidum, the causative agent of sexually transmitted syphilis, detected in clinical samples from France, was subjected to molecular typing using the recently developed Multilocus Sequence Typing system. The samples (n = 133) used in this study were collected from 2010-2016 from patients with diagnosed primary or secondary syphilis attending outpatient centers or hospitals in several locations in France. Altogether, 18 different allelic profiles were found among the fully typed samples (n = 112). There were five allelic variants identified for TP0136, 12 for TP0548, and eight for TP0705. Out of the identified alleles, one, seven, and three novel alleles were identified in TP0136, TP0548, and TP0705, respectively. Partial allelic profiles were obtained from 6 samples. The majority of samples (n = 110) belonged to the SS14-like cluster of TPA isolates while 7 clustered with Nichols-like isolates. Patients infected with Nichols-like samples were more often older (p = 0.041) and more often diagnosed with secondary syphilis (p = 0.033) compared to patients infected with SS14-like samples. In addition, macrolide resistance caused by the A2058G mutation was found to be associated with allelic profile 1.3.1 or with strains belonging to the 1.3.1 lineage (p<0.001). The genetic diversity among TPA strains infecting the European population was surprisingly high, which suggests that additional studies are needed to reveal the full genetic diversity of TPA pathogens infecting humans.

Human Treponema pallidum 11q/j isolate belongs to subsp. endemicum but contains two loci with a sequence in TP0548 and TP0488 similar to subsp. pertenue and subsp. pallidum, respectively.

BACKGROUND: Treponema pallidum subsp. endemicum (TEN) is the causative agent of endemic syphilis (bejel). An unusual human TEN 11q/j isolate was obtained from a syphilis-like primary genital lesion from a patient that returned to France from Pakistan. METHODOLOGY/PRINCIPAL FINDINGS: The TEN 11q/j isolate was characterized using nested PCR followed by Sanger sequencing and/or direct Illumina sequencing. Altogether, 44 chromosomal regions were analyzed. Overall, the 11q/j isolate clustered with TEN strains Bosnia A and Iraq B as expected from previous TEN classification of the 11q/j isolate. However, the 11q/j sequence in a 505 bp-long region at the TP0488 locus was similar to Treponema pallidum subsp. pallidum (TPA) strains, but not to TEN Bosnia A and Iraq B sequences, suggesting a recombination event at this locus. Similarly, the 11q/j sequence in a 613 bp-long region at the TP0548 locus was similar to Treponema pallidum subsp. pertenue (TPE) strains, but not to TEN sequences. CONCLUSIONS/SIGNIFICANCE: A detailed analysis of two recombinant loci found in the 11q/j clinical isolate revealed that the recombination event occurred just once, in the TP0488, with the donor sequence originating from a TPA strain. Since TEN Bosnia A and Iraq B were found to contain TPA-like sequences at the TP0548 locus, the recombination at TP0548 took place in a treponeme that was an ancestor to both TEN Bosnia A and Iraq B. The sequence of 11q/j isolate in TP0548 represents an ancestral TEN sequence that is similar to yaws-causing treponemes. In addition to the importance of the 11q/j isolate for reconstruction of the TEN phylogeny, this case emphasizes the possible role of TEN strains in development of syphilis-like lesions.

TLR-2 Recognizes Propionibacterium acnes CAMP Factor 1 from Highly Inflammatory Strains.

BACKGROUND: Propionibacterium acnes (P. acnes) is an anaerobic, Gram-positive bacteria encountered in inflammatory acne lesions, particularly in the pilosebaceous follicle. P. acnes triggers a strong immune response involving keratinocytes, sebocytes and monocytes, the target cells during acne development. Lipoteicoic acid and peptidoglycan induce the inflammatory reaction, but no P. acnes surface protein interacting with Toll-like receptors has been identified. P. acnes surface proteins have been extracted by lithium stripping and shown to induce CXCL8 production by keratinocytes. METHODOLOGY AND PRINCIPAL FINDINGS: Far-western blotting identified two surface proteins, of 24.5- and 27.5-kDa in size, specifically recognized by TLR2. These proteins were characterized, by LC-MS/MS, as CAMP factor 1 devoid of its signal peptide sequence, as shown by N-terminal sequencing. Purified CAMP factor 1 induces CXCL8 production by activating the CXCL8 gene promoter, triggering the synthesis of CXCL8 mRNA. Antibodies against TLR2 significantly decreased the CXCL8 response. For the 27 P. acnes strains used in this study, CAMP1-TLR2 binding intensity was modulated and appeared to be strong in type IB and II strains, which produced large amounts of CXCL8, whereas most of the type IA1 and IA2 strains presented little or no CAMP1-TLR2 binding and low levels of CXCL8 production. The nucleotide sequence of CAMP factor displays a major polymorphism, defining two distinct genetic groups corresponding to CAMP factor 1 with 14 amino-acid changes from strains phylotyped II with moderate and high levels of CAMP1-TLR2 binding activity, and CAMP factor 1 containing 0, 1 or 2 amino-acid changes from strains phylotyped IA1, IA2, or IB presenting no, weak or moderate CAMP1-TLR2 binding. CONCLUSIONS: Our findings indicate that CAMP factor 1 may contribute to P. acnes virulence, by amplifying the inflammation reaction through direct interaction with TLR2.

Clinical Aspects of Syphilis Reinfection in HIV-Infected Patients.

BACKGROUND: The incidence of HIV-syphilis co-infection has risen since 2000, especially among men having sex with men (MSM). Syphilis reinfection can occur, but the clinical features of such events remain poorly characterized. OBJECTIVE: To compare the cutaneous lesions seen with syphilis reinfections with those of first episodes in HIV-infected patients. METHODS: In a cohort of HIV-infected patients, syphilis reinfection was established both clinically and biologically by evaluating changes in Venereal Disease Research Laboratory titers. Photographs and medical records were studied in order to determine the type of skin lesions and their quantification. RESULTS: Among 533 HIV-infected patients, 42 (8%) experienced a first syphilis infection. Thirteen episodes of reinfection occurred in 12/42 (28%) patients, all MSM. In 78% of cases, reinfections were less symptomatic than first episodes. All patients presented classical syphilis lesions. CONCLUSIONS: We observed a high rate of reinfection, but with less severe skin manifestations during reinfection episodes.

Molecular subtyping of Treponema pallidum in Paris, France.

Two major Treponema pallidum subtypes, 14 d/g and 14 d/f, were identified in a population of 119 patients with syphilis in Paris, France, characterized by a high proportion of men who have sex with men. A new subtype named 11 q/j was characterized, and a reinfection case was determined in 1 patient having consecuitve syphilis infection at 19-month interval.

[Suicide: epidemiological data]. / Epidémiologie du suicide.

Within developed countries suicide rates in France are one of the highest, and are a matter of public health concern. In 2008 (latest available year) suicide figure is 10,313 (7,589 men and 2,724 women), which represents rates of 25.2 for men and 8.5 for women (16.6 p.100,000 persons). About 20% of suicide death are not registered. Suicide has increased since the mid 70'until the mid 80' and has declined since that period, but remains still higher than before for women (about 5% more). For baby-boomers the increase of suicide with age is more important until 50 years old, but seems less after. This new age-trend is becoming the same as in northern Europe and western countries. Suicide is prevalent by the age of 30 years old: suicide death is the first cause of death by that age, and suicide attempts are at the top incidence rate. Suicide is an important cause of premature mortality, and attempted suicide a frequent cause for hospitalization especially for young people (more than 180,000 each year, all ages). According to way of live, those who are alone are more at risk, especially the widowers. Mental diseases are prevalent in past life for suicide patients. Risk for those persons is higher, and each year about 1% of suicide attempters make suicide.

Cancer mortality in patients with schizophrenia: an 11-year prospective cohort study.

BACKGROUND: Schizophrenia has been associated with a rate of premature mortality that is 2 to 3 times higher than that in the general population. Although the role of cancer in this excess mortality remains unclear, previous incidence or mortality studies found contradictory results. METHODS: In 1993, a large prospective study was initiated in a cohort of 3470 patients with schizophrenia to examine cancer-related mortality and predictors. Standardized mortality ratios (SMRs) were calculated, adjusting for age and sex relative to a representative sample of the French general population. RESULTS: During the 11-year follow-up, 476 (14%) patients died; the mortality rate was thus nearly 4-fold higher than in the general population. Cancer was the second most frequent cause of mortality (n=74), with a global SMR of 1.5 (95% confidence interval [95% CI], 1.2-1.9). For all cancers, the SMRs were 1.4 (not significant) for men and 1.9 (95% CI, 1.4-2.8) for women. For men, lung cancer was the most frequent localization (n=23; 50%), with an SMR of 2.2 (95% CI, 1.6-3.3). For women, breast cancer was the most frequent localization (n=11; 39%), with an SMR of 2.8 (95% CI, 1.6-4.9). In comparison with patients who did not die of cancer, there were 2 significant baseline predictors of death by lung cancer in the final logistic regression model: duration of smoking and age>38 years. CONCLUSIONS: The results of the current study demonstrated an increased risk of mortality by cancer in patients with schizophrenia, especially for women from breast cancer and for men from lung cancer.

Ten-year prospective follow-up study of the mortality by suicide in schizophrenic patients.

This ten-year follow-up study examined the prevalence and the most relevant baseline predictors of suicide in schizophrenic patients. In 1993, 3470 patients meeting the ICD-10 criteria for schizophrenia were assessed. We used national death certificate data to identify patients that had died by suicide for each year included in the study. In this way, we calculated standardized mortality ratios, adjusting for age and sex relative to the general population. We used Cox's proportional hazards models to investigate potential sociodemographic and clinical risk factors. There were 141 suicides in the cohort during the follow-up period, corresponding to a risk of suicide that was approximately 16 times higher than that of the general population. Women had slightly higher standardized mortality ratios than men. Suicide was the cause of death in more than half (53.9%) of deaths occurring during the first year of follow-up and nearly one-third (31.8%) of those occurring in the ten-year period of the study. There were four significant baseline predictors of suicide remaining in the final logistic regression model: male gender, drug abuse, previous suicide attempts, and short duration of illness. Sex, age, history of suicide attempt should be particularly considered in the assessment of suicide risk in schizophrenic patients. Our findings also emphasize the need for detection and effective management of associated comorbid drug abuse.

Effect of telephone contact on further suicide attempts in patients discharged from an emergency department: randomised controlled study.

OBJECTIVE: To determine the effects over one year of contacting patients by telephone one month or three months after being discharged from an emergency department for deliberate self poisoning compared with usual treatment. DESIGN: Multicentre, randomised controlled trial. SETTING: 13 emergency departments in the north of France. PARTICIPANTS: 605 people discharged from an emergency department after attempted suicide by deliberate self poisoning. INTERVENTION: The intervention consisted of contacting patients by telephone at one month or three months after discharge from an emergency department for attempted suicide to evaluate the success of recommended treatment or to adjust treatment. Control patients received treatment as usual, in most cases referral back to their general practitioner. MAIN OUTCOME MEASURES: The primary outcome measures were proportion of participants who reattempted suicide, number of deaths by suicide, and losses to follow-up at 13 months' follow-up. Secondary outcome measures were types and number of contacts with health care. RESULTS: On an intention to treat basis, the three groups did not differ significantly for further suicide attempts, deaths by suicide, or losses to follow-up: contact at one month (intervention 23% (34/147) v controls 30% (93/312), difference 7%, 95% confidence interval - 2% to 15%), three months (25% (36/146) v 30%, difference 5%, - 4% to 14%). Participants contacted at one month were less likely at follow-up to report having reattempted suicide (12% v 22% in control group, difference 10%, 2% to 18%). CONCLUSION: Contacting people by telephone one month after being discharged from an emergency department for deliberate self poisoning may help reduce the number of reattempted suicides over one year.

Predictive factors of the number and the dose of anti-psychotics in a cohort of schizophrenic patients.

BACKGROUND: Anti-psychotic prescription in schizophrenia is characterised in Europe by frequent associations and high doses. Nevertheless, few longitudinal epidemiological studies have explored anti-psychotic prescriptions. AIM: (1) To describe the evolution of prescription patterns across time; (2) to determine risk factors for prescription of high doses of anti-psychotics and for anti-psychotic combinations. MATERIALS AND METHODS: Three thousand four hundred seventy three subjects were included in 1993. Data collected in 1993 and subsequently in 1996 and 1999, provided information on demographics, clinical status and prescription. In 1996, the response rate was 68.5% and 56.7% in 1999. RESULTS: The number of anti-psychotics prescribed slightly decreased across time, while doses remained high for one-third of the patients. The factors predicting dose were: dose at previous evaluation, class of anti-psychotic and clinical severity. The factors predicting the number of anti-psychotics were: previous number and class of anti-psychotic and clinical severity. CONCLUSION: Higher dosage and combinations were related more to physicians' habits than to patient characteristics, as is frequently observed in chronic disease.

Neuroleptics and mortality in schizophrenia: prospective analysis of deaths in a French cohort of schizophrenic patients.

OBJECTIVE: The putative role of neuroleptics in the known excess mortality of subjects with schizophrenia remains disputed. The aim of this study was to assess the link between mortality and the class of neuroleptic. METHOD: Causes of death (suicide, cardiovascular, etc.) and exposure to neuroleptics were studied in a cohort of 3474 patients with schizophrenia followed from 1993 to 1997. RESULTS: From 1993 to 1997, 178 patients died. The risk of all-cause death (OR=1.59; 95% CI 1.02-2.50; p=0.04), and suicide (OR=2.22; 95% CI 1.24-3.97; p=0.006) were increased in users of thioxanthenes (alone or associated with other drugs), and increased risk of "other causes" of death was associated with use of atypical neuroleptics (OR=2.06; 95% CI 1.15-3.70; p=0.0016). CONCLUSION: Our findings suggest the existence of association between certain classes of neuroleptics and death, all cause or specific. This could be related to the drug itself or to patient selection.