Several C-β-d-glucopyranosyl azoles have recently been uncovered as among the most potent glycogen phosphorylase (GP) catalytic site inhibitors discovered to date. Toward further exploring their translational potential, ex vivo experiments have been performed for their effectiveness in reduction of glycogenolysis in hepatocytes. New compounds for these experiments were predicted in silico where, for the first time, effective ranking of GP catalytic site inhibitor potencies using the molecular mechanics-generalized Born surface area (MM-GBSA) method has been demonstrated. For a congeneric training set of 27 ligands, excellent statistics in terms of Pearson (RP) and Spearman (RS) correlations (both 0.98), predictive index (PI = 0.99), and area under the receiver operating characteristic curve (AU-ROC = 0.99) for predicted versus experimental binding affinities were obtained, with ligand tautomeric/ionization states additionally considered using density functional theory (DFT). Seven 2-aryl-4(5)-(β-d-glucopyranosyl)-imidazoles and 2-aryl-4-(β-d-glucopyranosyl)-thiazoles were subsequently synthesized, and kinetics experiments against rabbit muscle GPb revealed new potent inhibitors with best Ki values in the low micromolar range (5c = 1.97 μM; 13b = 4.58 μM). Ten C-β-d-glucopyranosyl azoles were then tested ex vivo in mouse primary hepatocytes. Four of these (5a-c and 9d) demonstrated significant reduction of glucagon stimulated glycogenolysis (IC50 = 30-60 μM). Structural and predicted physicochemical properties associated with their effectiveness were analyzed with permeability related parameters identified as crucial factors. The most effective ligand series 5 contained an imidazole ring, and the calculated pKa (Epik: 6.2; Jaguar 5.5) for protonated imidazole suggests that cellular permeation through the neutral state is favored, while within the cell, there is predicted more favorable binding to GP in the protonated form.

Understanding how ambulance incidents are spatially distributed can shed light to the epidemiological dynamics of geographic areas and inform healthcare policy design. Here we analyze a longitudinal dataset of more than four million ambulance calls across a region of twelve million residents in the North West of England. With the aim to explain geographic variations in ambulance call frequencies, we employ a wide range of data layers including open government datasets describing population demographics and socio-economic characteristics, as well as geographic activity in online services such as Foursquare. Working at a fine level of spatial granularity we demonstrate that daytime population levels and the deprivation status of an area are the most important variables when it comes to predicting the volume of ambulance calls at an area. Foursquare check-ins on the other hand complement these government sourced indicators, offering a novel view to population nightlife and commercial activity locally. We demonstrate how check-in activity can provide an edge when predicting certain types of emergency incidents in a multi-variate regression model.

3-(β-d-Glucopyranosyl)-5-substituted-1,2,4-triazoles have been revealed as an effective scaffold for the development of potent glycogen phosphorylase (GP) inhibitors but with the potency very sensitive to the nature of the alkyl/aryl 5-substituent (Kun et al., Eur. J. Med. Chem. 2014, 76, 567). For a training set of these ligands, quantum mechanics-polarized ligand docking (QM-PLD) demonstrated good potential to identify larger differences in potencies (predictive index PI = 0.82) and potent inhibitors with Ki's < 10 μM (AU-ROC = 0.86). Accordingly, in silico screening of 2335 new analogues exploiting the ZINC docking database was performed and nine predicted candidates selected for synthesis. The compounds were prepared in O-perbenzoylated forms by either ring transformation of 5-β-d-glucopyranosyl tetrazole by N-benzyl-arenecarboximidoyl chlorides, ring closure of C-(β-d-glucopyranosyl)formamidrazone with aroyl chlorides, or that of N-(β-d-glucopyranosylcarbonyl)arenethiocarboxamides by hydrazine, followed by deprotections. Kinetics experiments against rabbit muscle GPb (rmGPb) and human liver GPa (hlGPa) revealed five compounds as potent low μM inhibitors with three of these on the submicromolar range for rmGPa. X-ray crystallographic analysis sourced the potency to a combination of favorable interactions from the 1,2,4-triazole and suitable aryl substituents in the GP catalytic site. The compounds also revealed promising calculated pharmacokinetic profiles.

Dipteran larvae from a number of families feed in aggregations. Rotation of blow fly (Diptera: Calliphoridae) larvae within an aggregation has been reported anecdotally many times. However, there is a lack of quantitative data on such larval movement, which is necessary to better understand the advantage of this gregarious behaviour. A recent development in tagging methods provided an opportunity to address this gap in knowledge. In 15 aggregations of 500 Lucilia sericata (Meigen) (Diptera: Calliphoridae) larvae, the location of four-tagged individuals was recorded at 10-minute intervals. All larvae were seen to rotate, alternating between the periphery and within. There was much variation in the relative proportions that larvae were seen in these two locations among aggregations ((2)=78.4, df=58, P=0.038), perhaps as a result of differences in mass shape and, therefore, surface area: volume ratio. There were also differences between larvae within aggregations ((2)=25.6, df=14, P=0.029), which may give rise to differences in development rate, perhaps as a result of intraspecific competition. Further work would be required to verify this competition, and to establish whether the limited resource is temperature, food, oxygen, or some other requirement.

As a result of criminal activity, human bodies are sometimes dismembered and concealed within sealed, plastic waste sacks. Consequently, due to the inhibited ingress of insects and dismemberment, the rate of decomposition of the body parts within may be different to that of whole, exposed bodies. Correspondingly, once found, an estimation of the postmortem interval may be affected and lead to erroneous inferences. This study set out to determine whether insects were excluded and how rate of decomposition was affected inside such plastic sacks. The limbs, torsos and heads of 24 dismembered pigs were sealed using nylon cable ties within plastic garbage sacks, half of which were of a type claimed to repel insects. Using a body scoring scale to quantify decomposition, the body parts in the sacks were compared to those of ten exposed, whole pig carcasses. Insects were found to have entered both types of plastic sack. There was no difference in rate of decomposition in the two types of sack (F 1,65 = 1.78, p = 0.19), but this was considerably slower than those of whole carcasses (F 1,408 = 1453, p < 0.001), with heads showing the largest differences. As well as a slower decomposition, sacks resulted in formation of some adipocere tissue as a result of high humidity within. Based upon existing methods, postmortem intervals for body parts within sealed sacks would be significantly underestimated.

Megyesi et al.'s (J Forensic Sci, 2005, 50, 618) paper was important to forensic anthropology as it introduced a quantitative framework for estimating time since death in human cadavers, based upon physical appearance by way of scoring on a novel scale. However, errors concerning rounding, temperature scale, and incorrect use of a statistical regression model render their predictive formula unusable. Based upon only their more reliable data, a more appropriate regression model to predict accumulated degree days (ADD) from total body score (TBS) is presented. The new model is also a superior fit (r(2) = 0.91) and produces markedly narrower confidence intervals than the original, which also allowed impossible, negative ADD values. Explanations of the shortcomings in the original analysis and calculations are presented, which it is hoped will help forensic scientists avoid making similar mistakes.

The length or stage of development of blow fly (Diptera: Calliphoridae) larvae may be used to estimate a minimum postmortem interval, often by targeting the largest individuals of a species in the belief that they will be the oldest. However, natural variation in rate of development, and therefore length, implies that the size of the largest larva, as well as the number of larvae longer than any stated length, will be greater for larger cohorts. Length data from the blow flies Protophormia terraenovae and Lucilia sericata were collected from one field-based and two laboratory-based experiments. The field cohorts contained considerably more individuals than have been used for reference data collection in the literature. Cohorts were shown to have an approximately normal distribution. Summary statistics were derived from the collected data allowing the quantification of errors in development time which arise when different sized cohorts are compared through their largest larvae. These errors may be considerable and can lead to overestimation of postmortem intervals when making comparisons with reference data collected from smaller cohorts. This source of error has hitherto been overlooked in forensic entomology.

Accurate prediction of the postmortem interval requires an understanding of the decomposition process and the factors acting upon it. A controlled experiment, over 60days at an outdoor site in the northwest of England, used 20 freshly killed pigs (Sus scrofa) as human analogues to study decomposition rate and pattern. Ten pigs were hung off the ground and ten placed on the surface. Observed differences in the decomposition pattern required a new decomposition scoring scale to be produced for the hanging pigs to enable comparisons with the surface pigs. The difference in the rate of decomposition between hanging and surface pigs was statistically significant (p=0.001). Hanging pigs reached advanced decomposition stages sooner, but lagged behind during the early stages. This delay is believed to result from lower variety and quantity of insects, due to restricted beetle access to the aerial carcass, and/or writhing maggots falling from the carcass.

Phosphorylase kinase (PhK) has been linked with a number of conditions such as glycogen storage diseases, psoriasis, type 2 diabetes and more recently, cancer (Camus et al., 2012 [6]). However, with few reported structural studies on PhK inhibitors, this hinders a structure based drug design approach. In this study, the inhibitory potential of 38 indirubin analogues have been investigated. 11 of these ligands had IC50 values in the range 0.170-0.360μM, with indirubin-3'-acetoxime (1c) the most potent. 7-Bromoindirubin-3'-oxime (13b), an antitumor compound which induces caspase-independent cell-death (Ribas et al., 2006 [20]) is revealed as a specific inhibitor of PhK (IC50=1.8μM). Binding assay experiments performed using both PhK-holo and PhK-γtrnc confirmed the inhibitory effects to arise from binding at the kinase domain (γ subunit). High level computations using QM/MM-PBSA binding free energy calculations were in good agreement with experimental binding data, as determined using statistical analysis, and support binding at the ATP-binding site. The value of a QM description for the binding of halogenated ligands exhibiting σ-hole effects is highlighted. A new statistical metric, the 'sum of the modified logarithm of ranks' (SMLR), has been defined which measures performance of a model for both the "early recognition" (ranking earlier/higher) of active compounds and their relative ordering by potency. Through a detailed structure activity relationship analysis considering other kinases (CDK2, CDK5 and GSK-3α/β), 6'(Z) and 7(L) indirubin substitutions have been identified to achieve selective PhK inhibition. The key PhK binding site residues involved can also be targeted using other ligand scaffolds in future work.

Twenty Sus scrofa carcasses were used to study the effect the presence of clothing had on decomposition rate and colonization locations of Diptera species; 10 unclothed control carcasses were compared to 10 clothed experimental carcasses over 58 days. Data collection occurred at regular accumulated degree day intervals; the level of decomposition as Total Body Score (TBSsurf), pattern of decomposition, and Diptera present was documented. Results indicated a statistically significant difference in the rate of decomposition, (t(427) = 2.59, p = 0.010), with unclothed carcasses decomposing faster than clothed carcasses. However, the overall decomposition rates from each carcass group are too similar to separate when applying a 95% CI, which means that, although statistically significant, from a practical forensic point of view they are not sufficiently dissimilar as to warrant the application of different formulae to estimate the postmortem interval. Further results demonstrated clothing provided blow flies with additional colonization locations.

Journal of Forensic SciencesVolume 59, Issue 4 p. 1161-1161 Letter to the Editor Commentary on: Humphreys MK, Panacek E, Green W, Albers E. Comparison of protocols for measuring and calculating postmortem submersion intervals for human analogs in fresh water. J Forensic Sci 2013;58(2):513-7 Colin Moffatt Ph.D., Colin Moffatt Ph.D. [email protected] School of Forensic & Investigative Sciences, University of Central Lancashire, Preston, PR6 8RJ U.K.Search for more papers by this authorTal Simmons Ph.D., Tal Simmons Ph.D. School of Forensic & Investigative Sciences, University of Central Lancashire, Preston, PR6 8RJ U.K.Search for more papers by this author Colin Moffatt Ph.D., Colin Moffatt Ph.D. [email protected] School of Forensic & Investigative Sciences, University of Central Lancashire, Preston, PR6 8RJ U.K.Search for more papers by this authorTal Simmons Ph.D., Tal Simmons Ph.D. School of Forensic & Investigative Sciences, University of Central Lancashire, Preston, PR6 8RJ U.K.Search for more papers by this author First published: 27 June 2014 https://doi.org/10.1111/1556-4029.12498Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onEmailFacebookTwitterLinkedInRedditWechat No abstract is available for this article. References 1Humphreys MK, Panacek E, Green W, Albers E. Comparison of protocols for measuring and calculating postmortem submersion intervals for human analogs in fresh water. J Forensic Sci 2013; 58(2): 513–7. 2Heaton V, Lagden A, Moffatt C, Simmons T. Predicting the post-mortem submersion interval for human remains recovered from U.K. waterways. J Forensic Sci 2010; 55(2): 302–7. 3Megyesi MS, Nawrocki SP, Haskell NH. Using accumulated degree-days to estimate the postmortem interval for decomposed human remains. J Forensic Sci 2005; 50: 1–9. 4Simmons T, Adlam RE, Moffatt C. Debugging decomposition data — comparative taphonomic studies and the influence of insects and carcass size on decomposition rate. J Forensic Sci 2010; 55(1): 8–13. Volume59, Issue4July 2014Pages 1161-1161 ReferencesRelatedInformation

Five pairs of mass graves, each containing carcasses of 21 rabbits, were used to examine differential decomposition at four locations within the burial: surface, deep, mid-outer, and core. Every 100 accumulated degree days (ADD), a pair of graves was exhumed, and total body score (TBS) and internal carcass temperature of each rabbit were recorded. Decomposition did not differ for core- and deep-positioned carcasses (p = 0.13); differences were significant (p < 0.001) for all other location comparisons. Decomposition occurred fastest in shallow carcasses, followed by mid-outer carcasses; deep and core carcasses decomposed slowest and at rates not significantly different from one another. Adipocere formation was minimal and confined to deep carcasses. Carcass location within the mass grave significantly influenced internal carcass temperature (p < 0.001); a mean internal temperature difference of ca. 1°C existed between deep and shallow carcasses (30 cm apart). Effects of mass compactness and oxygenation require further investigation.

Numerous Calliphoridae species have been observed to form larval aggregations during the feeding stage of development, resulting in localized increases in temperature. This study investigates the relationship between maggot numbers in a mass and heat generation. Single-species aggregations (Lucilia sericata) of various sizes (50-2500 individuals) were reared in the laboratory at a constant ambient temperature of 22 degrees C. Internal mass temperatures were recorded every 5min throughout the feeding stage of development. Results showed that mass temperatures increased with mass numbers (p-value <0.001), ranging from 2.5 to 14 degrees C above ambient. A minimum mass size of 1200 produced overall temperatures that were significantly warmer than ambient, diverging away from 22 degrees C after c. 26h. These results indicate that the microclimate of a mass has the potential to differ significantly from ambient, which may be influencing larval development rates and should therefore be factored into mPMI estimates to increase accuracy.

From each of four Arabic countries; Morocco, Tunisia, Jordan and Oman, 150 participants produced handwriting samples which were examined to assess whether national characteristics were discernible. Ten characters, which have different configurations depending upon their position in the word, along with one short word, were classified into distinguishable forms, and these forms recorded for each handwriting sample. Tests of independence showed that character forms used were not independent of country (p<0.001) for all but one character-position (this was dropped from subsequent analyses). A correspondence analysis ordination plot and analysis of similarity (R=0.326, p=0.0002) showed that whole samples were discernibly grouped by country, and a tree analysis produced a classification which was 71% accurate for the original data and 83% accurate for 80 new handwriting samples that underwent ‘blind’ classification. When the countries were combined into two regions, North Africa and Middle East, the grouping was more marked. Thus, there appears to be some scope for narrowing down the nationality, and particularly the wider geographical region of an author based upon the character forms they use in Arabic handwriting.

In this study, we have developed a PCR multiplex that can be used to assess DNA degradation and at the same time monitor for inhibition: primers have been designed to amplify human, pig, and rabbit DNA, allowing pig and rabbit to be used as experimental models for taphonomic research, but also enabling studies on human DNA persistence in forensic evidence. Internal amplified controls have been added to monitor for inhibition, allowing the effects of degradation and inhibition to be differentiated. Sequence data for single-copy nuclear recombination activation gene (RAG-1) from human, pig, and rabbit were aligned to identify conserved regions and primers were designed that targeted amplicons of 70, 194, 305, and 384 bp. Robust amplification in all three species was possible using as little as 0.3 ng of template DNA. These have been combined with primers that will amplify a bacterial DNA template within the PCR. The multiplex has been evaluated in a series of experiments to gain more knowledge of DNA persistence in soft tissues, which can be important when assessing what material to collect following events such as mass disasters or conflict, when muscle or bone material can be used to aid with the identification of human remains. The experiments used pigs as a model species. When whole pig bodies were exposed to the environment in Northwest England, DNA in muscle tissue persisted for over 24 days in the summer and over 77 days in the winter, with full profiles generated from these samples. In addition to time, accumulated degree days (ADD) were also used as a measure that combines both time and temperature24 days was in summer equivalent to 295 ADD whereas 77 days in winter was equivalent to 494 ADD.

Visible Implant Elastomer (VIE) has previously been used successfully to tag individuals in a variety of marine and amphibious animals, earthworms, and scorpions. Visible Implant Elastomer tags were injected into third instars of the blow fly Calliphora vicina Robineau-Desvoidy (Diptera: Calliphoridae) to test its compatibility and retention across life stages. Injecting into the dorsal midline of the 11th segment (seventh abdominal segment) produced survival rates of 80%, with no significant difference in the subsequent rate of development (z=0.21, P=0.83) as compared with untagged insects. Tags remained visible and allowed identification of individuals within a feeding, intermingling aggregation (maggot mass), especially when a high-contrast fluorescent colour was used. Tags were retained across life-stage changes and were easily found in dissected adults.

The synthesis of a variety of N-alkylated 2,3,3-trimethylindolenines and 2-methylbenzothiazoles is reported herein. Their potential as antifungal agents is evaluated by preliminary screening against Saccharomyces cerevisiae (S. cerevisiae), Schizosaccharomyces pombe (S. pombe), and Candida albicans (C. albicans). Statistical analyses illustrate a strong relationship between chain length and growth inhibition for S. cerevisiae and S. pombe (p < 0.0001 in every case). Of particular interest is the activity of both sets of compounds against S. cerevisiae, as this is emerging as an opportunistic pathogen, especially in immunosuppressed and immunocompromised patients. Bioassays were set up to compare the efficacy of our range of N-alkylated compounds against classic antifungal agents; Amphotericin B and Thiabendazole.