Impaired autophagy in macrophages promotes inflammatory eye disease

Autophagy is critical for maintaining cellular homeostasis. Organs such as the eye and brain are immunologically privileged. Here, we demonstrate that autophagy is essential for maintaining ocular immune privilege. Deletion of multiple autophagy genes in macrophages leads to an inflammation-mediated eye disease called uveitis that can cause blindness. Loss of autophagy activates inflammasome-mediated IL1B secretion that increases disease severity. Inhibition of caspase activity by gene deletion or pharmacological means completely reverses the disease phenotype. Of interest, experimental uveitis was also increased in a model of Crohn disease, a systemic autoimmune disease in which patients often develop uveitis, offering a potential mechanistic link between macrophage autophagy and systemic disease. These findings directly implicate the homeostatic process of autophagy in blinding eye disease and identify novel pathways for therapeutic intervention in uveitis.     

Effect of temperature on defense parameters in manila clam Ruditapes philippinarum challenged with Vibrio tapetis

Brown Ring Disease (BRD), a vibriosis affecting the clam Ruditapes philippinarum, is present on the Atlantic coasts of Western Europe and is considered to be a cold water disease. The present work investigated the effect of temperature on immune response and its relationships with BRD development. Clams maintained at different temperatures (8, 14 and 21 degrees C) were experimentally challenged with the pathogen Vibrio tapetis, the etiologic agent of BRD. Results demonstrated significant effects of temperature on disease development and on hemolymph immune parameters including total and viable hemocyte counts, lysozyme and leucine aminopeptidase activities. Thirty days after challenge, clams maintained at 21 degrees C displayed significantly higher values for all the measured immune parameters in comparison to specimens incubated at 14 degrees C. Improved performance of the immune system was associated with a low BRD prevalence. The recovery process, which occured mainly at 21 degrees C, was associated with high percentages of viable hemocytes and high activities of leucine amino-peptidase and lysozyme. This laboratory study clearly demonstrates that temperature strongly affects BRD development and clam immune response during infection. Favourable immune status at higher temperature may confer upon the clam a better capacity to fight the disease agent, and therefore to recover more easily.     

Genetic diversity assessment of Tunisian <Emphasis Type="Italic">Mycobacterium bovis</Emphasis> population isolated from cattle

Background

The genetic diversity of M. bovis in Tunisia is still underestimated despite the implementation of an eradication program. The lack of data about spatial distribution of the M. bovis population hinders the control of bovine tuberculosis (bTB) progress. This study represents the largest molecular analysis of M. bovis isolates in Tunisia. It is aimed to upgrade the understanding of bTB epidemiology and the geographical distribution of the infection. Tuberculosis research was performed in cattle (n?=?149) with TB-compatible lesions collected over 5 months from a slaughterhouse located in Sfax, Tunisia.

Results

Ninety-four animals were found to be infected by M. bovis and two others by M. caprae. Spoligotyping revealed twenty-five patterns, SB0120, SB0134, and SB0121 being the most prevalent profiles (36.4%, 11.4%, and 7.2%, respectively). Three new spoligotypes were detected: SB2345, SB2344 and SB2343. MIRU-VNTR analysis classified the isolates in seventy-three profiles and showed a large genotypic variety observed within the main spoligotype which was split into several MIRU-VNTR types: 29 in SB0120 (h?=?0.983), 10 in SB0134 (h?=?0.981) and 7 in SB0121 (h?=?1). Genotyping revealed a common pattern in different geographic regions. It also showed that Sfax, located in southern-Tunisia, represents a high-risk area with an elevated genetic diversity.

Conclusions

Spatial analysis may provide insights into disease transmission, which affects the effectiveness of eradication campaigns in cattle.

     

Evaluation of an <Emphasis Type="Italic">in silico</Emphasis> predicted specific and immunogenic antigen from the OmcB protein for the serodiagnosis of <Emphasis Type="Italic">Chlamydia trachomatis</Emphasis> infections

Background  

The OmcB protein is one of the most immunogenic proteins in C. trachomatis and C. pneumoniae infections. This protein is highly conserved leading to serum cross reactivity between the various chlamydial species. Since previous studies based on recombinant proteins failed to identify a species specific immune response against the OmcB protein, this study evaluated an in silico predicted specific and immunogenic antigen from the OmcB protein for the serodiagnosis of C. trachomatis infections.     

Broad-range PCR,cloning and sequencing of the full 16S rRNA gene for detection of bacterial DNA in synovial fluid samples of Tunisian patients with reactive and undifferentiated arthritis

Introduction

Broad-range rDNA PCR provides an alternative, cultivation-independent approach for identifying bacterial DNA in reactive and other form of arthritis. The aim of this study was to use broad-range rDNA PCR targeting the 16S rRNA gene in patients with reactive and other forms of arthritis and to screen for the presence of DNA from any given bacterial species in synovial fluid (SF) samples.

Methods

We examined the SF samples from a total of 27 patients consisting of patients with reactive arthritis (ReA) (n = 5), undifferentiated arthritis (UA) (n = 9), rheumatoid arthritis (n = 7), and osteoarthritis (n = 6) of which the latter two were used as controls. Using broad-range bacterial PCR amplifying a 1400 bp fragment from the 16S rRNA gene, we identified and sequenced at least 24 clones from each SF sample. To identify the corresponding bacteria, DNA sequences were compared to the EMBL (European Molecular Biology Laboratory) database.

Results

Bacterial DNA was identified in 20 of the 27 SF samples (74, 10%). Analysis of a large number of sequences revealed the presence of DNA from more than one single bacterial species in the SF of all patients studied. The nearly complete sequences of the 1400 bp were obtained for most of the detected species. DNA of bacterial species including Shigella species, Escherichia species, and other coli-form bacteria as well as opportunistic pathogens such as Stenotrophomonas maltophilia and Achromobacter xylosoxidans were shared in all arthritis patients. Among pathogens described to trigger ReA, DNA from Shigella sonnei was found in ReA and UA patients. We also detected DNA from rarely occurring human pathogens such as Aranicola species and Pantoea ananatis. We also found DNA from bacteria so far not described in human infections such as Bacillus niacini, Paenibacillus humicus, Diaphorobacter species and uncultured bacterium genera incertae sedis OP10.

Conclusions

Broad-range PCR followed by cloning and sequencing the entire 16S rDNA, allowed the identification of the bacterial DNA environment in the SF samples of arthritic patients. We found a wide spectrum of bacteria including those known to be involved in ReA and others not previously associated with arthritis.     

Retentissement de l’infection genitale a chlamydia trachomatis sur le sperme chez les hommes consultant pour infertilite du couple

Chlamydia trachomatis (CT) genital infection is one of the most frequent causes of infertility. Its repercution on semen parameters and male infertility is controversial. The objective of this study was to evaluate the impact of CT genital infection on semen parameters in male partners of infertile couples. Ninety-seven infertile couples were studied. Semen, urethral and cervical samples were tested for CT by means of direct fluorescence antibodies assay (DFA), cell culture, polymerase chain reaction (PCR) and FLISA. Sera from both parteners were tested for immunoglobulin M, A and G antibodies to Chlamydia by means of the microimmunofluorescence MIF). For all mens, standard semen parameters were analysed according to the guidlines of the word health organisation. CT infection was identified in 34% of the male partners. In 76% of cases, the infection was asymptomatic. 60,6% of infected patients’s wives were also infected by CT. There was no significant difference between the mean values of concentration, motility and morphology of spermatozoa in both groups of male patients, infected by CT (CT+ group) and lacked infection (CT-group). The mean values of motility, vitality, concentration and normal forms of spermatozoa, in both CT+ and CT- groups were respectively: 39,6%±17,5% vs 40,4% ± 14,9%, 61,9% ±18,1% vs 62,4% ± 18,5%, 80,7×10⁶±67,5×10⁶ vs 67,1×10⁶ ±65,2×10⁶ and 34,7% ± 16,7% vs 33% ± 0,1%. Oligospermia was significantly more frequent in CT+ group (54,9%) than in CT-group (26,9%). High levels of coiled flagella (≥20) were more frequently observed in CT+ group (18,5%) than in CT-group (7,4%), but the difference was not significant. We found in this study a high prevalence of genital chlamydial infection into infertile couples. This infection has no repercution on sperm quality, suggesting that there is no effect of CT upon the spermatozoa. But, we can not exclude any impact on fertilisation ability and/or ultrastructure of these gametes. The finding that oligospermia was more frequent in CT+group, leds us to suggest thas chlamydial infection has a repercution on the gametogenesis or on genital ducts permeability. Another hypothesis would be that oligospermia, reflect of spermatogenesis disorder would be associated with reduction of local immunity. Other studies with wide exploration of spermatic functions and of different parts of genital tract are needed to specify the real impact of genital chlamydial infection upon men reproduction function.     

Diagnostic value of an enzyme‐linked immunosorbent assay using the recombinant CT694 species‐specific protein of Chlamydia trachomatis

Aim: To study the performance of the CT694 protein in relation to the microimmunofluorescence (MIF) and the pELISA tests for the serodiagnosis of Chlamydia trachomatis infections. Methods and Results: The CT694 protein was produced as recombinant protein and was used as antigen in ELISA test for the detection of C. trachomatis IgG antibodies. The performance of the developed ELISA test was compared to the MIF test at two cut‐off values of 16 and 64, and to the specific pELISA test using a panel of 342 sera. These sera were from children MIF C. trachomatis and Chlamydophila pneumoniae negative, patients MIF C. pneumoniae positive, patients MIF C. trachomatis positive, patients suspected to have chlamydial infections diagnosed by the Cobas Amplicor test, healthy blood donors and prostitutes. Our results indicate that the developed ELISA test has performed better compared with the MIF and the pELISA tests. The highest performance was obtained when comparing the developed ELISA test in relation to the pELISA, yielding an overall sensitivity and specificity of 85% and 87% respectively. Conclusions: The CT694 ELISA showed the best performance when compared to the species‐specific pELISA test and may be used for the serodiagnosis of C. trachomatis infections. Significance and Impact of the Study: The CT694 ELISA test responds to the criteria of both sensitivity and specificity according to the MIF and pELISA tests and may be used for serodiagnosis of C. trachomatis infections.     

Genetic Diversity of Food-Isolated Salmonella Strains through Pulsed Field Gel Electrophoresis (PFGE) and Enterobacterial Repetitive Intergenic Consensus (ERIC-PCR)

All over the world, the incidence of Salmonella spp contamination on different food sources like broilers, clams and cow milk has increased rapidly in recent years. The multifaceted properties of Salomnella serovars allow the microorganism to grow and multiply in various food matrices, even under adverse conditions. Therefore, methods are needed to detect and trace this pathogen along the entire food supply network. In the present work, PFGE and ERIC-PCR were used to subtype 45 Salmonella isolates belonging to different serovars and derived from different food origins. Among these isolates, S. Enteritidis and S. Kentucky were found to be the most predominant serovars. The Discrimination Index obtained by ERIC-PCR (0.85) was slightly below the acceptable confidence value. The best discriminatory ability was observed when PFGE typing method was used alone (DI = 0.94) or combined with ERIC-PCR (DI = 0.93). A wide variety of profiles was observed between the different serovars using PFGE or/and ERIC-PCR. This diversity is particularly important when the sample origins are varied and even within the same sampling origin.     

Intérêt de l’étude de l’oxydation de l’ADN des spermatozoïdes par marquage de la 8-oxo-guanine en cytométrie en flux chez l’homme infertile

Oxidative stress in semen is essentially due to excessive production of oxygen-reactive species (ORS) essentially derived from leukocytes. DNA oxidation is due to the direct action of ORS which produce several adducts the most extensively studied of which is 8-oxo-guanine. Integrity of DNA is essential for the fertility of sperm and is an important subject of research for scientists and clinicians all over the world. Although evaluation of the global integrity of sperm DNA has considerably developed over recent years, few tests are available to document oxidative DNA damage. This study was designed to review the various tests of sperm DNA integrity commonly used in the literature and to present the results of our study on DNA oxidation with 8-oxo-guanine labelling by flow cytometry in infertile men. This study was based on 15 semen samples that were submitted to sperm analysis according to WHO guidelines, with determination of the leukocyte concentration by a cytochemical method revealing peroxidase in cytoplasmic granulations. The DNA oxidation study was performed with 8-oxo-guanine labelling by flow cytometry. Linear regression analysis showed a strong correlation between DNA oxidation and leukocyte count in the semen (p = 0.006, r = 0.7). A leukocyte cut-off of 250,000/ml of semen was associated with a significant increase of DNA oxidation (p = 0.03). 8-oxo-guanine can therefore be considered to be a biological marker of the direct action of oxidative stress on sperm DNA which appears to be susceptible to relatively low levels of ORS produced by leukocytes present at concentrations well below the limit of leukospermia defined by WHO.