Snorkelers impact on fish communities and algae in a temperate marine protected area

Multiple-use marine protected areas (MPAs) are used to manage marine resources, allocate space to different users and reduce conflicts while protecting marine biodiversity. In the Mediterranean, MPA managers are increasingly interested in containing the effects of coastal recreation within underwater trails, but snorkelers impacts on the surrounding ecosystem remain largely unknown. In a Mediterranean MPA, an underwater snorkeling trail was established to concentrate snorkelers and increase their awareness of marine habitats and species. The high level of summer frequentation may have negative impacts on the surrounding environment through trampling on the sessile flora or disturbance to the vagile fauna. We used a before-after-control-impact (BACI) design to analyse these potential human impacts. The structure of macroalgae and fish assemblages were used as indicators. Permutational multivariate analyses of variance (PERMANOVA) were carried out to assess potential temporal and spatial changes of the indicators between the trail and a control location within the adjacent no-take/no-use area. Fish communities and macroalgae were subjected to natural temporal trends but no significant impacts of snorkelers were found. Four reasons could explain the absence of snorkelers impact on the surrounding marine environment: (1) the absence of very fragile organisms within the trail (and the control no-take/no-use area) such as gorgonians or bryozoans; (2) the life cycle of the algae with a natural decreasing trend in summer, corresponding to the trail opening period; (3) only a few snorkelers are practicing apnoea; and (4) the information at the entrance and along the trail may influence the snorkelers’ behaviour.     

The Trw Type IV Secretion System of Bartonella Mediates Host-Specific Adhesion to Erythrocytes

Bacterial pathogens typically infect only a limited range of hosts; however, the genetic mechanisms governing host-specificity are poorly understood. The α-proteobacterial genus Bartonella comprises 21 species that cause host-specific intraerythrocytic bacteremia as hallmark of infection in their respective mammalian reservoirs, including the human-specific pathogens Bartonella quintana and Bartonella bacilliformis that cause trench fever and Oroya fever, respectively. Here, we have identified bacterial factors that mediate host-specific erythrocyte colonization in the mammalian reservoirs. Using mouse-specific Bartonella birtlesii, human-specific Bartonella quintana, cat-specific Bartonella henselae and rat-specific Bartonella tribocorum, we established in vitro adhesion and invasion assays with isolated erythrocytes that fully reproduce the host-specificity of erythrocyte infection as observed in vivo. By signature-tagged mutagenesis of B. birtlesii and mutant selection in a mouse infection model we identified mutants impaired in establishing intraerythrocytic bacteremia. Among 45 abacteremic mutants, five failed to adhere to and invade mouse erythrocytes in vitro. The corresponding genes encode components of the type IV secretion system (T4SS) Trw, demonstrating that this virulence factor laterally acquired by the Bartonella lineage is directly involved in adherence to erythrocytes. Strikingly, ectopic expression of Trw of rat-specific B. tribocorum in cat-specific B. henselae or human-specific B. quintana expanded their host range for erythrocyte infection to rat, demonstrating that Trw mediates host-specific erythrocyte infection. A molecular evolutionary analysis of the trw locus further indicated that the variable, surface-located TrwL and TrwJ might represent the T4SS components that determine host-specificity of erythrocyte parasitism. In conclusion, we show that the laterally acquired Trw T4SS diversified in the Bartonella lineage to facilitate host-restricted adhesion to erythrocytes in a wide range of mammals.     

A Bayesian method to estimate the optimal threshold of a longitudinal biomarker

The objective of this study was to develop methods to estimate the optimal threshold of a longitudinal biomarker and its credible interval when the diagnostic test is based on a criterion that reflects a dynamic progression of that biomarker. Two methods are proposed: one parametric and one non‐parametric. In both the cases, the Bayesian inference was used to derive the posterior distribution of the optimal threshold from which an estimate and a credible interval could be obtained. A numerical study shows that the bias of the parametric method is low and the coverage probability of the credible interval close to the nominal value, with a small coverage asymmetry in some cases. This is also true for the non‐parametric method in case of large sample sizes. Both the methods were applied to estimate the optimal prostate‐specific antigen nadir value to diagnose prostate cancer recurrence after a high‐intensity focused ultrasound treatment. The parametric method can also be applied to non‐longitudinal biomarkers.     

Differences in boar sperm head shape and dimensions recorded by computer-assisted sperm morphometry are not related to chromatin integrity

Although sperm head shape and relative dimensions are considered reliable indicators of sperm quality, their quantification is most often operator-driven, e.g., subjective. Artificial insemination semen doses from 35 mature stud boars of known fertility and belonging to three breeds and two hybrid breeds (Duroc, Large White, Landrace, respectively, Yorker and Risco) were used in this study. Sperm samples were extended to 100x10(6) cells per mL and 10microL of the sperm suspension used to made smears which, stained, were examined using phase contrast microscopy interfaced with an automated sperm morphology analyzer (ASMA, ISAS). Each sperm head was measured for four primary parameters [area (A) microm(2), perimeter (P) mum, length (L) microm, width (W) microm], and four derived parameters of head shape [(L/W), (4piA/P(2)), ((L-W)/(L+W)), (piLW/4A)]. Definition of head size was statistically performed. The threshold for each class was established on the basis of the area values, considering the 25th percentile as small and the 75th percentile as large spermatozoa. In a second step, sperm head shape was determined as normal, elliptic, abnormal (rugose) contour, long or irregular and percentiles set as above to define spermatozoa with normal values for each shape parameter. Significant differences were found among breeds in the size of morphologically normal spermatozoa, which were significantly larger and more elliptic (P<0.001) in the Duroc breed. Sperm chromatin integrity was studied using the SCSA-assay, with significant differences observed in the degree of fragmentation intensity (DFI) although this value was consistently low in all animals studied. The hereby-validated ASMA was able to determine significant differences in sperm shape and dimensions among breeds, which were not accompanied by deviations in chromatin structure neither within nor between fertile AI-boars.     

Interest of RGD-containing linear or cyclic peptide targeted tetraphenylchlorin as novel photosensitizers for selective photodynamic activity

Destruction of the neovasculature is essential for tumor eradication by photodynamic therapy. Since the over-expression of integrins is correlated with tumor angiogenesis, we conjugated a photosensitizer (5-(4-carboxyphenyl)-10,15,20-triphenylchlorin or porphyrin) to the alpha(v)beta(3) integrin specific peptide RGD (H-Arg-Gly-Asp-OH) motif as a common sequence. We reported an efficient solid-phase synthesis of a new family of peptidic photosensitizers with linear or cyclic[RGDfK] RGD motif and compared conjugates in vitro selectivity and photodynamic activity. The conjugates were characterized by (1)H NMR, MALDI, UV-visible spectroscopy and singlet oxygen formation was performed. Chlorins containing linear and constrained RGD motif were incorporated up to 98- and 80-fold more, respectively, than the unconjugated photosensitizer over a 24-h exposure in human umbilical vein endothelial cells (HUVEC) over-expressing alpha(v)beta(3) integrin. Peptidic moiety also led to a non-specific increased cellular uptake by murine mammary carcinoma cells (EMT-6), lacking RGD binding receptors. Survival measurements demonstrated that HUVEC were greatly sensitive to conjugates-mediated photodynamic therapy.     

iNKT cells require CCR4 to localize to the airways and to induce airway hyperreactivity

iNKT cells are required for the induction of airway hyperreactivity (AHR), a cardinal feature of asthma, but how iNKT cells traffic to the lungs to induce AHR has not been previously studied. Using several models of asthma, we demonstrated that iNKT cells required the chemokine receptor CCR4 for pulmonary localization and for the induction of AHR. In both allergen-induced and glycolipid-induced models of AHR, wild-type but not CCR4-/- mice developed AHR. Furthermore, adoptive transfer of wild-type but not CCR4-/- iNKT cells reconstituted AHR in iNKT cell-deficient mice. Moreover, we specifically tracked CCR4-/- vs wild-type iNKT cells in CCR4-/-:wild-type mixed BM chimeric mice in the resting state, and when AHR was induced by protein allergen or glycolipid. Using this unique model, we showed that both iNKT cells and conventional T cells required CCR4 for competitive localization into the bronchoalveolar lavage/airways compartment. These results establish for the first time that the pulmonary localization of iNKT cells critical for the induction of AHR requires CCR4 expression by iNKT cells.     

African trypanosomiasis: naturally occurring regulatory T cells favor trypanotolerance by limiting pathology associated with sustained type 1 inflammation

Tolerance to African trypanosomes requires the production of IFN-gamma in the early stage of infection that triggers the development of classically activated macrophages controlling parasite growth. However, once the first peak of parasitemia has been controlled, down-regulation of the type 1 immune response has been described. In this study, we have evaluated whether regulatory T cells (Tregs) contribute to the limitation of the immune response occurring during Trypanosoma congolense infection and hereby influence the outcome of the disease in trypanotolerant C57BL/6 host. Our data show that Foxp3+ Tregs originating from the naturally occurring Treg pool expanded in the spleen and the liver of infected mice. These cells produced IL-10 and limited the production of IFN-gamma by CD4+ and CD8+ effector T cells. Tregs also down-regulated classical activation of macrophages resulting in reduced TNF-alpha production. The Treg-mediated suppression of the type 1 inflammatory immune response did not hamper parasite clearance, but was beneficial for the host survival by limiting the tissue damages, including liver injury. Collectively, these data suggest a cardinal role for naturally occurring Tregs in the development of a trypanotolerant phenotype during African trypanosomiasis.     

Development of a novel method to determine very low density lipoprotein kinetics

Isotopic tracer methods of determining triglyceride-rich lipoprotein (TRL) kinetics are costly, time-consuming, and labor-intensive. This study aimed to develop a simpler and cost-effective method of obtaining TRL kinetic data, based on the fact that chylomicrons compete with large VLDL (VLDL(1); S(f) = 60-400) for the same catalytic pathway. Ten healthy subjects [seven men; fasting triglyceride (TG), 44.3-407.6 mg/dl; body mass index, 21-35 kg/m(2)] were given an intravenous infusion of a chylomicron-like TG emulsion (Intralipid; 0.1 g/kg bolus followed by 0.1 g/kg/h infusion) for 75-120 min to prevent the clearance of VLDL(1) by lipoprotein lipase. Multiple blood samples were taken during and after infusion for separation of Intralipid, VLDL(1), and VLDL(2) by ultracentrifugation. VLDL(1)-apolipoprotein B (apoB) and TG production rates were calculated from their linear increases in the VLDL(1) fraction during the infusion. Intralipid-TG clearance rate was determined from its exponential decay after infusion. The production rates of VLDL(1)-apoB and VLDL(1)-TG were (mean +/- SEM) 25.4 +/- 3.9 and 1,076.7 +/- 224.7 mg/h, respectively, and the Intralipid-TG clearance rate was 66.9 +/- 11.7 pools/day. Kinetic data obtained from this method agree with values obtained from stable isotope methods and show the expected relationships with indices of body fatness and insulin resistance (all P < 0.05). The protocol is relatively quick, inexpensive, and transferable to nonspecialist laboratories.