Identification and characterisation of microRNAs in young adults of Angiostrongylus cantonensis via a deep-sequencing approach

Angiostrongylus cantonensis is an important causative agent of eosinophilic meningitis and eosinophilic meningoencephalitis in humans. MicroRNAs (miRNAs) are small non-coding RNAs that participate in a wide range of biological processes. This study employed a deep-sequencing approach to study miRNAs from young adults of A. cantonensis. Based on 16,880,456 high-quality reads, 252 conserved mature miRNAs including 10 antisense miRNAs that belonging to 90 families, together with 10 antisense miRNAs were identified and characterised. Among these sequences, 53 miRNAs from 25 families displayed 50 or more reads. The conserved miRNA families were divided into four groups according to their phylogenetic distribution and a total of nine families without any members showing homology to other nematodes or adult worms were identified. Stem-loop real-time polymerase chain reaction analysis of aca-miR-1-1 and aca-miR-71-1 demonstrated that their level of expression increased dramatically from infective larvae to young adults and then decreased in adult worms, with the male worms exhibiting significantly higher levels of expression than female worms. These findings provide information related to the regulation of gene expression during the growth, development and pathogenesis of young adults of A. cantonensis.

Comparative clinical study of different multiplex real time PCR strategies for the simultaneous differential diagnosis between extrapulmonary tuberculosis and focal complications of brucellosis

Background: Both brucellosis and tuberculosis are chronic-debilitating systemic granulomatous diseases with a high incidence in many countries in Africa, Central and South America, the Middle East and the Indian subcontinent. Certain focal complications of brucellosis and extrapulmonary tuberculosis are very difficult to differentiate clinically, biologically and radiologically. As the conventional microbiological methods for the diagnosis of the two diseases have many limitations, as well as being time-consuming, multiplex real time PCR (M RT-PCR) could be a promising and practical approach to hasten the differential diagnosis and improve prognosis. Methodology/Principal Findings: We designed a SYBR Green single-tube multiplex real-time PCR protocol targeting bcsp31 and the IS711 sequence detecting all pathogenic species and biovars of Brucella genus, the IS6110 sequence detecting Mycobacterium genus, and the intergenic region senX3-regX3 specifically detecting Mycobacterium tuberculosis complex. The diagnostic yield of the M RT-PCR with the three pairs of resultant amplicons was then analyzed in 91 clinical samples corresponding to 30 patients with focal complications of brucellosis, 24 patients with extrapulmonary tuberculosis, and 36 patients (Control Group) with different infectious, autoimmune or neoplastic diseases. Thirty-five patients had vertebral osteomyelitis, 21 subacute or chronic meningitis or meningoencephalitis, 13 liver or splenic abscess, eight orchiepididymitis, seven subacute or chronic arthritis, and the remaining seven samples were from different locations. Of the three pairs of amplicons (senX3-regX3+ bcsp3, senX3-regX3+ IS711 and IS6110+ IS711) only senX3-regX3+ IS711 was 100% specific for both the Brucella genus and M. tuberculosis complex. For all the clinical samples studied, the overall sensitivity, specificity, and positive and negative predictive values of the M RT-PCR assay were 89.1%, 100%, 85.7% and 100%, respectively, with an accuracy of 93.4%, (95% CI, 88.3—96.5%). Conclusions/Significance: In this study, a M RT-PCR strategy with species-specific primers based on senX3-regX3+IS711 sequences proved to be a sensitive and specific test, useful for the highly efficient detection of M. tuberculosis and Brucella spp in very different clinical samples. It thus represents an advance in the differential diagnosis between some forms of extrapulmonary tuberculosis and focal complications of brucellosis.

Infeccions del sistema nerviós central pel virus varicel.la-zoster

Una associació inusual de casos d’infecció del SNC pel VVZ en un periode de 16 mesos i l’absència de lesions cutànies en la majoria d’ells ens va portar a revisar els casos d’infecció del SNC pel VVZ al nostre centre en els darrers 6 anys. Hem trobat 5 meningitis, 5 encefalitis i 2 mielitis. Les meningitis afecten a joves no immunodeprimits i solen cursar sense lesions d’herpes zòster. A més, sovint cursen amb hipoglucorràquia, el que inicialment pot fer pensar en una meningitis aguda bacteriana. Les encefalitis afecten a pacients més grans, immunodeprimits i s’acompanyen de lesions d’herpes zòster.

Cryptococcosis in Colombian children and literature review

Cryptococcosis is reported in adults and is often acquired immune deficiency syndrome (AIDS)-associated; however, its frequency in children is low. Based on the National Survey on Cryptococcosis conducted in Colombia, an epidemiological and clinical analysis was performed on cases of the disease observed in children less than 16 years old between 1993-2010. We found 41 affected children (2.6% prevalence) from the 1,578 surveys received. The country mean annual incidence rate was 0.017 cases/100,000 children under 16 years, while in Norte de Santander the incidence rate was 0.122 cases/100,000 (p < 0.0001). The average age of infected children was 8.4 and 58.5% were male. In 46.3% of cases, a risk factor was not identified, while 24.4% had AIDS. The most frequent clinical manifestations were headache (78.1%), fever (68.8%), nausea and vomiting (65.6%), confusion (50%) and meningeal signs (37.5%). Meningitis was the most frequent clinical presentation (87.8%). Amphotericin B was given to 93.5% of patients as an initial treatment. Positive microbiological identification was accomplished by India ink (94.7%), latex in cerebrospinal fluid (100%) and culture (89.5%). Out of 34 isolates studied, Cryptococcus neoformans var. grubii (VNI 85.3%, VNII 8.8%) was isolated in 94.1% of cases and Cryptococcus gattii (VGII) was isolated in 5.9% of cases. These data are complemented by a literature review, which overall suggests that cryptococcosis in children is an unusual event worldwide.

Postoperative epidural hematoma contributes to delayed upper cord tethering after decompression of Chiari malformation type I.

Symptomatic arachnoiditis after posterior fossa surgical procedures such as decompression of Chiari malformation is a possible complication. Clinical presentation is generally insidious and delayed by months or years. It causes disturbances in the normal flow of cerebrospinal fluid and enlargement of a syrinx cavity in the upper spinal cord. Surgical de-tethering has favorable results with progressive collapse of the syrinx and relief of the associated symptoms. Case Description: A 30-year-old male with Chiari malformation type I was treated by performing posterior fossa bone decompression, dura opening and closure with a suturable bovine pericardium dural graft. Postoperative period was uneventful until the fifth day in which the patient suffered intense headache and progressive loose of consciousness caused by an acute posterior fossa epidural hematoma. It was quickly removed with complete clinical recovering. One year later, the patient experienced progressive worsened of his symptoms. Upper spinal cord tethering was diagnosed and a new surgery for debridement was required. Conclusions: The epidural hematoma compressing the dural graft against the neural structures contributes to the upper spinal cord tethering and represents a nondescribed cause of postoperative fibrosis, adhesion formation, and subsequent recurrent hindbrain compression.

Comparison of polyethylene wear in anatomical and reversed shoulder prostheses.

Wear of polyethylene is associated with aseptic loosening of orthopaedic implants and has been observed in hip and knee prostheses and anatomical implants for the shoulder. The reversed shoulder prostheses have not been assessed as yet. We investigated the volumetric polyethylene wear of the reversed and anatomical Aequalis shoulder prostheses using a mathematical musculoskeletal model. Movement and joint stability were achieved by EMG-controlled activation of the muscles. A non-constant wear factor was considered. Simulated activities of daily living were estimated from in vivo recorded data. After one year of use, the volumetric wear was 8.4 mm(3) for the anatomical prosthesis, but 44.6 mm(3) for the reversed version. For the anatomical prosthesis the predictions for contact pressure and wear were consistent with biomechanical and clinical data. The abrasive wear of the polyethylene in reversed prostheses should not be underestimated, and further analysis, both experimental and clinical, is required.

Résistance aux antibiotiques chez les pneumocoques [Pneumococcal antibiotic resistance].

In 1875, 7 years prior to the description of the Koch bacillus, Klebs visualized the first Streptococcus pneumoniae in a pleural fluid. Since then, this organism has played a determinant role in biomedical science. From a biological point of view, it was largely implicated in the development of passive and active immunization by serotherapy and vaccination, respectively. Genetic transformation was also first observed in S. pneumoniae, leading to the discovery of DNA. From a clinical point of view, S. pneumoniae is still today a prime cause of otitis media in children and of pneumonia in all age groups, as well as a predominant cause of meningitis and bacteremia. In adults, bacteremia is still entailed with a mortality of over 25%. Although S. pneumoniae remained very sensitive to penicillin for many years, penicillin-resistance has emerged and increased dramatically over the last 15 years. During this period of time, the frequency of penicillin-resistant isolates has increased from &lt; or = 1% to frequencies varying from 20 to 60% in geographic areas as diverse as South Africa, Spain, France, Hungary, Iceland, Alaska, and numerous regions of the United States and South America. In Switzerland, the current frequency of penicillin-resistant pneumococci ranges between 5 and &gt; or = 10%. The increase in penicillin-resistant pneumococci correlates with the intensive use of beta-lactam antibiotics. The mechanism of resistance is not due to bacterial production of penicillinase, but to an alteration of the bacterial target of penicillin, the so-called penicillin-binding proteins. Resistance is subdivided into (i) inter mediate level resistance (minimal inhibitory concentration [MIC] of penicillin of 0.1-1 mg/L) and (ii) high level resistance (MCI &gt; or = 2 mg/L). The clinical significance of intermediate resistance remains poorly defined. On the other hand, highly resistant strains were responsible for numerous therapeutical failures, especially in cases of meningitis. Antibiotics recommended against penicillin-resistant pneumococci include cefotaxime, ceftriaxone, imipenem and in some instances vancomycin. However, penicillin-resistant pneumococci tend to present cross-resistances to all the antibotics of the beta-lactam family and could even become resistant to the last resort drugs mentioned above. Thus, in conclusion, the explosion of resistance to penicillin in pneumococci is a ubiquitous phenomenon which must be fought against by (i) a strict utilization of antibiotics, (ii) the practice of microbiological sampling of infected foci before treatment, (iii) the systematic surveillance of resistance profiles of pneumococci against antibiotics and (iv) the adequate vaccination of populations at risk.

Dynamic regulation of notch 1 and notch 2 surface expression during T cell development and activation revealed by novel monoclonal antibodies.

It is well established that Notch signaling plays a critical role at multiple stages of T cell development and activation. However, detailed analysis of the cellular and molecular events associated with Notch signaling in T cells is hampered by the lack of reagents that can unambiguously measure cell surface Notch receptor expression. Using novel rat mAbs directed against the extracellular domains of Notch1 and Notch2, we find that Notch1 is already highly expressed on common lymphoid precursors in the bone marrow and remains at high levels during intrathymic maturation of CD4(-)CD8(-) thymocytes. Notch1 is progressively down-regulated at the CD4(+)CD8(+) and mature CD4(+) or CD8(+) thymic stages and is expressed at low levels on peripheral T cells. Immunofluorescence staining of thymus cryosections further revealed a localization of Notch1(+)CD25(-) cells adjacent to the thymus capsule. Notch1 was up-regulated on peripheral T cells following activation in vitro with anti-CD3 mAbs or infection in vivo with lymphocytic chorio-meningitis virus or Leishmania major. In contrast to Notch1, Notch2 was expressed at intermediate levels on common lymphoid precursors and CD117(+) early intrathymic subsets, but disappeared completely at subsequent stages of T cell development. However, transient up-regulation of Notch2 was also observed on peripheral T cells following anti-CD3 stimulation. Collectively our novel mAbs reveal a dynamic regulation of Notch1 and Notch2 surface expression during T cell development and activation. Furthermore they provide an important resource for future analysis of Notch receptors in various tissues including the hematopoietic system.

False-negative PCR result due to gene polymorphism: the example of Neisseria meningitidis.

Early treatment of meningococcal meningitis is mandatory but may negate the cerebrospinal fluid culture. Etiological diagnosis then mainly relies on PCR. Here, we report a case of false-negative results for real-time PCR for a Neisseria meningitidis serogroup B isolate with a polymorphism in the ctrA gene.

Total knee arthroplasty - a clinical and numerical study of the micromovements of the tibial implant

Introduction The importance of the micromovements in the mechanism of aseptic loosening is clinically difficult to evaluate. To complete the analysis of a series of total knee arthroplasties (TKA), we used a tridimensional numerical model to study the micromovements of the tibial implant.Material and Methods Fifty one patients (with 57 cemented Porous Coated Anatomic TKAs) were reviewed (mean follow-up 4.5 year). Radiolucency at the tibial bone-cement interface was sought on the AP radiographs and divided in 7 areas. The distribution of the radiolucency was then correlated with the axis of the lower limb as measured on the orthoradiograms.The tridimensional numerical model is based on the finite element method. It allowed the measurement of the cemented prosthetic tibial implant's displacements and the microvements generated at bone-ciment interface. A total load (2000 Newton) was applied at first vertically and asymetrically on the tibial plateau, thereby simulating an axial deviation of the lower limbs. The vector's posterior inclination then permitted the addition of a tangential component to the axial load. This type of effort is generated by complex biomechanical phenomena such as knee flexion.Results 81 per cent of the 57 knees had a radiolucent line of at least 1 mm, at one or more of the tibial cement-epiphysis jonctional areas. The distribution of these lucent lines showed that they came out more frequently at the periphery of the implant. The lucent lines appeared most often under the unloaded margin of the tibial plateau, when axial deviation of lower limbs was present.Numerical simulations showed that asymetrical loading on the tibial plateau induced a subsidence of the loaded margin (0-100 microns) and lifting off at the opposite border (0-70 microns). The postero-anterior tangential component induced an anterior displacement of the tibial implant (160-220 microns), and horizontal micromovements with non homogenous distribution at the bone-ciment interface (28-54 microns).Discussion Comparison of clinical and numerical results showed a relation between the development of radiolucent lines and the unloading of the tibial implant's margin. The deleterious effect of lower limbs' axial deviation is thereby proven. The irregular distribution of lucent lines under the tibial plateau was similar of the micromovements' repartition at the bone-cement interface when tangential forces were present. A causative relation between the two phenomenaes could not however be established.Numerical simulation is a truly useful method of study; it permits to calculate micromovements which are relative, non homogenous and of very low amplitude. However, comparative clinical studies remain as essential to ensure the credibility of results.

Tigecycline in combination with other antimicrobials: a review of in vitro, animal and case report studies.

Tigecycline has been investigated in combination with other antibacterials against a wide range of susceptible and multiresistant Gram-positive and Gram-negative bacteria. Combinations have been analysed in vitro, in animal models and in human case reports. In vitro, tigecycline combined with other antimicrobials produces primarily an indifferent response (neither synergy nor antagonism). Nevertheless, synergy occurred when tigecycline was combined with rifampicin against 64-100% of Enterococcus spp., Streptococcus pneumoniae, Enterobacter spp. and Brucella melitensis isolates. Combinations of tigecycline with amikacin also showed synergy for 40-100% of Enterobacter spp., Klebsiella pneumoniae, Proteus spp. and Stenotrophomonas maltophilia isolates. Moreover, bactericidal synergisms occurred with tigecycline plus amikacin against problematic Acinetobacter baumannii and Proteus vulgaris, and with colistin against K. pneumoniae. Data from animal experiments and case reports, although limited, displayed consistent beneficial activity of tigecycline in combination with other antibacterials against multiresistant organisms, including vancomycin against penicillin-resistant S. pneumoniae in experimental meningitis, gentamicin against Pseudomonas aeruginosa in experimental pneumonia, daptomycin against Enterococcus faecium endocarditis, and colistin against K. pneumoniae bacteraemia and P. aeruginosa osteomyelitis. Antagonism was extremely rare in vitro and was not reported in vivo. Thus, tigecycline may be combined with a second antimicrobial as part of a combination regimen.

Varicella Zoster Virus CNS disease in hematopoietic cell transplantation: A single center experience

Background: Varciella Zoster Virus (VZV) can lead to serious complications in Hematopoietic Cell Transplant (HCT) recipients. Central nervous system (CNS) VZV can be one of the most devastating infections in transplant recipients, yet little is known about this rare disease. Objectives: To describe CNS VZV in the post-transplant period and to define potential risk factors in the HCT population. Methods: We reviewed the course of all patients who received a first HCT at the Fred Hutchinson Cancer Center (FHCRC) in Seattle, WA from 1/1996 through 12/2007. Data were collected retrospectively using the Long-Term Follow-Up database, which includes on-site examinations, outside records, laboratory tests, and yearly questionnaires. Patients were classified as CNS VZV if they had laboratory confirmation of VZV in the cerebrospinal fluid (CSF), or had zoster with associated clinical and laboratory findings consistent with CNS disease. Results: A total of six patients developed VZV CNS disease during the evaluation period (table 1). Diagnosis was confirmed in 3/6 by detection of VZV in CSF by PCR. All other patients had a clinical diagnosis based on the presence of CNS symptoms, zoster, lymphocytic pleiocytosis, and response to IV acyclovir. Patients who developed CNS disease had a mean age of 42 years (range 34-51) at time of transplant. CNS disease developed at a mean of 9 months posttransplantation (range 0.5-24 months), and severity varied, ranging from meningitis (3/6) to encephalitis/myelitis (3/6). All had active graft-versus host disease (GHVD) and all were being treated with immunosuppressive therapy at time of diagnosis. Fever and headache were the most common symptoms, but patients who developed focal CNS findings or seizures (3/6) had a more complicated clinical course. While most patients presented with classic VZV/zoster skin lesions, 2/6 patients had no dermatologic findings associated with their presentation. Four (66%) of patients who developed VZV CNS disease died, two related to VZV complications despite aggressive antiviral therapy. Conclusions: In this cohort of HCT patients, VZV CNS disease was a rare complication. Mortality due to CNS VZV is high, particularly in patients who develop focal neurologic findings or seizures. Even in the absence of skin lesions, VZV CNS disease should be considered in patients who develop fevers and neurologic symptoms.

Reversed shoulder arthroplasty baseplate fixed according to the three major columns principle

Introduction: Glenoid bone volume and bone quality can render the fixation of a reversed shoulder arthroplasty (RSA) basis plate hazardous. Cadaveric study at our institution has demonstrated that optimal baseplate fixation could be achieved with screws in three major columns. Our aim is to review our early rate of aseptic glenoid loosening in a series of baseplates fixed according to this principle. Methods: Between 2005 and 2008, 48 consecutive RSA (Reversed Aequalis) were implanted in 48 patients with an average age of 74.4 years (range, 56 to 86 years). There were 37 women and 11 men. Twenty-seven primary RSAs were performed for cuff tear arthropathy, 3 after failed rotator cuff surgery, 6 for failed arthroplasties, 7 for acute fractures and 5 after failed ORIF. All baseplate fixations were done using a nonlocking posterior screw in the scapular spine, a nonlocking anterior screw in the glenoid body, a locking superior screw in the coracoid and a locking inferior screw in the pillar. All patients were reviewed with standardized radiographs. We reported the positions of the screws in relation to the scapular spine and the coracoid process in two different views. We defined screw positions as totally, partially or out of the target. Finally, we reported aseptic glenoid loosening which was defined as implant subsidence. Results: Four patients were lost to follow-up. Thus 44 shoulders could be reviewed after a mean follow-up of 16 months (range, 9 to 32 months). Thirty-seven (84%) screws were either partially or totally in the spine. Thus, 7 (16%) scapular spine screws were out of the target. No coracoid screw was out of the target. At final follow-up control, we reported no glenoid loosening. Conclusion: Early glenoid loosening occurred before the two years follow-up and is most of time related to technical problems and/or insufficient glenoid bone stock and bone quality. Our study demonstrate that baseplate fixation of a RSA according to the three columns principle is a reproducible technique and a valuable way to prevent early glenoid loosening.

Cerebrospinal fluid PCR analysis and biochemistry in bodies with severe decomposition.

The aim of this study was to assess whether Neisseria meningitidis, Listeria monocytogenes, Streptococcus pneumoniae and Haemophilus influenzae can be identified using the polymerase chain reaction technique in the cerebrospinal fluid of severely decomposed bodies with known, noninfectious causes of death or whether postmortem changes can lead to false positive results and thus erroneous diagnostic information. Biochemical investigations, postmortem bacteriology and real-time polymerase chain reaction analysis in cerebrospinal fluid were performed in a series of medico-legal autopsies that included noninfectious causes of death with decomposition, bacterial meningitis without decomposition, bacterial meningitis with decomposition, low respiratory tract infections with decomposition and abdominal infections with decomposition. In noninfectious causes of death with decomposition, postmortem investigations failed to reveal results consistent with generalized inflammation or bacterial infections at the time of death. Real-time polymerase chain reaction analysis in cerebrospinal fluid did not identify the studied bacteria in any of these cases. The results of this study highlight the usefulness of molecular approaches in bacteriology as well as the use of alternative biological samples in postmortem biochemistry in order to obtain suitable information even in corpses with severe decompositional changes.

Infection of the central nervous system caused by varicella zoster virus reactivation: a retrospective case series study.

BACKGROUND: Recent data suggest that varicella zoster virus (VZV)-associated complications of the central nervous system (CNS) are more common and diverse than previously thought. The main purpose of this article is to describe the clinical characteristics and the outcome of patients suffering from meningitis and encephalitis caused by VZV reactivation. METHODS: A retrospective case study of adult patients (≥16 years old) diagnosed with a VZV reactivation in the CNS was performed. The cases were identified by a qualitative PCR DNA assay of the cerebrospinal fluid (CSF) at the Regional Hospital of Lugano between January 1, 2003 and July 31, 2010. RESULTS: Eleven out of 519 CSF samples (2.1%), submitted from patients with a clinical diagnosis of viral meningitis or encephalitis, were positive for VZV. A vesiculo-pustular skin eruption was observed in only five patients (45%). In six cases (55%), a systemic inflammatory syndrome was absent. The clinical outcome was favorable in eight patients (73%). Only one out of 11 patients (9%) died. The four patients with encephalitis had a less favorable prognosis: one patient recovered without residual neurological sequelae; two had a chronic neuropsychological handicap, speech difficulties, facial nerve palsy, and focal seizures; one patient died. We estimated an annual incidence rate of VZV infection of the CNS of 1.02/100 000 inhabitants for southern Switzerland. CONCLUSIONS: Screening of CSF for VZV by PCR is recommended for all patients with encephalitis and for those with viral meningitis of unclear origin in order to better target antiviral treatment.