The body size of the New Zealand orb-weaving spider Waitkera waitakerensis (Uloboridae) is directly related to temperature and affects fecundity

Abstract. Waitkera waitakerensis occupies lowland forests of New Zealand's North Island, where temperatures decrease in a southwestward direction. The mean annual temperatures of 18 collecting sites, as extracted from GIS data, are directly related to the first femur length of adult females. Neither site elevation nor phylogeny affected spider size or other variables examined. The direct relationship between spider body size and environmental temperature followed a pattern observed in other terrestrial arthropods with a univoltine life cycle and can probably be explained by the longer growing season of warmer regions. Egg diameter was uniform across the species. Site temperature and female first femur length were each directly related to the number of eggs deposited in egg sacs. The date of egg sac collection was inversely related to egg number, suggesting that clutch size declines during the reproductive season. Females deposit eggs beneath a triangular platform and then cover them with a lower silk sheet. The area of this upper platform and the volume of the egg sac were each directly related to egg number, but not to female first femur length. The depth of the lower covering was not related to egg number or to spider first femur length. This suggests that spiders use information about the volume of eggs in their abdomens to construct an egg sac whose volume will accommodate the volume of eggs to be laid and that females do so principally by adjusting the size of the sac's upper triangular platform.     

Dealing with uncertain absences in habitat modelling: a case study of a rare ground-dwelling parrot

In the development of a species distribution model based on regression techniques such as generalized linear or additive modelling (GLM/GAM), a basic assumption is that records of species presence and absence are real. However, a common concern in many studies examining species distributions is that absences cannot be inferred with certainty. This is particularly the case where the species is rare, difficult to detect and/or does not occupy all available habitat considered suitable. The western ground parrot ( Pezoporus wallicus flaviventris ) of southern Western Australia, Australia, is a case in point, as not only is it rare and difficult to detect, but it is also unlikely to occupy all available suitable habitat. A recent survey of ground parrots provided the opportunity to develop a predictive distribution model. As the data were susceptible to false absences, these were replaced with randomly selected 'pseudo' absences and modelled using GLM. As a comparison, presence-only information was modelled using a relatively new approach, MAXENT, a machine-learning technique that has been shown to perform comparatively well. The predictive performance of both models, as assessed by the receiver operating characteristic plot (ROC) was high (AUC > 0.8), with MAXENT performing only marginally better than the GLM. These approaches both indicated that the ground parrot prefers areas relatively high in altitude, distant from rivers, gently sloping to level habitat, with an intermediate cover of vegetation and where there is a mosaic of vegetation ages. In this case, the use of presence-only information resulted in the identification of important environmental attributes defining the occurrence of the ground parrot, but additional factors that account for the inability of the bird to occupy all suitable habitat should be a component of model refinement.     

Effect of intratumoral injection of carboplatin combined with pluronic P85 or L61 on experimental colorectal carcinoma in rats

Pluronic, a poly(ethylene oxide)-poly(propylene oxide)-poly (ethylene oxide) block copolymer, has been shown to enhance the cytotoxic activity of anticancer drugs in various cell lines. In the current study the effect of Pluronic P85 (P85) and Pluronic L61 (L61) on the intratumoral chemotherapy of an experimental adenocarcinoma in rats was examined. A total of 120 subcutaneous tumors (4 per rat) were inoculated in 30 BDIX rats and were treated weekly for 4 weeks with intratumoral injection of carboplatin (CPt) alone or with either P85 or L61. Tumors were monitored weekly and were excised at the endpoint for histologic evaluation. The effect of Pluronic on levels of intracellular ATP was explored as a possible mechanism of sensitization. Results showed that tumors treated with low-dose CPt (2.8 mg/kg) and P85 or L61 exhibited significant reductions in tumor volume after 28 days relative to Day 0 (112.7% +/- 34.4%, n = 15; 131.3% +/- 55.6%, n = 8) compared with tumors treated with free drug (339.4% +/- 75.0%, n = 16). Control tumors treated with either P85 or L61 alone or with saline showed volume increases of 1079.4% +/- 143.6% (n = 16), 729.4% +/- 202.2% (n = 7), and 1119.2% +/- 6.1% (n = 16), respectively. Treatment with high-dose CPt (20.7 mg/kg) led to a 79.3% +/- 4.2% reduction in tumor volume, and no differences were noted with addition of P85 or L61. In vitro ATP measurements showed that 28.0 mg/kg of P85 significantly reduced levels of intracellular ATP to 44.7% +/- 1.5% of controls, whereas L61 at this concentration depleted ATP levels completely. Results confirm that Pluronic P85 and L61 act as potent sensitizers to carboplatin chemotherapy of the experimental colorectal carcinoma, leading to a significant reduction of tumor growth compared to carboplatin alone. ATP depletion is a possible mechanism for these observed differences.     

Enhancing the Detection of Giardia duodenalis Cysts in Foods by Inertial Microfluidic Separation

The sensitivity and specificity of current Giardia cyst detection methods for foods are largely determined by the effectiveness of the elution, separation, and concentration methods used. The aim of these methods is to produce a final suspension with an adequate concentration of Giardia cysts for detection and a low concentration of interfering food debris. In the present study, a microfluidic device, which makes use of inertial separation, was designed and fabricated for the separation of Giardia cysts. A cyclical pumping platform and protocol was developed to concentrate 10-ml suspensions down to less than 1 ml. Tests involving Giardia duodenalis cysts and 1.90-μm microbeads in pure suspensions demonstrated the specificity of the microfluidic chip for cysts over smaller nonspecific particles. As the suspension cycled through the chip, a large number of beads were removed (70%) and the majority of the cysts were concentrated (82%). Subsequently, the microfluidic inertial separation chip was integrated into a method for the detection of G. duodenalis cysts from lettuce samples. The method greatly reduced the concentration of background debris in the final suspensions (10-fold reduction) in comparison to that obtained by a conventional method. The method also recovered an average of 68.4% of cysts from 25-g lettuce samples and had a limit of detection (LOD) of 38 cysts. While the recovery of cysts by inertial separation was slightly lower, and the LOD slightly higher, than with the conventional method, the sample analysis time was greatly reduced, as there were far fewer background food particles interfering with the detection of cysts by immunofluorescence microscopy.     

Model misspecification confounds the estimation of rates and exaggerates their time dependency

While welcoming the comment of Ho et al. ( 2015 ), we find little that undermines the strength of our criticism, and it would appear they have misunderstood our central argument. Here we respond with the purpose of reiterating that we are (i) generally critical of much of the evidence presented in support of the time‐dependent molecular rate (TDMR) hypothesis and (ii) specifically critical of estimates of μ derived from tip‐dated sequences that exaggerate the importance of purifying selection as an explanation for TDMR over extended timescales. In response to assertions put forward by Ho et al. ( 2015 ), we use panmictic coalescent simulations of temporal data to explore a fundamental assumption for tip‐dated tree shape and associated mutation rate estimates, and the appropriateness and utility of the date randomization test. The results reveal problems for the joint estimation of tree topology, effective population size and μ with tip‐dated sequences using beast . Given the simulations, beast consistently obtains incorrect topological tree structures that are consistent with the substantial overestimation of μ and underestimation of effective population size. Data generated from lower effective population sizes were less likely to fail the date randomization test yet still resulted in substantially upwardly biased estimates of rates, bringing previous estimates of μ from temporally sampled DNA sequences into question. We find that our general criticisms of both the hypothesis of time‐dependent molecular evolution and Bayesian methods to estimate μ from temporally sampled DNA sequences are further reinforced.     

Lipopolysaccharide transport to the cell surface: biosynthesis and extraction from the inner membrane

The cell surface of most Gram-negative bacteria is covered with lipopolysaccharide (LPS). The network of charges and sugars provided by the dense packing of LPS molecules in the outer leaflet of the outer membrane interferes with the entry of hydrophobic compounds into the cell, including many antibiotics. In addition, LPS can be recognized by the immune system and plays a crucial role in many interactions between bacteria and their animal hosts. LPS is synthesized in the inner membrane of Gram-negative bacteria, so it must be transported across their cell envelope to assemble at the cell surface. Over the past two decades, much of the research on LPS biogenesis has focused on the discovery and understanding of Lpt, a multi-protein complex that spans the cell envelope and functions to transport LPS from the inner membrane to the outer membrane. This paper focuses on the early steps of the transport of LPS by the Lpt machinery: the extraction of LPS from the inner membrane. The accompanying paper (May JM, Sherman DJ, Simpson BW, Ruiz N, Kahne D. 2015 Phil. Trans. R. Soc. B 370, 20150027. (doi:10.1098/rstb.2015.0027)) describes the subsequent steps as LPS travels through the periplasm and the outer membrane to its final destination at the cell surface.     

Lack of support for the time‐dependent molecular evolution hypothesis

There is increasing momentum surrounding the hypothesis that rates of molecular evolution between individuals within contemporary populations are high, and that these rates decrease as a function of time, perhaps over several millions of years, before reaching stationarity. The implications of this are powerful, potentially reshaping our view of how climate history impacts upon both species distribution patterns and the geographic structuring of genetic variation within species. However, our assessment of the hypothesis reveals a lack of theoretical support and empirical evidence for hypothesized magnitudes of time‐dependent rates of molecular evolution, with much of the apparent rate changes coming from artefacts and biases inherent in the methods of rate estimation. Our assessment also reveals a problem with how serial sampling is implemented for mutation rate estimation using ancient DNA samples, rendering published estimates unreliable.     

Use of a small molecule cell cycle inhibitor to control cell growth and improve specific productivity and product quality of recombinant proteins in CHO cell cultures

The continued need to improve therapeutic recombinant protein productivity has led to ongoing assessment of appropriate strategies in the biopharmaceutical industry to establish robust processes with optimized critical variables, that is, viable cell density (VCD) and specific productivity (product per cell, qP). Even though high VCD is a positive factor for titer, uncontrolled proliferation beyond a certain cell mass is also undesirable. To enable efficient process development to achieve consistent and predictable growth arrest while maintaining VCD, as well as improving qP, without negative impacts on product quality from clone to clone, we identified an approach that directly targets the cell cycle G1‐checkpoint by selectively inhibiting the function of cyclin dependent kinases (CDK) 4/6 with a small molecule compound. Results from studies on multiple recombinant Chinese hamster ovary (CHO) cell lines demonstrate that the selective inhibitor can mediate a complete and sustained G0/G1 arrest without impacting G2/M phase. Cell proliferation is consistently and rapidly controlled in all recombinant cell lines at one concentration of this inhibitor throughout the production processes with specific productivities increased up to 110 pg/cell/day. Additionally, the product quality attributes of the mAb, with regard to high molecular weight (HMW) and glycan profile, are not negatively impacted. In fact, high mannose is decreased after treatment, which is in contrast to other established growth control methods such as reducing culture temperature. Microarray analysis showed major differences in expression of regulatory genes of the glycosylation and cell cycle signaling pathways between these different growth control methods. Overall, our observations showed that cell cycle arrest by directly targeting CDK4/6 using selective inhibitor compound can be utilized consistently and rapidly to optimize process parameters, such as cell growth, qP, and glycosylation profile in recombinant antibody production cultures. Biotechnol. Bioeng. 2015;112: 141–155. © 2014 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc.