A randomized phase 2 study of MK-2206 versus everolimus in refractory renal cell carcinoma.

Background: Activation of the phosphoinisitide-3 kinase (PI3K) pathway through mutation and constitutive upregulation has been described in renal cell carcinoma (RCC), making it an attractive target for therapeutic intervention. We performed a randomized phase II study in vascular endothelial growth factor (VEGF) therapy refractory patients to determine whether MK-2206, an allosteric inhibitor of AKT, was more efficacious than the mammalian target of rapamycin inhibitor everolimus. Patients and methods: A total of 43 patients were randomized in a 2:1 distribution, with 29 patients assigned to the MK-2206 arm and 14 to the everolimus arm. Progression-free survival (PFS) was the primary endpoint. Results: The trial was closed at the first futility analysis with an observed PFS of 3.68 months in the MK-2206 arm and 5.98 months in the everolimus arm. Dichotomous response rate profiles were seen in the MK-2206 arm with one complete response and three partial responses in the MK-2206 arm versus none in the everolimus arm. On the other hand, progressive disease was best response in 44.8% of MK2206 versus 14.3% of everolimus-treated patients. MK-2206 induced significantly more rash and pruritis than everolimus, and dose reduction occurred in 37.9% of MK-2206 versus 21.4% of everolimus-treated patients. Genomic analysis revealed that 57.1% of the patients in the PD group had either deleterious TP53 mutations or ATM mutations or deletions. In contrast, none of the patients in the non-PD group had TP53 or ATM defects. No predictive marker for response was observed in this small dataset. Conclusions: Dichotomous outcomes are observed when VEGF therapy refractory patients are treated with MK-2206, and MK-2206 does not demonstrate superiority to everolimus. Additionally, mutations in DNA repair genes are associated with early disease progression, indicating that dysregulation of DNA repair is associated with a more aggressive tumor phenotype in RCC.

Characterization of Peroxidase Changes in Tolerant and Susceptible Soybeans Challenged by Soybean Aphid (Hemiptera: Aphididae).

Changes in protein content, peroxidase activity, and isozyme profiles in response to soybean aphid feeding were documented at V1 (fully developed leaves at unifoliate node, first trifoliate leaf unrolled) and V3 (fully developed leaf at second trifoliate node, third trifoliate leaf unrolled) stages of soybean aphid-tolerant (KS4202) and -susceptible (SD76R) soybeans. Protein content was similar between infested and control V1 and V3 stage plants for both KS4202 and SD76R at 6, 16, and 22 d after aphid introduction. Enzyme kinetics studies documented that control and aphid-infested KS4202 V1 stage and SD76R V1 and V3 stages had similar levels of peroxidase activity at the three time points evaluated. In contrast, KS4202 aphid-infested plants at the V3 stage had significantly higher peroxidase activity levels than control plants at 6 and 22 d after aphid introduction. The differences in peroxidase activity observed between infested and control V3 stage KS4202 plants at these two time points suggest that peroxidases may be playing multiple roles in the tolerant plant. Native gels stained for peroxidase were able to detect differences in the isozyme profiles of aphid-infested and control plants for both KS4202 and SD76R.

Evaluation of late vegetative and reproductive stage soybeans for resistance to soybean aphid (Hemiptera: Aphididae).

The soybean aphid, Aphis glycines Matsumura, has become the most significant soybean [Glycine max (L.) Merrill] insect pest in the north central soybean production region of North America. The objectives of this research were to measure selected genotypes for resistance to the soybean aphid in the later vegetative and reproductive stages under field conditions, and confirm the presence of tolerance in KS4202. The results from 2007 to 2011 indicate that KS4202 can support aphid populations with minimal yield loss at levels where significant yield loss would be expected in most other genotypes. The common Nebraska cultivar, 'Asgrow 2703', appears to show signs of tolerance as well. None of the yield parameters were significantly different between the aphid infested and noninfested treatments. Based on our results, genotypes may compensate for aphid feeding in different ways. Asgrow 2703 appears to produce a similar number of seeds as its noninfested counterpart, although the seeds produced are slightly smaller. Field evaluation of tolerance in KS4202 indicated a yield loss of only 13% at 34,585-53,508 cumulative aphid-days, when 24-36% yield loss would have been expected.

Effects of thiamethoxam seed treatments on soybean aphid (Hemiptera: Aphididae) feeding behavior.

Since its discovery in North America in 2000, the soybean aphid, Aphis glycines Matsumura (Hemiptera: Aphididae), has rapidly become an important pest of soybean [Glycine max (L.) Merrill], sometimes resulting in significant yield losses. Previous research has documented the toxicity of neonicotinoid seed treatments to soybean aphids, but control under field conditions has been inconsistent. Imidacloprid, a popular neonicotinoid insecticide, has been shown to exhibit antifeedant effects on aphids. Antifeedant activity has not been demonstrated for other neonicotinoids, including thiamethoxam. This research investigated the effects of a thiamethoxam seed treatment on soybean aphid feeding behavior by using electronic penetration graphs (EPG) to visualize stylet penetration behavior. Soybean aphid feeding behavior was assessed for 9 h on thiamethoxam-treated and untreated soybeans (V2 and V4 stages). Because results were inconclusive from initial experiments, a study was conducted to document the effects of thiamethoxam-treated soybeans on soybean aphid survival. The seed treatment was shown to negatively affect aphid survival at 4, 8, and 11 d after aphid introduction. A subsequent EPG study then was designed to document soybean aphid feeding behavior for 15 h, after an initial exposure of 9 h to thiamethoxam-treated soybeans. In this study, the exposed aphids exhibited significant differences in feeding behavior compared with those aphids feeding on untreated soybeans. Soybean aphids on thiamethoxam-treated soybeans spent significantly less time feeding in the sieve element phase, with a greater duration of nonprobing events. These studies suggest soybean aphids are unable to ingest phloem sap, which may be another important element in seed treatment protection.

Reliability and Validity Testing of the Persian Version of the Derriford Appearance Scale 24 in a Sample of Individuals with Craniofacial Irregularity and Amputation.

BACKGROUND: Despite the recent advancements in the design and manufacture of prostheses for individuals with craniofacial irregularity and amputation, these individuals tend to become self-conscious about their appearance. The aim of this study was to investigate the reliability and validity of Persian version of the Derriford Appearance Scale24 (P-DAS24) for a sample of individuals with craniofacial irregularity and limb loss. METHODOLOGY: Reliability of the P-DAS24 was determined by computing internal consistency and test-retest reliability utilizing Cronbach's alpha coefficient and Pearson's correlation coefficient. Discriminant validity was investigated with comparing the total score of the P-DAS24 between disfigured participants and those with no appearance problem. Known-groups validity was evaluated regarding the participants' gender and their level of involvement. FINDINGS: The sample size comprised of 251 individuals with disfigurement and 101 without disfigurement who were deemed normal in appearance. The P-DAS24 showed satisfactory internal consistency (Cronbach's alpha = 0.89) and excellent test-retest reliability (r = 0.96). The total score of the P-DAS24 showed a statistically significant difference between individuals deemed disfigured or normal (P=0.01). The total scores P-DAS24 in individuals with different levels of involvement were significantly different (P<0.001). The scores of the DAS2, DAS18, DAS21, and DAS24 were significantly different between men and women (P<0.01, <0.01, 0.03, and 0.01, respectively). CONCLUSION: The P-DAS24 is a valid and reliable tool that may be utilized in clinical practice and researches to assess the outcomes of prosthetic reconstructions in individuals with disfigurement.

Evaluation of the effectiveness of Face IT, a computer-based psychosocial intervention for disfigurement-related distress.

Some people who have a visible difference (disfigurement) experience psychosocial adjustment problems that can lead to social anxiety and isolation. The aim of this study was to assess the effectiveness of a new computerised CBT-based intervention (Face IT) in reducing anxiety and appearance-related distress for individuals with visible differences. Face IT was tested against a non-intervention control group and standard CBT-based face-to-face delivery. Eighty-three participants were assessed at four time points using the Hospital Anxiety and Depression Scales, Derriford Appearance Scale-24, Body Image Quality of Life Inventory and Fear of Negative Evaluation (FNE). The findings indicate a significant reduction in anxiety and appearance-related distress in both the Face IT intervention and the face-to-face condition. Similar findings were reported for depressive symptoms and FNE. Results at the three and six months follow-up demonstrate increased improvements in psychological functioning with both interventions. This new online psychosocial intervention has been found to be effective at reducing anxiety, depression and appearance concerns amongst individuals with disfigurements, whilst increasing positive adjustment. A remote-access, computer-based intervention offers the potential to provide psychosocial support more easily and in a cost-effective manner to adults with appearance-related distress.

Dose-response relationships of clothianidin, imidacloprid, and thiamethoxam to Blissus occiduus (Hemiptera: Blissidae).

The western chinch bug, Blissus occiduus Barber (Hemiptera: Blissidae), has emerged as a serious pest of buffalograss, Buchlod dactyloides (Nuttall) Engelmann. In general, neonicotinoid insecticides effectively control a variety of turfgrass insects, particularly phloem-feeding pests. However, because of well documented inconsistencies in control, these compounds are generally not recommended for chinch bugs. This study was designed to document the contact and systemic toxicity of three neonicotinoid insecticides (clothianidin, imidacloprid, and thiamethoxam) to B. occiduus. In contact bioassays, thiamethoxam was approximately 20-fold less toxic than clothianidin or imidacloprid to B. occiduus nymphs and three-fold more toxic to adults. In adult systemic bioassays, thiamethoxam was up to five-fold more toxic than clothianidin or imidacloprid. Interestingly, thiamethoxam was significantly more toxic to adults than to nymphs in both contact and systemic bioassays. This was not observed with clothianidin or imidacloprid. Bifenthrin, used for comparative purposes, exhibited 1844-fold and 122-fold increase in toxicity to nymphs and adults, respectively. These results provide the first documentation of the relative toxicity of these neonicotinoid insecticides to B. occiduus.

Categorizing the resistance of soybean genotypes to the soybean aphid (Hemiptera: Aphididae).

We evaluated selected soybean, Glycine max (L.) Merr., genotypes during their reproductive stages for resistance to the soybean aphid, Aphis glycines Matsumura (Hemiptera: Aphididae), under greenhouse conditions and documented the categories of aphid-resistant soybean. Two screening studies were performed to assess the level of resistance to the soybean aphid on six soybean genotypes during the reproductive stages of development. Significant differences in aphid damage ratings were detected among the soybean evaluated in the screening studies. Three genotypes (KS4202, K-1639-2, and K1621) were considered moderately resistant based on the assessed damage ratings. Two of these genotypes (K-1639-2 and KS4202), along with a commercial variety ('Asgrow 2703') were used in a follow-up greenhouse study to test for antibiosis and tolerance. For the antibiosis evaluation, KS4202 had significantly more nymphs than Asgrow 2703 and K-1639-2. In fact, KS4202 had a threefold difference in the number of nymphs compared with Asgrow 2703 (81.8 +/- 14.7 and 26.2 +/- 13.9 nymphs, respectively) and a fivefold difference compared with K-1639-2 (15.6 +/- 13.9). Although not significant, Asgrow 2703 had more nymphs than K-1639-2. The lower aphid numbers on infested K-1639-2 plants compared with aphid numbers on Asgrow 2703 and KS4202 plants indicates antibiosis for this genotype. No significant differences in average seed weight, number of seeds per pod, or plant damage were observed between infested and control KS4202 plants; however, significant differences in biomass, total seed weight, number of pods per plant, and number of seeds per plant were detected.

Short-range DNA looping by the Xenopus HMG-box transcription factor, xUBF.

Xenopus UBF (xUBF) interacts with DNA by way of multiple HMG-box domains. When xUBF binds to the ribosomal promoter, the carboxyl-terminal acidic tail and amino-terminal HMG-box interact. Binding also leads to negative DNA supercoiling and the formation of a disk-like structure, the enhancesome. Within the enhancesome, an xUBF dimer makes a low-density protein core around which DNA is looped into a single 180-base pair turn, probably by in-phase bending. The enhancesome structure suggests a mechanism for xUBF-dependent recruitment of the TATA box-binding protein complex without direct interaction between the two factors.

Gene expression profiling of tolerant barley in response to Diuraphis noxia (Hemiptera: Aphididae) feeding.

Aphids are, arguably, the single most damaging group of agricultural insect pests throughout the world. Plant tolerance, which is a plant response to an insect pest, is viewed as an excellent management strategy. Developing testable hypotheses based on genome-wide and more focused methods will help in understanding the molecular underpinnings of plant tolerance to aphid herbivory. As a first step in this process, we undertook transcript profiling with Affymetrix GeneChip Barley Genome arrays using RNA extracted from tissues of tolerant and susceptible genotypes collected at three hours, three days and six days after Diuraphis noxia introduction. Acquired data were compared to identify changes unique to the tolerant barley at each harvest date. Transcript abundance of 4086 genes was differentially changed over the three harvest dates in tolerant and susceptible barley in response to D. noxia feeding. Across the three harvest dates, the greatest number of genes was differentially expressed in both barleys at three days after aphid introduction. A total of 909 genes showed significant levels of change in the tolerant barley in response to D. noxia feeding as compared to susceptible plants infested with aphids. Many of these genes could be assigned to specific metabolic categories, including several associated with plant defense and scavenging of reactive oxygen species (ROS). Interestingly, two peroxidase genes, designated HvPRXA1 and HvPRXA2, were up-regulated to a greater degree in response to D. noxia feeding on tolerant barley plants, indicating that specific peroxidases could be important for the tolerance process. These findings suggest that the ability to elevate and sustain levels of ROS-scavenging enzymes could play an important role in the tolerant response.

The carotid bodies influence growth responses to moderate maternal undernutrition in late-gestation fetal sheep.

OBJECTIVE: To determine the role of carotid sinus innervation on differential fetal organ growth during maternal nutrient restriction in late pregnancy. DESIGN: Randomised controlled study. SETTING: University research facility. SAMPLE: Thirty-nine Merino ewes. METHODS: At 113 days gestational age (dGA), fetuses were bilaterally carotid sinus denervated or sham denervated. From 118 dGA, the surgery groups were subdivided into two dietary groups, and their ewes were fed 100% of nutrient requirements or 50% until tissue collection at 140 dGA. This provided four groups (sham/control diet, sham/restricted diet, denervated/control diet and denervated/restricted diet). MAIN OUTCOME MEASURES: Fetal organ weights and hormone levels and maternal weight change during the dietary restriction. RESULTS: Adrenal glands were larger in sham/restricted diet fetuses than in sham/control diet or denervated/restricted diet fetuses (P < 0.05). Fetal adrenal weight and brain-to-liver weight ratio were positively related to maternal weight change during the nutritional challenge in sham fetuses only (P < 0.05). Fetal liver weight was negatively related to maternal weight change during nutritional challenge in sham fetuses only (P < 0.05). CONCLUSIONS: We have shown a reduction in liver growth but sparing of adrenal growth in response to moderate maternal undernutrition, which is dependent on intact carotid body innervation. This suggests a new role for the carotid bodies in the control of differential organ growth during such undernutrition.

Hereditary multi-infarct dementia of the Swedish type is a novel disorder different from NOTCH3 causing CADASIL.

Several hereditary small vessel diseases (SVDs) of the brain have been reported in recent years. In 1977, Sourander and Wålinder described hereditary multi-infarct dementia (MID) in a Swedish family. In the same year, Stevens and colleagues reported chronic familial vascular encephalopathy in an English family bearing a similar phenotype. These disorders have invariably been suggested to be cerebral autosomal dominant arteriopathy with subcortical infarcts and leucoencephalopathy (CADASIL) but their genetic identities remain unknown. We used molecular, radiological and neuropathological methods to characterize these disorders. Direct DNA sequencing unexpectedly confirmed that affected members of the English family carried the R141C mutation in the NOTCH3 gene diagnostic of CADASIL. However, we did not detect any pathogenic mutations in the entire 8091 bp reading frame of NOTCH3 or find clear evidence for NOTCH3 gene linkage in the Swedish DNA. This was consistent with the lack of hyperintense signals in the anterior temporal pole and external capsule in Swedish subjects upon magnetic resonance imaging. We further found no evidence for granular osmiophilic material in skin biopsy or post-mortem brain samples of affected members in the Swedish family. In addition, there was distinct lack of NOTCH3 N-terminal fragments in the cerebral microvasculature of the Swedish hereditary MID subjects compared to the intense accumulation in the English family afflicted with CADASIL. Several differences in arteriosclerotic changes in both the grey and white matter were also noted between the disorders. The sclerotic index values, density of collagen IV immunoreactivity in the microvasculature and number of perivascular macrophages were greater in the English CADASIL samples compared to those from the Swedish brains. Multiple approaches suggest that the Swedish family with hereditary MID suspected to be CADASIL has a different novel disorder with dissimilar pathological features and belongs to the growing number of genetically uncharacterized familial SVDs.

The impact of treatment for head and neck cancer on positive psychological change within a year of completing treatment.

Head and neck cancer carries a high level of morbidity and mortality. So why could anyone find having such a disease a positive event? The adversity hypothesis of "what doesn't kill you makes you stronger" suggests that people can use adversity to develop as human beings. This positive psychological change has received little attention in relation to head and neck cancer. Responses to the Silver Lining Questionnaire, University of Washington Quality of Life Questionnaire, and Short-Form 12 were collected from a postal survey, 3 to 12 months after the completion of treatment for head and neck cancer. Fifty-two (63%) people returned the survey and were included in the analysis. Time since completion of therapy did not show any relationship with positive psychological change. Tumour stage and treatment regimen both had a relationship with positive change. Participants with lower stage tumours had higher levels of positive change than those with tumours of higher stages. Participants who had surgery alone reported more positive change than those who had surgery with radiotherapy. A social factor related to greater change was being married or living with a partner when compared to living alone. Further research would aid the identification of bio-psychosocial factors that influence the development of positive psychological change and inform the development of rehabilitation interventions.

Documenting Resistance and Physiological Changes in Soybean Challenged by Aphis glycines Matsumura (Hemiptera: Aphididae).

The soybean aphid, Aphis glycines Matsumura (Hemiptera: Aphididae), is a limiting factor in soybean production in the North Central region of the USA. The objectives of this work were to identify sources of resistance to A. glycines in 14 soybean genotypes, and also document changes in total protein, peroxidase, and chlorophyll in response to aphid feeding. A reduced number of A. glycines was observed on the genotypes UX 2569-159-2-01 and UX 2570-171- 04, indicating the presence of antixenosis and/or antibiosis. UX 2569-159-2-01 expressed the highest level of resistance; whereas, UX 2570-171-04 had moderate levels of resistance to A. glycines. Chlorophyll content was relatively unaffected by A. glycines, except for a reduction in UX 2569-159-2-01 infested plants at 5 and 15 days after infestation (DAI). No changes were detected in total protein content between infested and control plants for the genotypes analyzed; however, peroxidase activity was higher in infested UX 2570-171-04 at both 5 and 10 DAI. This improvement in peroxidase content in infested UX 2570-171-04 may be playing multiple roles in the plant tolerance.

Gamma spectrometry in the ITWG CMX-4 exercise.

Low enriched uranium samples of unknown origin were analyzed by 16 laboratories in the context of a Collaborative Materials Exercise (CMX), organized by the Nuclear Forensics International Technical Working Group (ITWG). The purpose was to compare and prioritize nuclear forensic methods and techniques, and to evaluate attribution capabilities among participants. This paper gives a snapshot of the gamma spectrometric capabilities of the participating laboratories and summarizes the results achieved by gamma spectrometry.

Characterization and Identification of Methods for Phenotyping Soybean Populations With Tolerance to the Soybean Aphid (Hemiptera: Aphididae).

The development of soybeans tolerant to the soybean aphid [Aphis glycines Matsumura (Hemiptera: Aphididae)] remains unexplored. The objectives of this research were to determine the susceptibility of two high-yielding soybean [Glycine max (L.) Merrill (Fabales: Fabaceae)] genotypes involved in a breeding platform to develop aphid-tolerant recombinant inbred lines (RILs); characterize the peroxidase activity and relative expression of peroxidase transcripts in the parents of RILs; and identify an assay to phenotype aphid-tolerant RILs. Enzyme kinetic assays documented the total peroxidase activity for tolerant (KS4202), susceptible (SD76R), and two high-yielding (U09-105007 and U11-611112) soybeans during two vegetative stages (V1 and V3) at three sampling days (D4, D6, and D8 after aphid introduction). Enzyme kinetic assays showed that V3 infested tolerant and U11-611112 plants had significantly higher peroxidase activity than their respective control plants at D4, and infested tolerant plants were also higher than control plants at D6. There were no apparent trends when comparing the expression of peroxidase-specific transcripts in the absence of aphids (basal levels) in both V1 and V3. Relative expression analyses of two peroxidase transcripts (PRX52 and PRX2) performed to compare differences among the soybean genotypes indicated that, despite basal levels being similar for the treatments analyzed, tolerant soybeans had a tendency for a higher expression of PRX52 in the presence of aphids. Based on the different patterns observed and the feasibility of analyses performed in this study, enzyme kinetics using V3 infested plants may be a marker for screening RILs in a breeding program targeting the development of aphid-tolerant soybeans.

The Xenopus laevis ribosomal gene terminator contains sequences that both enhance and repress ribosomal transcription.

A DNA segment approximately 200 base pairs upstream of the Xenopus laevis ribosomal promoter acts both as an upstream promoter element that augments transcription and as a transcription terminator. It is, however, unclear to what extent these two activities are related. A segment of the X. laevis ribosomal DNA, containing the terminator and the upstream promoter element, was subjected to point mutation, and the effects of the resulting mutations were investigated by oocyte microinjection. Analysis of 26 point mutants revealed not only sequences that augment 40S transcription but also those that repress it. The sequences that augmented transcription lay within the T3 homology box and also near the site of 3'-end formation. These sequences also played a role in termination. The sequences that repressed transcription lay within the G+C-rich DNA flanking the T3 box. It can be concluded that termination is probably essential but may not be sufficient for the activity of the upstream promoter element.

Light-induced variability in development of forensically important blow fly Phormia regina (Diptera: Calliphoridae).

The use of the postmortem interval (PMI) in practical applications of forensic entomology is based upon developmental data of blow flies (Diptera: Calliphoridae) generated under controlled environmental conditions. Careful review of the published forensic entomology data sets showed that experimental (environmental) parameters differed between studies. Despite the differences in study design, there are no empirical data on the effect of photoperiod on blow fly development; yet, photoperiod has been shown to alter some insect development and behavior among noncalliphorids. Consequently, will differences in photoperiod alter the developmental times of calliphorids, and thereby alter PMI estimates? To answer this question, we used a replicated design with precise temperature measurement to examine the effects of photoperiod on the forensically important blow fly Phormia regina (Meigen). We concluded that inaccurate temperature recordings by using set-chamber temperatures over rearing-container temperatures would have overshadowed any affect light had on development. Second, constant light increased variation in overall adult developmental time and significantly delayed development compared with cyclic light. Finally, not accounting for delayed development induced by photoperiod underestimated the initial empirical estimate of the PMI. These sources of variation need to be included in forensic estimates because this variation can compromise predictions of PMI based upon current data sets. Without pinpointing optimal photoperiods with which to test development, we must assume that potentially large sources of variability exist within current estimates of the PMI.

Immunization information systems progress - United States, 2005.

Immunization registries are confidential, computerized information systems that collect and consolidate vaccination data from multiple health-care providers, generate reminder and recall notifications, and assess vaccination coverage within a defined geographic area. A registry with added capabilities, such as vaccine management, adverse event reporting, lifespan vaccination histories, and linkages with electronic data sources, is called an immunization information system (IIS). This report summarizes data from CDC's 2005 Immunization Information System Annual Report (IISAR), a survey of grantees in 50 states, five cities, and the District of Columbia (DC) that receive funding under section 317b of the Public Health Service Act. These data indicated that approximately 56% of U.S. children aged <6 years participated in an IIS, an increase from 48% in 2004. Moreover, 75% percent of public vaccination provider sites and 44% of private vaccination provider sites submitted vaccination data to an IIS during July-December 2005. These findings underscore the need to increase the number of participating children, from the current 13 million to approximately 21 million, to assure 95% participation of children aged <6 years and improve the effectiveness of U.S. immunization programs.

Effects of temperature on development of Phormia regina (Diptera: Calliphoridae) and use of developmental data in determining time intervals in forensic entomology.

Precise developmental data for forensic indicator blow fly species are essential for accuracy in the estimate of the post-mortem interval (PMI). Why, then, does the determination of the PMI result in conflicting time frames when published conspecific developmental data are used? To answer this question, we conducted constant temperature trials between the developmental minimum temperature and upper threshold temperatures (8-32 degrees C) on the forensically important blow fly species Phormia regina (Meigen) (Diptera: Calliphoridae). Flies were reared using two designs to quantify sources of variation. We measured rearing container temperatures and internal growth chamber temperatures by using thermocouples to accurately record temperatures experienced by larvae and to construct a degree-day model. Differences in experimental design, as seen across temperature studies for this fly species, did not significantly impact larval development. We also found that using set chamber temperatures rather than rearing container temperatures altered the final degree-day model. Using any minimum threshold (including an empirically determined true minimum) other than that from linear interpolation (x-intercept) violated degree-day assumptions and invalidated estimates of the PMI. We observed the minimum developmental temperature to be higher (14 degrees C) than that generated under the x-intercept method (5.46 degrees C) by using data from oviposition to adult emergence. This difference along with the noted difference in accumulated degree-days (using different base temperatures) suggests a need for additional experimentation on other forensically important fly species at low temperature thresholds to help with development of curvilinear models. Former and current estimates of the PMI may be inaccurate if the process to determine the time frame ignored degree-day model assumptions or was based upon questionable data sets.