75) to bring the total TCA percentage to 0.1% following the manufacturer’s direction. Samples RG7204 cost were then taken and added with the reconstituted luciferase/luciferin reagent mix from the kit in a sterile white 96-well plate (Nunc) and the ATP luminescence determined in a Biotek Synergy HT luminometer using KC4 software and compared to control cells not treated with 2-DG. For accumulation, cells

were treated with 10 mM 2-DG before incubation with [3H]nifurtimox as described above. In a series of experiments to assess the impact of CT on [3H]nifurtimox cellular accumulation, the clinically relevant concentrations of melarsoprol (30 μM), pentamidine (10 μM), suramin (150 μM) or eflornithine (250 μM) were added to accumulation buffer. DMSO was used to dissolve melarsoprol and pentamidine to give a final concentration of 0.05% DMSO. Control experiments here also contained 0.05% DMSO. For unlabelled eflornithine and suramin and the appropriate controls, no DMSO was used. There was no significant difference between accumulation of [3H]nifurtimox with or without 0.05% DMSO (data not shown). The cytotoxic effects of the drugs used in this study were assessed on confluent

monolayers of cells in 96 well plates using an MTT assay. Cells underwent 30 minute incubations with a 200 μl/well aliquot of each drug in accumulation buffer at the concentrations used in the experiments. After 30 min, the buffer was aspirated and replaced Chlormezanone with a 100 μl http://www.selleckchem.com/products/atezolizumab.html aliquot of 1 mg/ml MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, Sigma, UK) in DMEM without phenol red (Gibco, Invitrogen, UK). The cells were then incubated for 4 h at 37 °C, the solution removed and replaced with 100 μl propan-2-ol per well, and the absorbance was measured. Absorbance values were corrected by protein content (determined using a BCA assay) and expressed as percentage

viability compared to control untreated cells. The expression of P-gp and BCRP by the hCMEC/D3 and HepG2 cell lines was analysed by Western blot using Abcam primary mouse anti-P-gp/MDR1 [C219] (ab3364) and mouse anti-BCRP/ABCG2 [BXP-21] (ab3380) monoclonal antibodies at 1:80 and 1:1000 dilutions in PBS-Tween (PBS-T, PBS with 0.05% Tween 20) with 0.5% BSA, (Sigma) respectively. Mouse anti-GAPDH monoclonal antibody [6C5] (ab8245), was used as a loading control, 1:1000 in PBS-T with 0.5% BSA. Confluent monolayers of hCMEC/D3 cells and flasks of HepG2 cells (positive controls) were lysed in TGN lysis buffer (50 mM Tris, 150 mM NaCl, 10% glycerol, 50 mM glycerophosphate B, 1% Tween-20, 0.2% NP-40, all purchased from Sigma, UK), and 25 μg loaded per lane. For P-gp, a precast 4–20% gradient gel was used (Bio-Rad Europe, 456-1093S). For BCRP, a 10% SDS-PAGE acrylamide/bisacrylamide gel was used. Following electrophoresis, proteins were transferred using semi-dry transfer onto methanol activated Immobilon-P PVDF membranes (0.

These Bortezomib mw trials should also include an assessment of safety. For the most part, the new BLA references the original BLA, including the non-clinical, chemistry, manufacturing and controls data. Continuous post-licensure surveillance is used to confirm the safety of vaccines in the general population, including people with a variety of health backgrounds. Post-licensure studies of safety and effectiveness of vaccines are now considered increasingly important and are often based on national

immunisation programmes and safety surveillance. Due to the nature of surveillance methods, such as patient registers and call-backs, post-licensure data may not appear in the

literature until 5–10 years after a vaccine has been granted a licence. It is important to assess the background incidence in non-vaccinees of rare conditions and AI disorders that might be possibly diagnosed in temporal association with vaccination (Table 5.2). The background incidence is required to determine whether temporal associations with vaccination are in line with the natural expected incidence rate or if there is an increased incidence selleck products that may suggest a causal link with the vaccine, as described in the rotavirus case study (see case study 3). Vaccine pharmacovigilance is defined by the Council for International Organizations of Medical Sciences as ‘the science and activities relating to the

detection, assessment, understanding, prevention and communication of AEs following immunisation, or of any other vaccine- or immunisation-related issues’. This covers many activities such as continuous benefit–cost assessment, risk management or communication activities to improve vaccine safety. Pharmacovigilance activities include the collection, analysis and reporting of AEs following authorisation. These reports are received from different sources, with the most frequent being healthcare professionals. Reporting to the competent authorities Alanine-glyoxylate transaminase can be expedited or periodic. The expedited reporting of serious unexpected suspected adverse events (SUSARs) to regulatory authorities should be done no later than 15 days from their receipt. In Europe, life-threatening or fatal events must be reported within 7 days. Periodic safety reporting, which in Europe takes the form of a periodic safety update report (PSUR), should be submitted to regulatory authorities at 6-monthly intervals until a full 2 years of marketing experience has been completed, then one is due every year for the following 2 years and every 3 years thereafter. Examples of assessments, requirements and timings in vaccine pharmacovigilance are summarised in Figure 5.7. Case study 2.

Different strategies have been applied for linking antibodies with DNA templates, like streptavidin bridge combined with biotinylated antibody and biotinylated DNA template, or chemically conjugated antibody-DNA complexes ( Lind and Kubista,

2005 and Niemeyer et al., 2007). The amount of DNA amplified during PCR corresponds to the amount of target structure recognized by the antibody, Selleckchem MK-2206 and can be detected by electrophoresis ( Zhou et al., 1993) or by ELISA, utilizing digoxigenin- or biotin-labeled PCR products ( Niemeyer et al., 1997 and Smrž and Dráber, 2003). Later, immunodetection was combined with real-time PCR and used for quantification of vascular endothelial grow factor ( Sims et al., 2000). The method was further modified in such a way that both protein detection

and real-time PCR were performed in the same well of the TopYield strip ( Niemeyer et al., 2007). Furthermore, a gold nanoparticle (Au-NP)-based bio-bar code assay for ultrasensitive detection of proteins has been developed. The assay utilizes Au-NPs functionalized with both thiolated single-strand DNA oligonucleotide and an antibody to the target antigen ( Nam et al., 2003, Nam et al., 2004 and Georganopoulou et al., 2005). Finally, PCR assays based on antibody- and oligonucleotide-functionalized Au-NPs were used for detection of Hantaan virus nucleocapsid protein ( Chen et al., 2009) and respiratory syncytial viruses ( Perez et al., 2011). Although the assays showed high sensitivity for virus detection, they required two sets of wells (for immunodetection and PCR) and therefore were not suitable for high throughput screening and were fraught with high risk of contamination. Here check details we tested

the suitability of functionalized Au-NPs-based iPCR (Nano-iPCR) for detection of low concentrations of cytokines in cell culture supernatants, and changes in cytokine concentration in aging cultures of BMMCs. We defined the conditions for simplified detection of cytokines by Nano-iPCR, and compared the performance of assays based on antibodies anchored either directly on the plastic surface or through extravidin. The assays were carried out in PCR polypropylene wells or wells of TopYield polycarbonate strips which allow more efficient binding of antibodies. We further compared Nano-iPCR with iPCR and G protein-coupled receptor kinase ELISA; outline of the assays is shown in Fig. 1. For these comparisons we utilized identical immunoreagents in all assays. The data indicate that Nano-iPCR offers a sensitive, rapid and robust assay for detection of low concentrations of cytokines in complex biological fluids. Advantages and drawbacks of different assays are discussed. Rabbit anti-murine IL-3 and rabbit anti-murine SCF polyclonal antibodies and their biotinylated forms, recombinant (r) murine IL-3 and rSCF were all obtained from PeproTech (London, UK). Colloidal Au-NPs (30 nm), containing approximately 2 × 1011 Au-NPs/ml, were obtained from BBInternational (Cardiff, UK).

The other personnel costs are obtained by multiplying

the estimated 20 euro per capita by the number of days that the operation will take, which is obtained from the node Time for mechanical/manual removal. This is the last of the factors considered in the Shoreline clean-up cost node. It is assumed that the boats selleck products will be operating as long as the mechanical removal is taking place. The manual removal may take considerably more time, as the removal method covers the more sensitive areas of the shore, and, at this stage the assistance of the oil-recovery boats is most likely unnecessary. In Table 7, the costs for each boat type are presented, as well as the share that each boat type has of the total boat fleet. The costs are based Dabrafenib ic50 on the discussion at the workshop. According to the experts’ panel an average boat cost is about 250 euro per hour, and there are approximately 50 oil-recovery boats located along the Finnish shore of the Gulf of Finland at any given time. Considering that all of these boats would be sent to collect oil and help to protect the coastline with booms, we arrive at a total hourly Boat cost of 12,500 euro. By using this cost and multiplying it by the time from the Time for mechanical removal, we

calculate the probability table for the Boat cost, which has 28 states in total, defined as intervals. This variable is dependent on the following variables: Air surveillance cost, Cost of emptying tanks, Combating cost, Preparation cost and Booms. This node is estimated by using the hourly cost of the Dornier surveillance aircraft: 7000 euro per hour. The aircraft will make its two-hour surveillance runs three times per day, which means that the total daily cost amounts to 42,000 euro. The number of days during which runs are completed is taken from the Time to collect the oil, since it is assumed that surveillance of the movements of the oil slick is required as long as the oil-combating vessels are operating. The probability

table is calculated with the following expression: equation(8) Air surveillance cost=42,000ifC824<1C824·42,000otherwisewhere C8 is Time Rho to collect oil (h). Eq(8) specifies that even if the oil-combating vessels have less than one day to recover the spilt oil, the air surveillance will still take place, as it must estimate the damage and make sure that the authorities are well-informed about the location and the trajectory of the oil slick. In this case the air surveillance cost will automatically be 42,000 euro. Otherwise, the daily air surveillance costs are multiplied with the number of days the operation takes. This node refers to the cost arising from the combating vessels emptying their full tanks during the operation so that they can return to the oil slick to continue the oil-combating operation.

g., citric acid, malic acid), fruit dreg extracts (Shui and Leong, 2002 and Wudrich et al., 1993), etc. Traditional detection methods including sense organ appraisal, physical and chemical identification, mostly focusing on detecting the MK-2206 supplier main components (e.g., soluble solid state materials, total sugars, and total acids) or unique constituents (e.g., inorganic elements, amino acids, and organic acids) of fruit juice. HPLC, GC, MS, NIR, Electronic Tongue (ET) and other techniques have been used to detect food components (Gayo and Hale, 2007,

Hilt et al., 2003, Ogrinc et al., 2003 and Ruiz-Matute et al., 2007). However, as the means of ‘fake food’ production improve, it becomes much more difficult to detect these adulterations. Insufficient original juice contents, fake

juices, sterilization or the use of reconstituted juice concentrates to replace fresh juice are all considered fruit juice adulterations(Gurdeniz and Ozen, 2009 and Tripathi et al., 2004), but traditional methods cannot always identify these changes. Thus, manufacturers would make use of this technology gap to produce the fake juice and get benefits. Along with the advances in modern biotechnology, molecular biology methods have become widely applied in the field of food authenticity Selleck Veliparib identification, and the development of new methods is becoming a popular topic of research worldwide. Specifically, rapid PCR-based methods with high sensitivity and reproducibility (Lockley and Bardsley, 2000 and Mafra et al., 2008) have become especially common, for example, in the identification of genetically modified food (Wurz, Bluth, Zeltz, Pfeifer, & Willmund, 1999), meat (Kung et al.,

2010), milk and cheese (Sachinandan et al., 2011) and their by-products. Sass-Kiss (Sass-Kiss & Sass, 2002) isolated four tissue-specific peptides from grapefruit juice and peel and successfully tested commercial grapefruit juice products for adulteration. Ng, Chang, Wu, Kotwal, and Shyu (2006) designed primers based on the 18S and internal transcribed spacer (ITS) region of the orange as part of a rapid and accurate molecular approach to identify freshly squeezed and reconstituted orange juice. Mooney, Chappell, and Knight (2006) designed primers based on the chloroplast trnT-trnL intergenic BCKDHB spacer region in the orange and mandarin genomes; the heteroduplex resulting from the co-amplification of a fragment containing an 8 base-pair indel distinguished mixtures of orange and mandarin juice. Gimeenez, Piston, Martin, and Atienza (2010) used qRT-PCR and molecular markers for olive oil authentication. Above all, more and more researchers are inclined to adopt PCR methods for detecting food adulteration and doping. Endogenous reference gene analysis is broadly applied in food component source authentication and to qualitatively and quantitatively evaluate food samples. To date, endogenous reference genes of many species have been reported, including the LAT52 gene in the tomato ( Yang et al.

No differences were discernible in secondary Inhibitor Library mineralized trabecular surfaces (Fig. 5). Statistical comparisons based on unpaired t-tests indicated that there were significant differences

in this ratio in primary mineralized trabecular areas (Fig. 6a), with treated animals exhibiting a significantly higher PYD/divalent collagen cross-link ratio compared to the corresponding controls, regardless of treatment duration. Since this is a ratio, the observed increase could be due to several possibilities regarding the change in the individual factors. To further discern the reason for the observed increase in the treated animals, the relative % area of the individual underlying bands (1660 and 1690 cm− 1, representative of Pyd and divalent collagen cross-links, respectively) were plotted (Fig. 6b), revealing a disproportionate decrease in Pyd and divalent collagen Sunitinib supplier cross-links, in agreement with the biochemical analysis data. Similar findings were observed when the cortical periosteal surfaces were compared (Figs. 6c and d, respectively). The results thus far indicated that β-APN

treatment affected bone structural properties, collagen cross-links in anatomically confined areas (primary mineralized packets in trabecular, and periosteal cortical surfaces), and mechanical properties. The statistically Buspirone HCl significant correlations between these outcomes along with the Spearman’s rho value are listed in Table 5. Stiffness correlates well with biochemically, and spectroscopically determined trabecular Pyd/divalent collagen cross-links, and cortical thickness (Ct.Th). Maximum force to failure correlates well with biochemically, and spectroscopically determined trabecular pyd/divalent collagen cross-links, TriSmi, Tb.Th, and Ct.Th. Finally, maximum energy

to failure correlates well with biochemically determined Pyd/divalent collagen cross-link ratio, Ct.Th, and periosteal Pyd/divalent collagen cross-link ratio. The results of the present study employing a lathyritic rat animal model indicate that collagen cross-links coupled with structural changes are a major contributor to bone strength, in line with previously published reports in animal models and human tissue [22], [23], [34], [35], [36], [37], [38] and [39]. They also indicate a correlation with bone structural properties, in agreement with previously published results [40]. They additionally indicate that even when these changes are anatomically restricted (in the present case only in primary mineralized bone), coupled with changes in bone structural properties, they are sufficient to influence the mechanical performance of whole bone, even in the absence of concomitant mineral quantitative and/or qualitative properties alterations.

The similarity matrices generated from an analysis of both living and dead assemblages were examined using the RELATE routine in PRIMER

v6, which measures how closely related two sets of multivariate data are by calculating a rank correlation co-efficient (Clarke and Gorley, 2006). In order to determine whether there were differences between Foraminifera from pipeline and non-pipeline sites in each location, and between locations, the multivariate data were analysed using the PERMANOVA routine in PRIMER v6. Further, in order to determine which species were most responsible for the similarity within each location, a SIMPER (Similarity Percentage) analysis was performed, and the results have been graphically displayed. To see more explore the relationships between Foraminifera and the environment two further find more analyses were undertaken. Firstly Spearman rank order correlations (using STATISTICA v. 11 and Bonferroni correction of significant values) were calculated between environmental measures and species richness, diversity and abundance of live Foraminifera. Secondly, a distance-based linear model (DistLM) (Clarke and Gorley, 2006 and Anderson et al., 2008) was computed in an attempt to define those environmental variables that were most responsible for structuring the multivariate Foraminifera data. DistLM first conducts a marginal test, which determines the proportion of the variance in the distribution pattern of the foraminifera that can

be explained by each environmental variable, before portioning the variation according to a multiple regression model (step-wise), in order to provide a “best” solution (adjusted R2) for a combination of the environmental variables. A distance-based redundancy analysis (dbRDA) was then used to visualise the fitted model, where the length of the

vector overlays depicts the effect each variable had on the construction of the dbRDA axes. Note that because % N was only determined per site, and Sulfite dehydrogenase not per sample core, these data were not used in the above analyses. However, the average data (across cores) for all environmental variables and foraminiferal abundances per site were analysed together with the % N and these results were compared to those generated as described above: all are available in the Supplementary data. The nMMDS plot reveals that the physico-chemical environment at the two sampling locations was quite distinct and the overall stress value was sufficiently low (0.07) to allow ready interpretation (Fig. 1 and Supplementary data Fig. 4). While there was a difference between pipeline and non-pipeline sites in TB, this was less clear in SHB where a greater variability was observed. The results of the PERMANOVA indicate no significant differences in the environment between the two study areas (Table 1) but that significant differences were apparent between pipeline and non-pipeline sites: note the high level of intra-site sample variation (Table 1).

2C and F). However, envenomed neonate rats showed a 83.1% increase in the water channel AQP4 expression at 2 h (**p ≤ 0.01), a 58.8% increase at 5 h (**p ≤ 0.01) and a 23.5% non-significant increase at 24 h indicating that after an immediate rise the expression of AQP4 declined with time toward baseline. On the selleck kinase inhibitor other hand, relative to controls PNV-administered adult rats showed a 59.8% increase of AQP4 expression at 2 h (*p ≤ 0.05), 39.5% (not significant) at 5 h and 91.8% at 24 h (*p ≤ 0.05) indicating a prolonged

effect of PNV on the expression of the protein ( Fig. 3C). GFAP expression showed no significant change in response selleck inhibitor to PNV in P14 animals; however, in adult rats it induced a 71.2% increase at 2 h (***p ≤ 0.001) and 33.5% at 5 h (*p ≤ 0.05) and was close to baseline at 24 h ( Fig. 3F). The two-way analysis of variance showed that with regard to the granular layer the variable time after injection interfered in the expression of AQP4 (***p ≤ 0.001) and GFAP (***p ≤ 0.05) in neonates and AQP4 and GFAP (***p ≤ 0.001) in adults. Also, there was interaction between the

age variable and PNV treatment in the expression of AQP4 at 2 h (***p ≤ 0.001), 5 h (**p ≤ 0.01) and 24 h (**p ≤ 0.01) and GFAP at all time intervals (**p ≤ 0.01; *p ≤ 0.05; *p ≤ 0.05, respectively). The smallest value of AQP4 expression in Bergmann glia cells for neonate was 15.73 ± 2.61 and for adult rats was 16.39 ± 1.62, whereas the highest value was 23.95 ± 2.16 for neonates and 22.96 ± 3.45 for adults (Fig. 4C). The expression of GFAP was slightly higher in P14 animals than in

the adults ranging from 23.53 ± 2.19 to 29.31 ± 2.16 in P14 and 20.23 ± 1.51 to 23.83 ± 2.46 in adults (Fig. 4F). The effect of PNV on AQP4 expression was significant only after 24 h when a 52% upregulation was found for Bergman glia of 8-week-old rats (*p ≤ 0.05) ( Fig. 4C). In contrast, in 14-day-old rats a 44.2% Levetiracetam increase occurred earlier at 2 h (*p ≤ 0.05), but its level did not differ from the control at 5 h and then increased 101.6% at 24 h (*p ≤ 0.01) relative to the baseline ( Fig. 4C). GFAP expression showed no alteration in P14, whereas it rose significantly by 66.34% at 2 h (***p ≤ 0.001), 51.11% at 5 h (**p ≤ 0.01) and 58.59 at 24 h (**p ≤ 0.01) above baseline counterparts ( Fig. 4F). The two-way analysis of variance showed that the time elapsed between envenomation and animal euthanasia interfered with the expression of AQP4 in P14 (***p ≤ 0.001) and GFAP in adults (*p ≤ 0.05). Also, the age variable interacted with PNV treatment relative to AQP4 expression at 24 h (***p ≤ 0.001) and GFAP expression at 2 h (**p ≤ 0.01). Fig.

In our slices treatment with EtOH did not result in enhanced cytokine production. It seems likely that this treatment was not strong enough to induce an inflammatory cascade in the nbM. Indeed, http://www.selleckchem.com/products/isrib-trans-isomer.html EtOH-induced inflammation in humans has been shown after chronic alcoholism and is not

a short time effect. In addition, cytokines found in the brains of individuals after heavy EtOH consume are originally produced by the liver cells (Crews and Nixon, 2009). Thus, any lack of direct EtOH on inflammation is in line with such a peripheral inflammatory process. In hippocampal–entorhinal brain slice cultures EtOH induced inflammatory gene expression (Zou and Crews, 2010), suggesting that this region may be more sensitive to EtOH-induced cytokine upregulation than the nbM. Further studies are necessary to investigate if the lack of inflammation in our slice model is area-related or a methodological limitation. Taken together, our data show that EtOH-induced a decline of cholinergic neurons in vitro, which was partly counteracted by NGF. Inhibition of MAPK p38 and NOS ameliorated the EtOH effect suggesting a role in the underlying mechanism of EtOH-mediated effects in vitro. In conclusion, the data may suggest that direct EtOH exposure to cholinergic nbM neurons may transiently

suppress the enzyme ChAT and may not induce cell SRT1720 death of cholinergic neurons, but rather may reflect a form of neuronal plasticity in response to EtOH. Cholinergic neurons in organotypic brain slices were cultured, as described in detail previously (Humpel and Weis, 2002 and Weis et al., 2001). Briefly, the basal nucleus of Meynert (nbM) of postnatal day 10 (P10) rats else was dissected under aseptic conditions. Further, 400 μm slices were cut with a tissue chopper (McIlwain, USA) and placed on 30 mm Millicell-CM 0.4 μm pore membrane culture plate inserts (6–8 slices per membrane). It needs to be pointed out that a single experiment included approximately

8–12 pups. In one dissection experiment 4 pups were dissected and all brain slices were randomly distributed on all 6-wells. An experiment was normally repeated 3 times on different groups, so that a single treatment contained at least slices from 9 different rat pups. Slices were cultured in 6-well plates at 37 °C and 5% CO2 with 1.2 ml/well of slice medium (50% MEM/HEPES (Gibco), 25% heat inactivated horse serum (Gibco/Lifetech, Austria), 25% Hanks’ solution (Gibco), 2 mM NaHCO3 (Merck, Austria), 6.5 mg/ml glucose (Merck), 2 mM glutamine (Merck), pH 7.2) including 10 ng/ml nerve growth factor (NGF) for 2 weeks. It is well established that the 400 μm brain slices become thinner during the 2 weeks of incubation resulting in a thickness of approx. 100 μm, which is a sign of healthy cultures. Slices, which did not flatten were immediately removed from the experiments.

10). Of the 404 sequence of dams, 73% are closer than 100 km to each other. Results show that the 512 km

between the Garrison and Oahe Dam is not enough distance to consider these dams separately. We propose a conceptual model of how a sequence of interacting dams might impact river geomorphology (Fig. 11) based on our results. We call this morphologic sequence the Inter-Dam Sequence, and we present a simplified model based on the Upper Missouri River that could be easily adapted to other river reaches. Although the morphologic sequence is a useful conceptualization, there are clear limitations to these results. SCH727965 This model is likely only applies to large Volasertib dams on alluvial rivers. Dams on rivers that are controlled by bedrock or where morphologic adjustment is limited by vegetation or cohesive banks may respond completely different than the model presented here. Similarly, the downstream effects of small dams will likely attenuate

over much shorter distances. However, this framework is a helpful advancement in our understanding of longitudinal responses to multiple dams. One of the greatest influences that humans have had on the fluvial landscape is the construction of dams. Despite significant advancements in the study of the downstream and upstream impacts of dams, they are often considered separately from each other. The Garrison and Oahe Dama on the Missouri River are used to demonstrate ADAMTS5 that the effects of an upstream dam maintains significant geomorphic control over river morphology as the backwater effects of downstream reservoir begin to occur. The upstream–downstream interaction of multiple dams overlap to create a distinct morphologic sequence.

Five unique geomorphic gradational reaches were identified for the Garrison Reach, two of which are controlled solely by the upstream dam and three of which are controlled by the dam interaction termed: Dam Proximal, Dam-Attenuating, River-Dominated Interaction, Reservoir-Dominated Interaction, and Reservoir. A conceptual model was developed of a morphologic sequence of downstream dam impacts and dam interaction which can be adapted to other rivers. The current distribution of dams on the major rivers in the U.S. indicates that more than 80% of large rivers may have interacting between their dams. Given this widespread occurrence, we describe a generalized morphologic sequence termed the Inter-Dam Sequence and suggest it should be the focus of additional research. We would like to acknowledge project funding from the following sources: U.S. Army Corps of Engineers, ND State Water Commission, ND Department of Transportation, ND Game and Fish Department, ND Department of Health, City of Bismark, City of Mandan, Burleigh County WRB, Morton County WRB, and Lower Hart WRB.