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Vertebrate corpse decomposition provides an important stage in nutrient cycling in most terrestrial habitats, yet microbially mediated processes are poorly understood. Here we combine deep microbial community characterization, community-level metabolic reconstruction, and soil biogeochemical assessment to understand the principles governing microbial community assembly during decomposition of mouse and human corpses on different soil substrates. We find a suite of bacterial and fungal groups that contribute to nitrogen cycling and a reproducible network of decomposers that emerge on predictable time scales. Our results show that this decomposer community is derived primarily from bulk soil, but key decomposers are ubiquitous in low abundance. Soil type was not a dominant factor driving community development, and the process of decomposition is sufficiently reproducible to offer new opportunities for forensic investigations., (Copyright © 2016, American Association for the Advancement of Science.)

Little is known about how variables, such as carcass mass, affect the succession pattern of microbes in soils during decomposition. To investigate the effects of carcass mass on the soil microbial community, soils associated with swine (Sus scrofa domesticus) carcasses of four different masses were sampled until the 15th day of decomposition during the month of June in a pasture near Lincoln, Nebraska. Soils underneath swine of 1, 20, 40, and 50 kg masses were investigated in triplicate, as well as control sites not associated with a carcass. Soil microbial communities were characterized by sequencing the archaeal, bacterial (16S), and eukaryotic (18S) rRNA genes in soil samples. We conclude that time of decomposition was a significant influence on the microbial community, but carcass mass was not. The gravesoil associated with 1 kg mass carcasses differs most compared to the gravesoil associated with other carcass masses. We also identify the 15 most abundant bacterial and eukaryotic taxa, and discuss changes in their abundance as carcass decomposition progressed. Finally, we show significant decreases in alpha diversity for carcasses of differing mass in pre-carcass rupture (days 0, 1, 2, 4, 5, and 6 postmortem) versus post-carcass rupture (days 9 and 15 postmortem) microbial communities.

Body-associated microbes were recently shown to change significantly during decomposition, undergoing an ecological succession in experimental conditions using rodent and swine models. We investigated microbial succession in soils associated with swine carcasses under experimental field conditions in summer and winter. We demonstrate that these postmortem microbial communities change in a specific, reproducible fashion, and that soil microbes represent a significant component of the postmortem microbial community, contrary to widespread belief in forensic science. However, the effects of decomposition on soil microbial communities were different in summer and winter. We suggest that the microbial ecological succession will be useful in medicolegal death investigation; however, observations in winter might not be applicable to summer, which indicates a need for a greater understanding of the seasonality of decomposition.

Understanding the evolution of Australia's extinct marsupial megafauna has been hindered by a relatively incomplete fossil record and convergent or highly specialized morphology, which confound phylogenetic analyses. Further, the harsh Australian climate and early date of most megafaunal extinctions (39-52 ka) means that the vast majority of fossil remains are unsuitable for ancient DNA analyses. Here, we apply cross-species DNA capture to fossils from relatively high latitude, high altitude caves in Tasmania. Using low-stringency hybridization and high-throughput sequencing, we were able to retrieve mitochondrial sequences from two extinct megafaunal macropodid species. The two specimens, Simosthenurus occidentalis (giant short-faced kangaroo) and Protemnodon anak (giant wallaby), have been radiocarbon dated to 46-50 and 40-45 ka, respectively. This is significantly older than any Australian fossil that has previously yielded DNA sequence information. Processing the raw sequence data from these samples posed a bioinformatic challenge due to the poor preservation of DNA. We explored several approaches in order to maximize the signal-to-noise ratio in retained sequencing reads. Our findings demonstrate the critical importance of adopting stringent processing criteria when distant outgroups are used as references for mapping highly fragmented DNA. Based on the most stringent nucleotide data sets (879 bp for S. occidentalis and 2,383 bp for P. anak), total-evidence phylogenetic analyses confirm that macropodids consist of three primary lineages: Sthenurines such as Simosthenurus (extinct short-faced kangaroos), the macropodines (all other wallabies and kangaroos), and the enigmatic living banded hare-wallaby Lagostrophus fasciatus (Lagostrophinae). Protemnodon emerges as a close relative of Macropus (large living kangaroos), a position not supported by recent morphological phylogenetic analyses., (© The Authors 2014. Published by Oxford University Press on behalf of Molecular Biology and Evolution. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

A recent report warns that DNA extraction kits and other laboratory reagents are considerable sources of contamination in microbiome experiments. The issue of contamination is particularly problematic for samples of low biomass.

In a series of 4 experiments, we tested children's understanding that the causes of their actions must necessarily be attributed to information known prior to (i.e., "pre-action" information), rather than after (i.e., "post-action" information), the completion of their actions. For example, children were shown a dog, asked to get some cheese to feed the dog, and then returned to discover a mouse. In Experiment 1, the majority of 3-, 4-, and 5-year-olds claimed that they had gotten the cheese to feed the mouse. In Experiments 2 and 3, we ruled out the possibilities that (1) children had forgotten the critical "pre-action" information (e.g., "dog"), and (2) children had merely attributed the cause of their action to the most recent item (e.g., "dog") that they had seen. Finally, in Experiment 4, we determined that 7-year-olds, but not 6-year-olds, correctly attributed the cause of their action to the pre-action information, suggesting that this is the age at which children are no longer influenced by associative post-action information when explaining the causes of their actions. These results are discussed in terms of their relevance for causal reasoning, action explanation, and memory., (PsycINFO Database Record (c) 2014 APA, all rights reserved.)

The bacteria that colonize humans and our built environments have the potential to influence our health. Microbial communities associated with seven families and their homes over 6 weeks were assessed, including three families that moved their home. Microbial communities differed substantially among homes, and the home microbiome was largely sourced from humans. The microbiota in each home were identifiable by family. Network analysis identified humans as the primary bacterial vector, and a Bayesian method significantly matched individuals to their dwellings. Draft genomes of potential human pathogens observed on a kitchen counter could be matched to the hands of occupants. After a house move, the microbial community in the new house rapidly converged on the microbial community of the occupants' former house, suggesting rapid colonization by the family's microbiota., (Copyright © 2014, American Association for the Advancement of Science.)

Carrion decomposition is an ecologically important natural phenomenon influenced by a complex set of factors, including temperature, moisture, and the activity of microorganisms, invertebrates, and scavengers. The role of soil microbes as decomposers in this process is essential but not well understood and represents a knowledge gap in carrion ecology. To better define the role and sources of microbes in carrion decomposition, lab-reared mice were decomposed on either (i) soil with an intact microbial community or (ii) soil that was sterilized. We characterized the microbial community (16S rRNA gene for bacteria and archaea, and the 18S rRNA gene for fungi and microbial eukaryotes) for three body sites along with the underlying soil (i.e., gravesoils) at time intervals coinciding with visible changes in carrion morphology. Our results indicate that mice placed on soil with intact microbial communities reach advanced stages of decomposition 2 to 3 times faster than those placed on sterile soil. Microbial communities associated with skin and gravesoils of carrion in stages of active and advanced decay were significantly different between soil types (sterile versus untreated), suggesting that substrates on which carrion decompose may partially determine the microbial decomposer community. However, the source of the decomposer community (soil- versus carcass-associated microbes) was not clear in our data set, suggesting that greater sequencing depth needs to be employed to identify the origin of the decomposer communities in carrion decomposition. Overall, our data show that soil microbial communities have a significant impact on the rate at which carrion decomposes and have important implications for understanding carrion ecology., (Copyright © 2014, American Society for Microbiology. All Rights Reserved.)

A new study explores the ancient oral microbiome from the well-preserved dental calculus samples of four human individuals who lived during medieval times, using a suite of genomic, proteomic and microscopic approaches. The authors investigate the evolution of dental pathogens by reconstructing the genome of the periodontal pathogen Tannerella forsythia and also identify antibiotic resistance genes, bacterial virulence factors and host immune defense proteins.

Mammals have diversified into many dietary niches. Specialized myrmecophagous (ant- and termite-eating) placental mammals represent a textbook example of evolutionary convergence driven by extreme diet specialization. Armadillos, anteaters, aardvarks, pangolins and aardwolves thus provide a model system for understanding the potential role of gut microbiota in the convergent adaptation to myrmecophagy. Here, we expand upon previous mammalian gut microbiome studies by using high-throughput barcoded Illumina sequencing of the 16S rRNA gene to characterize the composition of gut microbiota in 15 species representing all placental myrmecophagous lineages and their close relatives from zoo- and field-collected samples. We confirm that both diet and phylogeny drive the evolution of mammalian gut microbiota, with cases of convergence in global composition, but also examples of phylogenetic inertia. Our results reveal specialized placental myrmecophages as a spectacular case of large-scale convergence in gut microbiome composition. Indeed, neighbour-net networks and beta-diversity plots based on UniFrac distances show significant clustering of myrmecophagous species (anteaters, aardvarks and aardwolves), even though they belong to phylogenetically distant lineages representing different orders. The aardwolf, which diverged from carnivorous hyenas only in the last 10 million years, experienced a convergent shift in the composition of its gut microbiome to become more similar to other myrmecophages. These results confirm diet adaptation to be a major driving factor of convergence in gut microbiome composition over evolutionary timescales. This study sets the scene for future metagenomic studies aiming at evaluating potential convergence in functional gene content in the microbiomes of specialized mammalian myrmecophages., (© 2013 John Wiley & Sons Ltd.)

DNA repair is essential for maintaining genomic stability, and defects in this process significantly increase the risk of cancer. Clear-cell renal cell carcinoma (CCRCC) caused by inactivation of the von Hippel-Lindau (VHL) tumor suppressor gene is characterized by high genomic instability. However, the molecular mechanism underlying the association between the loss of VHL and genomic instability remains unclear. Here, we show that suppressor of cytokine signaling 1 (SOCS1) promotes nuclear redistribution and K63-ubiquitylation of VHL in response to DNA double-strand breaks (DSBs). Loss of VHL or VHL mutations that compromise its K63-ubiquitylation attenuates the DNA-damage response (DDR), resulting in decreased homologous recombination repair and persistence of DSBs. These results identify VHL as a component of the DDR network, inactivation of which contributes to the genomic instability associated with CCRCC.

One of the grand goals of historical biogeography is to understand how and why species' population sizes and distributions change over time. Multiple types of data drawn from disparate fields, combined into a single modelling framework, are necessary to document changes in a species's demography and distribution, and to determine the drivers responsible for change. Yet truly integrated approaches are challenging and rarely performed. Here, we discuss a modelling framework that integrates spatio-temporal fossil data, ancient DNA, palaeoclimatological reconstructions, bioclimatic envelope modelling and coalescence models in order to statistically test alternative hypotheses of demographic and potential distributional changes for the iconic American bison (Bison bison). Using different assumptions about the evolution of the bioclimatic niche, we generate hypothetical distributional and demographic histories of the species. We then test these demographic models by comparing the genetic signature predicted by serial coalescence against sequence data derived from subfossils and modern populations. Our results supported demographic models that include both climate and human-associated drivers of population declines. This synthetic approach, integrating palaeoclimatology, bioclimatic envelopes, serial coalescence, spatio-temporal fossil data and heterochronous DNA sequences, improves understanding of species' historical biogeography by allowing consideration of both abiotic and biotic interactions at the population level.

Establishing the time since death is critical in every death investigation, yet existing techniques are susceptible to a range of errors and biases. For example, forensic entomology is widely used to assess the postmortem interval (PMI), but errors can range from days to months. Microbes may provide a novel method for estimating PMI that avoids many of these limitations. Here we show that postmortem microbial community changes are dramatic, measurable, and repeatable in a mouse model system, allowing PMI to be estimated within approximately 3 days over 48 days. Our results provide a detailed understanding of bacterial and microbial eukaryotic ecology within a decomposing corpse system and suggest that microbial community data can be developed into a forensic tool for estimating PMI. DOI:http://dx.doi.org/10.7554/eLife.01104.001.

Large-scale characterization of the human microbiota has largely focused on Western adults, yet these populations may be uncharacteristic because of their diets and lifestyles. In particular, the rise of "Western diseases" may in part stem from reduced exposure to, or even loss of, microbes with which humans have coevolved. Here, we review beneficial microbes associated with pathogen resistance, highlighting the emerging role of complex microbial communities in protecting against disease. We discuss ways in which modern lifestyles and practices may deplete physiologically important microbiota, and explore prospects for reintroducing or encouraging the growth of beneficial microbes to promote the restoration of healthy microbial ecosystems., (Copyright © 2013 WILEY Periodicals, Inc.)

In the last two decades, the widespread application of genetic and genomic approaches has revealed a bacterial world astonishing in its ubiquity and diversity. This review examines how a growing knowledge of the vast range of animal-bacterial interactions, whether in shared ecosystems or intimate symbioses, is fundamentally altering our understanding of animal biology. Specifically, we highlight recent technological and intellectual advances that have changed our thinking about five questions: how have bacteria facilitated the origin and evolution of animals; how do animals and bacteria affect each other's genomes; how does normal animal development depend on bacterial partners; how is homeostasis maintained between animals and their symbionts; and how can ecological approaches deepen our understanding of the multiple levels of animal-bacterial interaction. As answers to these fundamental questions emerge, all biologists will be challenged to broaden their appreciation of these interactions and to include investigations of the relationships between and among bacteria and their animal partners as we seek a better understanding of the natural world.

Many species are threatened with extinction and efforts are underway worldwide to restore imperilled species to their native ranges. Restoration requires knowledge of species' historical diversity and distribution. For some species, many populations were extirpated or individuals moved beyond their native range before native diversity and distribution were documented, resulting in a lack of accurate information for establishing restoration goals. Moreover, traditional taxonomic assessments often failed to accurately capture phylogenetic diversity. We illustrate a general approach for estimating regional native diversity and distribution for cutthroat trout in the Southern Rocky Mountains. We assembled a large archive of historical records documenting human-mediated change in the distribution of cutthroat trout (Oncorhynchus clarkii) and combined these data with phylogenetic analysis of 19th century samples from museums collected prior to trout stocking activities and contemporary DNA samples. Our study of the trout in the Southern Rocky Mountains uncovered six divergent lineages, two of which went extinct, probably in the early 20th century. A third lineage, previously declared extinct, was discovered surviving in a single stream outside of its native range. Comparison of the historical and modern distributions with stocking records revealed that the current distribution of trout largely reflects intensive stocking early in the late 19th and early 20th century from two phylogenetically and geographically distinct sources. Our documentation of recent extinctions, undescribed lineages, errors in taxonomy and dramatic range changes induced by human movement of fish underscores the importance of the historical record when developing and implementing conservation plans for threatened and endangered species., (© 2012 Blackwell Publishing Ltd.)

The processes by which cells sense and respond to ambient oxygen concentration are fundamental to cell survival and function, and they commonly target gene regulatory events. To date, however, little is known about the link between the microRNA pathway and hypoxia signaling. Here, we show in vitro and in vivo that chronic hypoxia impairs Dicer (DICER1) expression and activity, resulting in global consequences on microRNA biogenesis. We show that von Hippel-Lindau-dependent down-regulation of Dicer is key to the expression and function of hypoxia-inducible factor α (HIF-α) subunits. Specifically, we show that EPAS1/HIF-2α is regulated by the Dicer-dependent microRNA miR-185, which is down-regulated by hypoxia. Full expression of hypoxia-responsive/HIF target genes in chronic hypoxia (e.g. VEGFA, FLT1/VEGFR1, KDR/VEGFR2, BNIP3L, and SLC2A1/GLUT1), the function of which is to regulate various adaptive responses to compromised oxygen availability, is also dependent on hypoxia-mediated down-regulation of Dicer function and changes in post-transcriptional gene regulation. Therefore, functional deficiency of Dicer in chronic hypoxia is relevant to both HIF-α isoforms and hypoxia-responsive/HIF target genes, especially in the vascular endothelium. These findings have relevance to emerging therapies given that we show that the efficacy of RNA interference under chronic hypoxia, but not normal oxygen availability, is Dicer-dependent. Collectively, these findings show that the down-regulation of Dicer under chronic hypoxia is an adaptive mechanism that serves to maintain the cellular hypoxic response through HIF-α- and microRNA-dependent mechanisms, thereby providing an essential mechanistic insight into the oxygen-dependent microRNA regulatory pathway.

Rapidly developing sequencing methods and analytical techniques are enhancing our ability to understand the human microbiome, and, indeed, how the microbiome and its constituents are defined. This review highlights recent research that expands our ability to understand the human microbiome on different spatial and temporal scales, including daily time series datasets spanning months. Furthermore, emerging concepts related to defining operational taxonomic units, diversity indices, core versus transient microbiomes, and the possibility of enterotypes are discussed. Additional advances in sequencing technology and in our understanding of the microbiome will provide exciting prospects for exploiting the microbiota for personalized medicine., (© 2012 International Life Sciences Institute.)

Until recently, the study of microbial diversity has mainly been limited to descriptive approaches, rather than predictive model-based analyses. The development of advanced analytical tools and decreasing cost of high-throughput multi-omics technologies has made the later approach more feasible. However, consensus is lacking as to which spatial and temporal scales best facilitate understanding of the role of microbial diversity in determining both public and environmental health. Here, we review the potential for combining these new technologies with both traditional and nascent spatio-temporal analysis methods. The fusion of proper spatio-temporal sampling, combined with modern multi-omics and computational tools, will provide insight into the tracking, development and manipulation of microbial communities., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Oxygen is essential for eukaryotic life and is inextricably linked to the evolution of multicellular organisms. Proper cellular response to changes in oxygen tension during normal development or pathological processes, such as cardiovascular disease and cancer, is ultimately regulated by the transcription factor, hypoxia-inducible factor (HIF). Over the past decade, unprecedented molecular insight has been gained into the mammalian oxygen-sensing pathway involving the canonical oxygen-dependent prolyl-hydroxylase domain-containing enzyme (PHD)-von Hippel-Lindau tumour suppressor protein (pVHL) axis and its connection to cellular metabolism. Here we review recent notable advances in the field of hypoxia that have shaped a more complex model of HIF regulation and revealed unique roles of HIF in a diverse range of biological processes, including immunity, development and stem cell biology.

Advances in DNA sequencing have allowed us to characterize microbial communities--including those associated with the human body--at a broader range of spatial and temporal scales than ever before. We can now answer fundamental questions that were previously inaccessible and use well-tested ecological theories to gain insight into changes in the microbiome that are associated with normal development and human disease. Perhaps unsurprisingly, the ecosystems associated with our body follow trends identified in communities at other sites and scales, and thus studies of the microbiome benefit from ecological insight. Here, we assess human microbiome research in the context of ecological principles and models, focusing on diversity, biological drivers of community structure, spatial patterning and temporal dynamics, and suggest key directions for future research that will bring us closer to the goal of building predictive models for personalized medicine.

The rich fossil record of the family Equidae (Mammalia: Perissodactyla) over the past 55 MY has made it an icon for the patterns and processes of macroevolution. Despite this, many aspects of equid phylogenetic relationships and taxonomy remain unresolved. Recent genetic analyses of extinct equids have revealed unexpected evolutionary patterns and a need for major revisions at the generic, subgeneric, and species levels. To investigate this issue we examine 35 ancient equid specimens from four geographic regions (South America, Europe, Southwest Asia, and South Africa), of which 22 delivered 87-688 bp of reproducible aDNA mitochondrial sequence. Phylogenetic analyses support a major revision of the recent evolutionary history of equids and reveal two new species, a South American hippidion and a descendant of a basal lineage potentially related to Middle Pleistocene equids. Sequences from specimens assigned to the giant extinct Cape zebra, Equus capensis, formed a separate clade within the modern plain zebra species, a phenotypicically plastic group that also included the extinct quagga. In addition, we revise the currently recognized extinction times for two hemione-related equid groups. However, it is apparent that the current dataset cannot solve all of the taxonomic and phylogenetic questions relevant to the evolution of Equus. In light of these findings, we propose a rapid DNA barcoding approach to evaluate the taxonomic status of the many Late Pleistocene fossil Equidae species that have been described from purely morphological analyses.

Tumor hypoxia is associated with disease progression, resistance to conventional cancer therapies and poor prognosis. Hypoxia, by largely unknown mechanisms, leads to deregulated accumulation of and signaling via receptor tyrosine kinases (RTKs) that are critical for driving oncogenesis. Here, we show that hypoxia or loss of von Hippel-Lindau protein--the principal negative regulator of hypoxia-inducible factor (HIF)--prolongs the activation of epidermal growth factor receptor that is attributable to lengthened receptor half-life and retention in the endocytic pathway. The deceleration in endocytosis is due to the attenuation of Rab5-mediated early endosome fusion via HIF-dependent downregulation of a critical Rab5 effector, rabaptin-5, at the level of transcription. Primary kidney and breast tumors with strong hypoxic signatures show significantly lower expression of rabaptin-5 RNA and protein. These findings reveal a general role of the oxygen-sensing pathway in endocytosis and support a model in which tumor hypoxia or oncogenic activation of HIF prolongs RTK-mediated signaling by delaying endocytosis-mediated deactivation of receptors.

Nitration of N(alpha),N(1)-bis(trifluoroacetyl)-l-tryptophan methyl ester with HNO(3) in acetic anhydride at 0 degrees C provides N(alpha)-trifluoroacetyl-2-nitro-l-tryptophan methyl ester in 67% yield, whereas nitration in trifluoroacetic acid at 0 degrees C gives N(alpha)-trifluoroacetyl-6-nitro-l-tryptophan methyl ester in 69% yield.

Chromosomal abnormalities, such as structural and numerical abnormalities, are a common occurrence in cancer. The close association of homologous chromosomes during interphase, a phenomenon termed somatic chromosome pairing, has been observed in cancerous cells, but the functional consequences of somatic pairing have not been established. Gene expression profiling studies revealed that somatic pairing of chromosome 19 is a recurrent chromosomal abnormality in renal oncocytoma, a neoplasia of the adult kidney. Somatic pairing was associated with significant disruption of gene expression within the paired regions and resulted in the deregulation of the prolyl-hydroxylase EGLN2 [corrected] a key protein that regulates the oxygen-dependent degradation of hypoxia-inducible factor (HIF). Overexpression of EGLN2 [corrected] in renal oncocytoma increased ubiquitin-mediated destruction of HIF and concomitantly suppressed the expression of several HIF-target genes, including the pro-death BNIP3L gene. The transcriptional changes that are associated with somatic pairing of chromosome 19 mimic the transcriptional changes that occur following DNA amplification. Therefore, in addition to numerical and structural chromosomal abnormalities, alterations in chromosomal spatial dynamics should be considered as genomic events that are associated with tumorigenesis. The identification of EGLN2 as a significantly deregulated gene that maps within the paired chromosome region directly implicates defects in the oxygen-sensing network to the biology of renal oncocytoma., Competing Interests: The authors have declared that no competing interests exist.

Newly formed hybrid populations provide an opportunity to examine the initial consequences of secondary contact between species and identify genetic patterns that may be important early in the evolution of hybrid inviability. Widespread introductions of rainbow trout (Oncorhynchus mykiss) into watersheds with native cutthroat trout (Oncorhynchus clarkii) have resulted in hybridization. These introductions have contributed to the decline of native cutthroat trout populations. Here, we examine the pattern of hybridization between introduced rainbow trout and 2 populations of cutthroat trout native to Colorado. For this study, we utilized 7 diagnostic, codominant nuclear markers and a diagnostic mitochondrial marker to investigate hybridization in a population of greenback cutthroat trout (Oncorhynchus clarkii stomias) and a population of Colorado River cutthroat trout (Oncorhynchus clarkii pleuriticus). We infer that cutthroat-rainbow trout hybrid swarms have formed in both populations. Although a mixture of hybrid genotypes was present, not all genotype combinations were detected at expected frequencies. We found evidence that mitochondrial DNA introgression in hybrids is asymmetric and more likely from rainbow trout than from cutthroat trout. A difference in spawning time of the 2 species or differences in the fitness between the reciprocal crosses may explain the asymmetry. Additionally, the presence of intraspecific cytonuclear associations found in both populations is concordant with current hypotheses regarding coevolution of mitochondrial and nuclear genomes.

Accurate assessment of species identity is fundamental for conservation biology. Using molecular markers from the mitochondrial and nuclear genomes, we discovered that many putatively native populations of greenback cutthroat trout (Oncorhynchus clarkii stomias) comprised another subspecies of cutthroat trout, Colorado River cutthroat trout (Oncorhynchus clarkii pleuriticus). The error can be explained by the introduction of Colorado River cutthroat trout throughout the native range of greenback cutthroat trout in the late 19th and early 20th centuries by fish stocking activities. Our results suggest greenback cutthroat trout within its native range is at a higher risk of extinction than ever before despite conservation activities spanning more than two decades.

We examined the mating frequencies of queens in a social wasp, Dolichovespula arenaria (Vespinae) using DNA microsatellites. Five of the seven colonies supported the hypothesis of single mating by queens. The other two colonies supported two and three matings, with effective paternity of 1.48 and 1.91. Mean worker relatedness was 0.77 +/- 0.09. In two of the four male-containing colonies, all were likely progeny of the queen. In the other two colonies workers produced 8 and 14% of the male wasps. Overall, 94.3% of the male wasps were likely progeny of the queen. These patterns are consistent with published studies of vespine wasps.

Endometriosis, a disease of women and nonhuman primates in which endometrial tissue grows outside the uterus, can be mimicked surgically in rats and mice. The disease is related to infertility in women, and surgical induction of endometriotic lesions reduces fertility in rats according to some studies. Conversely, pregnancy appears to have a beneficial effect on endometriosis in women and some rat studies. Our objective was to evaluate a new mouse model of surgically induced endometriosis with respect to the effects of pregnancy on endometriosis and the effects of endometriosis on pregnancy. Female B6C3F1 mice were divided into four groups. Those in Group A and B underwent induction surgery for endometriosis and hemi-ovariectomy, those in Group C underwent sham surgery and hemi-ovariectomy, and animals in Group D received no surgery at all. Three weeks later, Group A, C, and D were bred. Eighteen days later one half of the dams in Groups A, B, and C only were sacrificed, and evaluations included endometriotic lesion diameter, number of pups, fetal weight, and various organ weights. The remaining dams delivered, and, 18 days after parturition, dams and pups were sacrificed. Evaluations included gestation length and those listed above. Endometriotic lesion diameter was significantly reduced in pregnant animals when compared with nonpregnant controls, but the reduction was not a full regression. Lactation returned the mean lesion diameter to pre-pregnancy dimensions. When effects of endometriosis on pregnancy were evaluated, no effects on the litter size, pup weight, or gestation length were found, but trends toward increased resorptions and malformations were evident. Thus, in the mouse model of induced endometriosis, pregnancy produced a significant reduction in endometriotic lesion diameter while fertility was largely unaffected by the surgically induced endometriosis. The mouse model of endometriosis thus appears more resistant than the rat model to effects of endometriosis on fertility.

Epidermal growth factor (EGF) and its receptor (EGF-R) have been implicated as mediators for estrogen induced cellular growth. This study examines whether the action of the estrogenic pesticide methoxychlor (MXC) parallels the action of 17 beta-estradiol (E2) on uterine EGF-R. Administration of 20 micrograms E2/sexually immature female rat increased 125I-EGF binding to membranes extracted from whole uteri 175% over endogenous levels, while 500 mg MXC/kg led to a 156% increase. E2 in both 20 and 40 micrograms/rat doses and 500 mg MXC/kg led to maximal stimulation over endogenous levels, 12-h posttreatment. Rats were treated with E2, MXC, or vehicle plus 100 micrograms actinomycin-D (ACT-D) or 100 micrograms cycloheximide (CYCLO) per rat to determine if mRNA transcription and translation are involved in the increased EGF-R binding following estrogenic treatment. Only ACT-D inhibited the estrogenic stimulation of EGF-R binding, resulting in a 44% decrease when given concurrently with E2 or MXC, suggesting transcription is required. Additionally, ACT-D decreased endogenous receptor levels by 55%. No other differences were detected. When EGF-R binding data were analyzed by the method of Scatchard, both E2 and MXC, at maximal dosages, elevated uterine EGF-R binding sites by over 200% after 12 h as measured by maximal binding (Bmax) with no significant difference in dissociation constant (Kd) values. These results demonstrate that both E2 and MXC can stimulate the number of EGF-R binding sites without significantly altering the receptor binding affinity (Kd). Further, this stimulation is time dependent and is affected by dose.

In the disease of endometriosis, endometrial tissue grows outside the uterus, usually in the peritoneal cavity. Rodent models of endometriosis allow a way to reproduce the disease, evaluate effects of chemicals, and study mechanisms. Twenty-one days prior to induction surgery which produces endometriosis, female Sprague-Dawley rats and B6C3F1 mice were pretreated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) at 0, 3 or 10 micrograms TCDD/kg. Animals were treated again at the time of surgery and at 3, 6, and 9 weeks following surgery. Evaluations were made at 3, 6, 9, and 12 weeks postsurgery. TCDD produced a dose-dependent increase in endometriotic site diameter when all time points were pooled within each dose in rats and a dramatic increase in site diameter in mice at 9 and 12 weeks. In rats but not mice, ovarian weight was decreased at 9 and 12 weeks, the occurrence of persistent vaginal estrus was increased at these times, and histological evaluation of the ovaries revealed ovulatory arrest at 12 weeks. In both species, thymic antrophy, indicating immune dysfunction, and hepatomegaly were observed as consequences of TCDD exposure. Body weight was reduced in rats but not in mice. Histological evaluations of endometriotic sites revealed fibrosis in control rats, necrotic and inflammatory changes in the sites from TCDD-treated rats, and predominantly fibrotic changes in sites from TCDD-treated mice. Differences observed between the rat and the mouse with respect to (a) the magnitude of the effect on endometrial site diameter (rats < mice), (b) measured effects on ovarian function (rats > mice) that may be based on the partial antiestrogenicity of TCDD, and (c) evidence that mice and rats differ in their immune response to TCDD suggest that the mechanisms mediating TCDD's action to promote endometriosis are complex and may be different in rats and mice. The mouse may be a better model for future studies necessary to elucidate these mechanisms.

Endometriosis is a disease of women where endometrial tissue is found growing at ectopic sites. While evidence suggesting a role for the ovarian hormones in endometriosis exists, no complete studies of the roles of estrogen and progesterone have heretofore been performed. Also, if estrogen has a role in the growth and/or maintenance of endometriosis, it is likely that the proestrogenic pesticide, methoxychlor (MXC), might also have such an effect. Sixty rats underwent surgery on Day 0 to induce endometriosis. On Day 21, all rats were ovariectomized. During surgery, the diameters of all endometriotic implants (which were fully developed) were measured. Starting on Day 21, groups of rats were treated daily, for 3 weeks, with (a) vehicle (b) estrone, 1 micrograms/rat, E;(c) progesterone, 2 mg/rat, P; (d) E + P, 1 micrograms + 2 mg; (e) MXC, 250 mg/kg; or (f) MXC + P, 250 mg/kg + 2 mg/rat. On Day 42, all rats were killed, and the diameters of all endometriotic sites were measured. While no differences in diameter were found across groups prior to ovariectomy, ovariectomy plus treatment altered the growth of endometriosis tissue. Progesterone and vehicle treatments produced results that were identical: regression of endometriotic sites. Both estrogen and MXC treatments maintained endometriotic site size at a level greater than that in the vehicle-treated group. The combination of progesterone with either estrone or MXC did not alter the effect of either chemical. We conclude that while estrogen promotes the growth of endometriosis, progesterone either produces regression or fails to maintain the sites. MXC, at a relatively high dose, supports the development of endometriosis. Concurrent progesterone treatment does not modulate the effects of estrone or MXC. These results suggest that exposure of women to high doses of MXC may exacerbate the development of endometriosis or contribute to its recurrence.

Endometriosis consists of the growth of endometrial tissue outside the uterus. A rat model of endometriosis is available to evaluate the potential for environmental chemicals to promote the disease but may be relatively insensitive for the evaluation of the hazard of certain compounds. Our objective, which was to develop a mouse model for endometriosis, was based on (a) the promotion of endometriosis in primates by 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD), (b) the apparent relationship between endometriosis and immunodeficiency, and (c) evidence that humoral immunity is suppressed in mice but not rats following TCDD exposure. In the mouse model, slices of uterus were sutured to intestinal mesenteric vessels. By 3 weeks after surgery, these sites were cyst-like structures. The growth of the sites was hormone dependent. In intact mice, sites measured 3.60 +/- 0.22 mm; vehicle and estrone (0.5 microgram/day) treatments produced site diameters of 0.95 +/- 0.128 and 5.28 +/- 0.355 mm, respectively. This new mouse model provides a sensitive and useful technique for future studies of the potential for specific xenobiotics to promote the development of endometriosis.

Methoxychlor (MXC), a proestrogenic pesticide, has adverse effects on fertility and uterine function in rodents. MXC is converted to an estrogenic substance, 2,2-bis-(p-hydroxyphenyl)-1,1,1-trichloroethane (HPTE), which binds to the estrogen receptor. We examined the similarities in mechanism between MXC (500 mg/kg) and estrogen (10 micrograms/rat) actions using the estrogen-induced protein, IP, also known as creatine kinase. Immature, female rats were treated with MXC or estradiol (E2). Concurrent treatment included actinomycin D (100 micrograms), cycloheximide (100 micrograms), or progesterone (0.5 mg). Uterine proteins were labeled in vitro with 3H for treated rats and with 14C for controls. The uteri were combined, cytosol was isolated, non-denaturing (ND) gels were run, and dpm/gel slice were plotted. In a follow-up study, labeled cytosols from MXC- and E2-treated rats were immunoprecipitated with a monoclonal antibody to creatine kinase. The immunoprecipitate was run on SDS gels. The data show that both MXC and E2 treatments result in ND gels with peaks in (a) induced protein and (b) the 3H/14C ratio, in the same position. The induction of IP by MXC is time- and dose-dependent. Concurrent treatment with MXC plus actinomycin D or cycloheximide blocked IP synthesis, a result parallel to E2 action signifying the necessity of RNA and protein synthesis for IP induction. Progesterone did not block either MXC or E2 induction of IP synthesis. Immunoprecipitation of creatine kinase revealed a single peak at a molecular weight of approximately 49,000. SDS gels of cytosol after MXC or E2 treatment also yielded protein and ratio peaks at molecular weights of approximately 49,000. This estimate is near the published estimated molecular weight of creatine kinase of 46,000. We conclude that MXC action parallels that of estradiol on the induction and regulation of the estrogen-induced protein in immature rat uterus.

Methoxychlor (MXC) has adverse effects on fertility and rat uteri via its active metabolite HPTE (2,2-bis(p-hydroxyphenyl)-1,1,1-trichloroethane). Uterine peroxidase, a marker of estrogen action, was used to probe potential mechanisms of MXC's adverse effects. Specifically, our objective was to compare the regulation of the effects of estrogen and MXC on uterine peroxidase. Immature female rats were treated with MXC (250 mg/kg; gavage) 24 h prior to the measurement of uterine peroxidase activity, with or without concurrent treatment with actinomycin D, cycloheximide, progesterone, or tamoxifen. MXC alone produced an increase in peroxidase activity. The prior and/or concurrent treatment with the compounds listed blocked the MXC-induced stimulation of peroxidase. These data show similarities between the mechanisms of estrogen MXC action. Both estrogen and MXC act to stimulate uterine peroxidase activity via increased RNA and protein synthesis and this stimulation can be blocked by progesterone and tamoxifen.

When high-birefringent fibers are used in polarization-sensitive devices, one must ensure the proper alignment of the optical axes. One nondestructive alignment method involves squeezing the fiber and noting the change in the resultant birefringent axes. We use the finite-element method to determine the change in the principal stress direction, and, hence, the extrinsic birefringent axis orientation, as a function of the fiber coating eccentricity. Fiber eccentricity can shift the extrinsic birefringent axis by more than 1.5 degrees , whch becomes important in high-performance device applications.