The quantitative PCR of n-damo 16S rRNA gene was performed with specific primers qP1F-qP1R described previously (Ettwig et al., 2009). Total bacterial numbers were quantified with the primer pair 616F-Eub338-IR specific for the 16S rRNA gene (Amann et al., 1990; Juretschko et al., 1998). Standard curves were obtained with serial dilutions of plasmid DNA containing the target genes. The sequences reported in this study have been deposited in the GenBank database under accession numbers JN704402–JN704415 (n-damo pmoA), JN704416–JN704466 (n-damo 16S rRNA ), and JN704467–JN704568 (anammox hzsB). Owing to the long-term fertilizations, Selleck Androgen Receptor Antagonist the concentrations of nitrogen compounds (, and total

nitrogen) and total organic matter (TOM) in soil were very high (Supporting Information, Fig. S1). Most of the highest values were observed in the upper 10-cm layers except for which was peaked at 10–20 cm (up to 158.8 mg kg−1 dry soil). For , the common electron acceptor for anammox and n-damo bacteria, the highest concentration (53.8 mg kg−1 dry soil) was present at 0–10 cm. After a rapid decrease at 10–30 cm (11.6 ± 0.3 mg kg−1 dry soil), a slight increase in was observed at 30–50 cm of 12.5 ± 0.3 mg kg−1 dry soil, providing a potentially suitable condition for the growth of anammox and

n-damo bacteria. In addition to the previous work exploiting the hzsA gene HKI-272 purchase (Harhangi et al., 2012), we focused on the hzsB gene in this study. A data set with hydrazine synthase β-subunit DNA and protein sequences from the known anammox bacteria of Candidatus genera ‘Jettenia’, Bumetanide ‘Brocadia’, ‘Scalindua’, ‘Kuenenia’, and Planctomycete KSU-1 available from metagenome sequencing projects and GenBank were aligned. Conserved regions of the aligned sequences were identified and used as the targets for designing degenerate primers (Fig. S2). Six forward and five reverse degenerate primers were designed based on the alignment. The sequences and positions on the gene were shown in Table S1 and Fig. S3. Different combinations of the designed primers were tested and evaluated with

template DNA extracted from anammox enrichment cultures. High intensities of specific band (c. 365 bp) were observed (Figs S4–S7) using the primer pair of hzsB_396F and hzsB_742R (at annealing temperature 59 °C and with 2–2.5 mM MgCl2) by single-step amplification instead of nested PCR which was previously required for soil samples (Humbert et al., 2010; Hu et al., 2011; Zhu et al., 2011b). The PCR products were cloned and sequenced, and a phylogenetic tree of the retrieved hzsB sequences from anammox enrichment cultures was constructed (Fig. S8a). The phylogeny of hzsB was consistent with that of the 16S rRNA gene (Fig. S8b) (Schmid et al., 2008) and the hzsA gene (Harhangi et al., 2012). For the molecular detection of anammox bacteria in soil, the 16S rRNA gene was the most common used biomarker (Humbert et al., 2010; Hu et al., 2011; Zhu et al., 2011b).

For C. hominis, differences in apparent mobility were related to the number of thymidine residues in the poly-T region, which ranged from 7 to 11 (Fig. 2). This study is the first to report on the application of capillary

electrophoresis analyses of SSCP for the differentiation of Cryptosporidium species. Although SSCP has been used previously to differentiate Cryptosporidum species, analyses were performed using conventional nondenaturing gel electrophoresis that relied on selleck chemicals manual scoring of band mobilities against a reference control (Jex et al., 2007a, b). In our hands, CE-SSCP provides a method for the differentiation of Cryptosporidium species both within host groups and between host groups. The Cryptosporidium CE-SSCP electropherograms comprise a major peak that corresponds to a single strand of a fluorescently labeled PCR template. For four species, additional minor peaks were detected.

Cloning and learn more sequencing confirmed that multiple peaks corresponded to polymorphism in the 18S rRNA gene. For C. parvum and C. fayeri the peaks corresponded to type A and type B copies of the 18S rRNA gene (Le Blancq Sylvie et al., 1997; Xiao et al., 1999a, b). A third peak in C. fayeri samples appeared to be a recombinant between type A and type B 18 s rRNA gene copies; however, it is also possible that this peak was a PCR chimera of type A and type B. Similarly, the two peaks observed in the Cryptosporidium possum genotype corresponded to the 18S rRNA gene polymorphism (Hill et al., 2008). For C. hominis isolates, the two peaks observed corresponded to variations in the poly-T region of the 18S rRNA gene. The inter- and intraisolate variation for the poly-T region has been shown to range in thymidine numbers from 8 to 12 (Gibbons-Matthews & Prescott, 2003). Inter- and intraspecies variations in the poly-T region, observed in clones from five C. hominis

isolates, corresponded to differences in CE-SSCP mobility. Although several Cryptosporidium species have heterogeneic copies of the 18S rRNA gene, the regions complementary to PCR primers are conserved, and hence a second fluorescent peak is present in amplicons and detectable by CE. The three species of concern to human health, C. parvum, C. hominis and C. meleagridis, were clearly discernable by CE-SSCP, and the multiple peaks observed in C. parvum and C. hominis provided extra Mirabegron discriminatory power. The ability of CE-SSCP to clearly identify multiple peaks in samples indicates that it may be applicable for the detection of mixed infections. However, current PCR protocols need to be optimized for mixed species detection because preferential amplification of C. parvum has been observed in the past. Mixes of C. parvum and C. hominis DNA over a range of concentrations have shown that C. parvum is preferentially amplified (Cama et al., 2007; Waldron et al., 2009). CE-SSCP could be applied to evaluate PCR protocols for the detection of mixed infections.

Also, a link between activity-regulated I-BET-762 mw cytoskeleton-associated protein (Arc), PKMζ and LTP has been proposed. Our previous results demonstrated that re-exposure to the withdrawal environment was able to evoke the memory acquired when the anxiety measured as a diazepam (DZ) withdrawal sign was experienced. In the present work we evaluated if the memory associated with DZ withdrawal could be affected by changes

in the contextual cues presented during withdrawal and by intrahippocampal administration of a PKMζ inhibitor. We found that the context was relevant for the expression of withdrawal signs as changes in contextual cues prevented the expression of the anxiety-like behavior observed during plus-maze (PM) re-exposure, the associated enhanced synaptic plasticity and the increase in Arc expression. Furthermore, intrahippocampal administration of PKMζ inhibitor previous to re-exposure to the PM test also impaired expression of anxiety-like behavior and the buy 5-FU facilitated LTP. These results support the relevance of the hippocampal synaptic plasticity in the maintenance of the memory trace during benzodiazepines withdrawal, adding new evidences for common mechanisms between memory and drug addiction that can be intervened for

treatment or prevention of this pathology. “
“The mouse has emerged as an advantageous species for studying the brain circuitry that underlies complex behavior and for modeling neuropsychiatric disease. The transition from flexible, goal-directed actions to inflexible, habitual responses is argued to be a valid and reliable behavioral model for studying a core aspect of corticostriatal systems that is implicated in certain forms of psychopathology. This transition is thought to correspond to a progression of behavioral control from associative to sensorimotor corticobasal ganglia networks. Habits form following extensive training and are characterized by reduced sensitivity of instrumental responding to reinforcer revaluation; few studies

have examined this form of behavioral control in mice. Here we examined the involvement of the dorsolateral and dorsomedial striatum in this transition in the C57BL/6 inbred mouse strain. nearly We provided evidence that damage to the dorsolateral striatum disrupted habitual responding, i.e. it preserved sensitivity to changes in outcome value following either outcome devaluation or, shown for the first time in mice, outcome inflation. Together, these data show that instrumental responding in lesioned mice tracks the current value of a reinforcer and provide evidence that neuroanatomical mechanisms underlying habit learning in rats are preserved in the mouse. This will allow for the genetic and molecular dissection of neural factors involved in decision-making and mechanisms of aberrant habit formation.

The overnight cultures were diluted into 50 mL of medium to an OD600 nm of 0.05 and grown for several hours to an OD600 nm of 0.2. To determine the relative amounts of glutathionylspermidine and of spermidine in each strain, cells were prelabeled with 1.25 μCi of [14C]-spermidine trihydrochloride (12.5 nmoles), and the incubation was continued for either 2 h (‘log-phase culture’ OD600 nm = 0.7) or 20 h (‘overgrown culture’). The cultures were rapidly centrifuged at room temperature. The pellets were washed

twice with medium and re-suspended in 10% perchloric acid (1 : 5 wt/vol); the supernatants were subjected to HPLC chromatography on a Shim-pack cation exchange Dasatinib column with the elution system described in the previous section but with 1.0 M NaCl-0.2 M Talazoparib sodium citrate as the elution buffer. The elutes were collected at 2-min intervals (0.7 mL min−1), and a 100-μL aliquot from each fraction was counted in a Beckman scintillation counter (LS6500). Three independent cultures (109–1010 cells) from the E. coli gss+ and Δgss cells (OD600 nm of 0.7–0.8) were

harvested and re-suspended in Tris-EDTA buffer (100 mM Tris, 10 mM EDTA, pH 8.0) containing 2 mg mL−1 lysozyme (Sigma). The cell suspensions were incubated for 5 min at room temperature to digest the cell wall. Total RNA was isolated according to the protocol described in the RNeasy mini kit (Qiagen, Germantown, MD). The mRNAs were enriched from total RNA by removing the 16S and 23S ribosomal RNAs using the MICROBExpress method and kit (part no. AM1905; Ambion). The quantity and quality of RNA were evaluated by OD260 nm/OD280 nm assays and by RNA capillary electrophoresis (Agilent Biotechnologies). Enriched mRNAs were reverse-transcribed by Superscript II and random hexanucleotide primer

(Invitrogen) and used for microarrays as described earlier (Chattopadhyay et al., 2009a) using Affymetrix (Santa Clara, CA) E. coli GeneChip arrays (Genome 2.0 array; n = 3 each for gss+ and Δgss). anova (analysis of variance) was performed, and P-values were calculated Mirabegron using Partek Pro-software (Partek, St. Louis, MO) and plotted in negative log scale on y-axis against the Affimetrix signal ratios for each probe set on x-axis. Up- and down-regulated genes were selected based on P-values of <0.05 and fold change > +2 or −2. The complete microarray data can be obtained from GEO (accession number GSE30679). Most striking is that sequences homologous to E. coli Gss are only found in Eubacteria and the very distantly related Kinetoplastids (plus two fungal species with relatively low homology; Table 2). No homologous sequences (as defined by the blast-p program) were found when the E. coli Gss sequence was compared with the human, rat, mouse, Arabidopsis, rice, worm, and Drosophila sequence databases (Table 2).

In both areas, these correlations were stronger in neurons showing delay selectivity than in those without delay selectivity. Notably, delay-selective neurons in A35 responded similarly to the optimal stimulus and its paired associate, whereas delay-selective PLX-4720 research buy neurons in A36 discriminated between them. However, these neurons in both areas discriminated the primary pair, consisting of the optimal stimulus and its paired associate, from other pairs, indicating that selectivity across pairs was maintained between the two areas. These results suggest that delay-selective neurons in A35 represent these paired stimuli as a single unitized item rather than two associated items. “
“The

activity of neurons in the rostral ventrolateral medulla (RVLM) is critical for the generation of vasomotor sympathetic tone. Multiple pre-sympathetic pathways converge on spinally projecting

RVLM neurons, but the origin and circumstances in which such inputs are active are poorly understood. We have previously shown that input from the contralateral brainstem contributes to the baseline activity of this population: in the current study we investigate the distribution, phenotype and functional properties of RVLM neurons with commissural projections in the rat. We firstly used retrograde transport of fluorescent microspheres to identify neurons that project to the contralateral RVLM. Labelled neurons were prominent in a longitudinal column that extended over 1 mm caudal from the facial nucleus and contained hybridisation products indicating enkephalin BAY 80-6946 (27%), GABA (15%) and adrenaline (3%) synthesis and

included 6% of bulbospinal neurons identified by transport of cholera toxin B. Anterograde transport of fluorescent dextran-conjugate from the contralateral RVLM revealed extensive inputs throughout the RVLM that frequently terminated in close NADPH-cytochrome-c2 reductase apposition with catecholaminergic and bulbospinal neurons. In urethane-anaesthetised rats we verified that 28/37 neurons antidromically activated by electrical stimulation of the contralateral pressor region were spontaneously active, of which 13 had activity locked to central respiratory drive and 15 displayed ongoing tonic discharge. In six tonically active neurons sympathoexcitatory roles were indicated by spike-triggered averages of splanchnic sympathetic nerve activity. We conclude that neurons in the RVLM project to the contralateral brainstem, form synapses with sympathetic premotor neurons, and have functional properties consistent with sympthoexcitatory function. “
“Gamma protocadherins (Pcdh-γs) resemble classical cadherins and have the potential to engage in cell–cell interactions with homophilic properties. Emerging evidence suggests non-conventional roles for some protocadherins in neural development. We sought to determine whether Pcdh-γ trafficking in neurons is consistent with an intracellular role for these molecules.

In such cases, the parameter k may reach a value close to 1. The parameter k, representing the nearshore wave energy in relation to the offshore wave energy for all encountered

wave conditions, is illustrated in Figure 6. Apart from coastal swell and wind waves, there may also be oscillatory motion of the water, characterized by longer periods. Such waves, called infragravity waves, are said to have a significant influence Copanlisib chemical structure on coastal morphodynamic processes (Aagaard and Greenwood, 2008, Coco et al., 1999, Coco et al., 2001 and Pruszak et al., 2007). The study of Pruszak et al. (2007) concerns the southern Baltic coast, and includes the Lubiatowo site considered in the present paper. It appears that both standing and progressive infragravity waves can occur in a multi-bar dissipative coastal zone. These latter waves are generally much smaller than gravity waves, and decrease rapidly in height seawards of the shoreline. Infragravity waves are

therefore likely to create and modify rhythmic shoreline forms, but are unlikely to affect the onshore and offshore movement of the entire shoreline (Pruszak et al. 2007). An important indicator of beach resilience, especially for dunes on the beach INCB018424 hinterland, is beach width. Owing to possible large variations at shorter time scales, the behaviour of beach width in the long term is of considerable importance. Long-term field data (1875–1979) collected from the ca 500 km long non-tidal coast of Poland suggest that a sandy seashore with dunes is relatively safe and stable when the beach width (ys–yd) is no less than 40 m ( Dubrawski & Zawadzka (eds.) 2006). Similar conclusions, defining the safety

of a sandy shore by a beach width of at least 40–50 m, can be drawn from investigations of other southern and south-eastern Baltic shores ( Boldyrev, 2008 and Bobykina and Boldyrev, 2008). Observations of the shore at Lubiatowo, comprising measurements of shoreline and dune toe positions carried out since 1983, indicate that this coastal section has been rather stable in the long term. Nevertheless, beach safety criteria are different for tidal shores where the hydrodynamic loads are more complicated. On tidal coasts, the mean beach width during the Thalidomide ebb tide can be 2–3 times larger than at high tide. Moreover, unlike dissipative non-tidal shores, the beach width is bigger in winter than in summer ( Quartel et al. 2008). Previous surveys at CRS Lubiatowo have shown that it is difficult to make out any clear seasonality of variations in the parameter (ys–yd): this can be assumed as evidence that the randomness of morphological processes plays a more important role than seasonal climatic fluctuations. A certain regularity is discernible only for the autumn months (decrease of beach width): this can probably be explained by the storms and other extreme events that usually occur at that time and cause periodic intensification of beach erosion and shoreline retreat.

Plates were washed six times and 100 μl of rabbit polyclonal anti-Hsp70 (1/400) diluted in PBS/T containing 4% mouse serum was added. After 1 h on shaker at 37 °C, plates were washed and incubated with 100 μl of an anti-rabbit immunoglobulin peroxidase conjugate in selleck screening library PBS/T/BSA (1/10,000) for 1h on shaker at 37 °C. Plates were then washed and 200 μl of o-phenylenediamine dihydrochloride (OPD) substrate

was added. After 45 min on shaker and at 37 °C, the reaction was stopped with 50 μl of sulphuric acid (1 N H2SO4) and the absorbance determined at 490 nm with background subtraction at 620 nm using a microplate reader (Ceres 900C, Bio-Tec Instruments, Inc., Belgium). Hsp70 concentrations in serum were detected by comparing sample absorbance with the absorbance of a reference purified human recombinant Hsp70 protein. The serum levels of 25-OH-vitamin-D were determined using the 25 hydroxyvitamin D125I RIA Kit (Diasorin Inc., Stillwater, USA; normal values: 16–74 μg/l). Vitamin B12 and folate were determined with the Simultrac Radioassay Kit (Becton Dickinson Immunodiagnostics, USA; normal values: 0.22–0.94 μg/l and 2.0–14.0 μg/l for vitamin B12 and folate, respectively). The serum levels of parathyroid hormone (PTH) were determined using the N-tact

PTH Irma Kit (Diasorin Inc, Stillwater, USA; normal values 15–65 ng/l). Calcium was measured in serum by the o-cresolphthalein complexone method (Bio Phase Diagnostics Laboratory, Ontario, CA; normal values 8.6–9.8 mg/dl). Selleck INK128 Oxalosuccinic acid Antimalarial antibody concentration was determined in the clinical laboratory of the Institute of Tropical Medicine (Antwerp, Belgium) as reported elsewhere (Njemini et al., 2002). Antimalarial antibodies were tested by an indirect immunofluorescence using antigens from the Institute of Tropical Medicine and an anti-human immunoglobulin (IgGAM) conjugate. Titers ≥ 1/40 were considered positive. Fresh skin snips, taken from the lower extremities, as well as fresh blood were screened microscopically for the presence of filarial parasites. All reagents were applied according to manufacturers’ recommendations. Column statistics (with statistical package prism 3.0) was used to test

the approximation of the population distribution to normality. Spearman’s rank test was used to examine the relationship between the serum concentrations of Hsp70 and the levels of the other parameters. For the comparison of Hsp70 levels between two groups, the nonparametric Mann–Whitney test was applied. A 2-sided p < 0.05 was considered statistically significant. Table 1 summarizes the data for women and men. The Hsp70 serum levels varied between 0 and 47 ng/ml (median 13 ng/ml) in female and between 0 and 78 ng/ml (median 13 ng/ml) in male. There were no relationships with gender. Hsp70 concentrations were found to be dependent on the degree of inflammation, as measured by the circulating CRP levels (r = 0.172, p = 0.044), as well as by the WBC count (r = 0.

Slides were evaluated Linsitinib chemical structure using a Leica DMR upright microscope equipped for epifluorescence microscopy. ST sections were stained for TH immunoreactivity (IR) using nickel enhancement and slides were scanned as 600 dpi, 8 bit grayscale tiff images

using a CanoScan8400F flatbed scanner with automatic settings disabled. One section at the level of the anterior commissure (−0.26 mm from Bregma) was chosen from each brain for fiber density measurement. Using Fiji software (http://imageJ.nih.gov/ij), each ST was outlined using the freehand tool, processed to correct for background and brightness, converted to a binary image and the number of pixels in the ST determined as a measure of TH-IR fiber density. ST fiber density data are expressed as a percentage of injected vs control ST for each treatment group. Sections of SN were stained for TH-IR without nickel enhancement. For the dose response study, the number of TH-IR neurons was counted in both injected and control SNs in one section from each brain at the level of the medial terminal nucleus accessory optic tract (−5.3 mm from Bregma) using Neurolucida software (Micro Bright Field Biosciences). selleck products For the efficacy study, unbiased stereology was used to determine the total number of TH-IR neurons in each SN. Seven sections at similar anterior to posterior levels were chosen throughout

the anterior-posterior extent of the SN in every brain. The number of TH-IR cells was counted in injected and control SN using StereoInvestigator™ software (Micro Bright Field Biosciences). Parameters were as follows: grid size (80×80), frame size (175×175), guard zones (3 μm), optical dissector height (10 μm). The Gundersen coefficient of error was ≤0.07 for TH neuronal counts in the

control SN and ≤0.11 for Carnitine palmitoyltransferase II TH neuronal counts in experimental SN. For both counting methods, only large TH-IR neurons (greater than ~15 μm in diameter) were counted to avoid counting interneurons or dying neurons. Histology images were collected using a Retiga 4000R digital camera on a Leica DMR upright microscope. Adobe Photoshop CS5 was used to compile multi-photo plate figures. Data were analyzed using Prism software. Kruskal–Wallis one-way ANOVAs followed by Dunn’s post hoc tests were used to compare treatment groups for forelimb behavior analysis. All other data were analyzed using a parametric one-way ANOVA followed by Tukey’s post hoc tests. Statistically significant differences were set at p≤0.05. Data are expressed as mean±SEM. This study was supported by the Department of Defense Neurotoxicology Program (NO06079001) and NIH grants (NS31957 and NS054989 to M.C.B. and T32 NS041234 to C.E.K.), the Harry F. and Elaine M. Chaddick Foundation and the Medical Research Institute Council of Ann and Robert H. Lurie Children’s Hospital of Chicago. The University of Pennsylvania Viral Vector Core is acknowledged for AAV production. The technical assistance of Jianping Xie and Brian Corstange (Ann and Robert H.

(21)), the ideal dissociation model (Eq. (26)), and the molality- and mole fraction-based ideal dilute models defined in Eqs. (22), (24), (23) and (25), respectively, Selleckchem Z VAD FMK were used to make predictions of solution osmolality in each of the ten multi-solute solution systems listed in Table 2. Fig. 1, Fig. 2, Fig. 3, Fig. 4, Fig. 5, Fig. 6, Fig. 7, Fig. 8, Fig. 9 and Fig. 10 show a representative isopleth and corresponding model predictions

from each of the considered solution systems. Table 6 and Table 7 give the average values of RRTO2 and %MRME, respectively, calculated over all isopleths within a given solution system for each of the six models considered. Each table also contains an overall (unweighted, e.g. with respect to number of isopleths) average value of its corresponding measure calculated over all the solution systems for each model. Before discussing the results in Table 6 and Table 7, an important point should be

made regarding one of the measures of model prediction accuracy used in this work, that is, RRTO2. As is discussed in greater detail in Appendix B, RRTO2 is not directly comparable to a “standard” R  2 statistic (i.e.   one with the total sum of squares calculated using Eq. (B3) instead of Eq. (B7)). In fact, RRTO2 values for a given prediction or fit will always be higher than the corresponding R  2 values. Thus, for example, while a value of R  2 = 0.9 might represent a respectable prediction, RRTO2=0.9 does not. From selleck chemical the results in Table 6 and Table 7 and Fig. 1, Fig. 2, Fig. 3, Fig. 4, Fig. 5, Fig. 6, Fig. 7, Fig. 8, Fig. 9 and Fig. 10, it is evident that the three non-ideal models perform considerably better than the three ideal models. However, none of the three non-ideal models Rapamycin mouse is clearly superior to the others. Each non-ideal model has solution systems where it is noticeably—at least, in terms of %MRME—more accurate than the other two (e.g. Me2SO + glycerol for the molality-based multi-solute osmotic virial equation, EG + NaCl + sucrose for the mole fraction-based multi-solute osmotic virial equation,

and NaCl + sucrose for the freezing point summation model), but overall the performance of all three non-ideal models is very close. In contrast to the non-ideal models, there is a distinct difference in the performance of one of the ideal models relative to the other two: the molality-based ideal dilute model and the ideal dissociation model clearly provide more accurate predictions than the mole fraction-based ideal dilute model in almost all of the solution systems considered (the lone exception being BSA + OVL, where all three ideal models provide equally poor predictions). Given that the main difference between the molality- and mole fraction-based ideal dilute models is the way in which concentration is defined, the gap in their prediction accuracy highlights the importance of the choice of concentration units in thermodynamic modeling.

Gorgonians such as E. verrucosa create complex elevated structures ( Jones et al., 1994), which provide settlement sites for larvae ( Howarth et al., 2011) and create habitats for associated organisms such as the whip fan nudibranch (Tritonia nilsodhneri) ( Hall-Spencer et al., 2007). The sessile RAS indicator species, and their associated biodiversity, produce planktonic larvae that support higher trophic levels. This bentho-pelagic coupling through a range of trophic links provides prey for birds (Grecian et al., 2010), and commercially

important fishes such as cod (G. morhua, Heath and Lough, 2007 and Lomond et al., 1998). For these reasons, sessile RAS are recognised by governments for their importance to ecosystem functionality, and receive protection under environmental legislation from destructive human activities. This includes species http://www.selleckchem.com/products/lgk-974.html such as E. verrucosa in the UK, which is protected Vincristine by the UK Biodiversity Action Plan. By their very nature, sessile RAS need to attach to hard substratum and therefore, indicate ‘reef’, which is often a protected feature of environmental legislation. Reef substratum can be observed by humans as rock, boulders or cobbles, and protected to allow recovery of RAS. However, where sediment overlies rock, reef cannot be identified through habitat assessment, but could be identified by the presence of sessile RAS. Our results indicate that sessile

RAS can only indicate such additional reef habitat if the area is protected from fishing, thereby giving sensitive species a chance to recover. This however, presents a difficult situation for marine managers. Site based protection which encompasses features, such Tortugas Ecological Reserve, and Buck Island Reef National Monument in the USA (Jeffrey et al., 2012 and Kendall et al., 2004), allows sessile RAS to colonise not only areas of visual reef but also areas that are functionally reef to these species i.e. they can find

attachment to hard substratum through overlying sediments. It is clear that by ‘Drawing lines at the sand’ where the visible rocky reef feature ends, managers limit the reef area, but by alternatively protecting sites that encompass features, the functional reef extent can expand and be fully protected. This effect Y-27632 solubility dmso observed here could occur with other protected features in MPAs such as seagrass beds. Our findings are currently of particular importance as improving, low cost GPS technology is allowing what some GIS experts may think is a ‘more intelligent’ detailed design of MPA boundaries rather than a simple box. However, in practice for ecosystem function, simplicity of enforcement and clarity to users (Great Barrier Reef Marine Park Authority 2002) would be the more intelligent design. For example, in Europe, Special Areas of Conservation management focuses on the features within designated sites (European Commission 2000), such as the physical reef habitat.