Posts tagged 'BIOTAQ'RSS Feed
"Regenerative medicine replaces or regenerates human cells, tissue or organs, to restore or establish normal function."- Mason, C. & Dunnill, P. (2008)
A brief definition of regenerative medicine. Regen. Med.(1), 1-5.
This week it was announced that Sir John B. Gurdon (Cambridge University) and Professor Shinya Yamanaka (Kyoto University and the Gladstone Institute in San Francisco) were joint recipients of the Nobel Prize in Physiology or Medicine 2012 for their pioneering work in the fields of cloning and stem cells.
Sir John B. Gurdon, joint 2012 Nobel Prize Winner (Photo: Creative Commons Attr. 2.0 Generic license)
Shinya Yamanaka, joint 2012 Nobel Prize Winner (Photo: Creative Commons Attr. 2.0 Generic license)
Their discoveries, made some 50 years apart, that mature specialised cells can be reprogrammed to become immature pluripotent cells capable of developing into any cell type are truly ground-breaking.
In a classic experiment, Gurdon replaced the immature cell nucleus in an egg cell of a frog (Xenopus laevis) with the nucleus from a mature intestinal cell. The modified egg still developed into a normal tadpole, overturning the dogma that cell differentiation was irreversible. Yamanaka, building on Gurdon’s work, discovered that mature cells can be reprogrammed to become pluripotent using a ‘reprogramming cocktail’ of just four genes (dubbed 'Yamanaka factors'). On an ethical level, Yamanaka's extension of this research has resulted in the ability to circumvent the need to derive stem cells from human embryos, a practice which is fraught with ethical implications and outlawed in a number of countries.
The Nobel Assembly said the scientists' findings had "revolutionised our understanding of how cells and organisms develop". Their work has led to a new field of regenerative medicine (RegenMed) using pluripotent stem cells, adult cells that have been reprogrammed to be able to develop into any cell type. It is hoped that Stem cell therapies may become effective in treating a variety of diseases by capitalising on the ability of transplanted cells to migrate toward target areas of injury and repair damaged tissues and organs.
Rapidly advancing developments — not only stem cell therapy, but also tissue engineering — hold the potential to revolutionise 21st century medicine. Aided by developments in nanomaterials and nanoparticles, tissue engineering has already seen patients successfully receive artificially synthesized veins, tracheas and bladders. Scientists are now attempting to build a variety of complete organs in the lab including kidneys, livers and hearts.
In this edition of Bioline Scholar, we focus on recently published papers in this exciting field. Bioline offers tried and trusted molecular biology tools to aid researchers working at the cutting-edge of RegenMed.
The demand for stem cells in drug discovery and clinical applications is growing quickly. However, to translate the promise of stem cells into therapeutic reality for patient populations, industrial-scale production under tightly regulated conditions is required. Researchers from the UCL Stem Cell and Regenerative Medicine Bioprocessing Unit, led by Prof Chris Mason*, tested a commercial ‘hollow fibre’ bioreactor for scale-up of human embryonic stem cell (hESC) derived cell therapies. The bioreactor was able to successfully expand hESCs over ten-fold (from 60-708 million), whilst maintaining high levels of pluripotency marker expression. 97.7 % of cells expressed the embryonic stem cell marker, SSEA-4 when harvested.
* Chris is a co-founder of the London Regenerative Medicine Network (LRMN) and an internationally renowned RegenMed expert.
Roberts, I. et al. (2012) - Scale-up of Human Embryonic Stem Cell Culture using a Hollow Fibre Bioreactor. Biotechnol Lett. doi: 10.1007/s10529-012-1033
The growth and differentiation of stem cells is controlled by intrinsic and extrinsic bioactive factors. The latter includes growth factors, which are typically included in culture media.• This paper from the Universities of Edinburgh, Politecnica delle Marche and Yale describes the use of affinity targeted biodegradable nanoparticles to mediate paracrine stimulation as an alternative approach to sustain the growth and pluripotency of mouse (ES) cells. The dynamic microenvironmental control of extrinsic factors will be of benefit for stem cell manufacturing.
Corradetti, B. et al. (2012) - Paracrine signalling events in embryonic stem cell renewal mediated by affinity targeted nanoparticles Biomaterials 33(28), 6634-6643
Human mesenchymal stromal/stem cells (MSC) isolated from fetal tissues are ethically contentious as a cell source and also technically challenging to obtain. In contrast, stem cells are more readily-obtainable from the placenta throughout pregnancy.
In this study, from the Fetal Stem Cell Therapy Group, Imperial College, the phenotype of first trimester and term fetal placental chorionic stem cells (e-CSC and l-CSC respectively) were compared to determine their suitability for cell-based therapies and tissue engineering applications. e-CSC showed an earlier state of ‘stemness’ compared to l-CSC, higher tissue repair in vivo (osteogenesis imperfecta murine model), increased bone quality and plasticity and accelerated skin healing. Their results suggest the more primitive characteristics of early compared to late gestation fetal chorionic stem cells may be translationally advantageous, and provide insight into the developmental ontogeny of fetal chorionic stem cells.
Jones G.N. et al. (2012) - Ontological Differences in First Compared to Third Trimester Human Fetal Placental Chorionic Stem Cells. PLoS ONE 7(9): e43395.
Further BioTaq DNA Polymerase references
Fujishiro, S.H. et al. (2012). Generation of Naive-like Porcine-Induced Pluripotent Stem Cells Capable of Contributing to Embryonic and Fetal Development. Stem Cells Dev. doi:10.1089/scd.2012.0173
Martínez-González, I. et al. (2012). Engraftment Potential of Adipose Tissue-Derived Human Mesenchymal Stem Cells After Transplantation in the Fetal Rabbit. Stem Cells Dev. doi:10.1089/scd.2012.0032
Jones, G.N. et al. (2012). Upregulating CXCR4 in Human Fetal Mesenchymal Stem Cells Enhances Engraftment and Bone Mechanics in a Mouse Model of Osteogenesis Imperfecta. Stem Cells Trans Med (1)1: 70-78
Progress in the use of hESCs derivatives for cellular therapies also necessitates the production of clinical-grade lines under the control of good manufacturing practice (GMP). Human fibroblast feeder cells are currently required for derivation of new hESC lines. This work from the North East England Stem Cell Institute in Newcastle is the first report of the production of a GMP-grade human fibroblast line (NclFed1A) derived from human foreskin with consent for derivation and culture of hESCs.
Prathalingam, N. et al. (2012) - Production and Validation of a Good Manufacturing Practice Grade Human Fibroblast Line for Supporting Human Embryonic Stem Cell Derivation and Culture. Stem Cell Res Ther 3:12
Prof. Molly Stevens* runs a large multidisciplinary group with interests in the directed differentiation of stem cells, design of novel bioactive scaffolds and new approaches towards tissue regeneration. Poly(γ-glutamic acid) (γ-PGA), a biocompatible natural polymer was investigated as a new and more versatile candidate for tissue engineering applications. Fibrous scaffolds were engineered from ethyl, propyl and benzyl esterifications of γ-PGA. γ-PGA-Benzyl showed increased cell adhesion of human mesenchymal stem cells (hMSCs), three-fold higher viability at day 14 compared to γ-PGA-ethyl and γ-PGA-propyl, and significantly higher adhesion compared with PLLA polyester scaffolds. γ-PGA-Bn scaffolds cultured in osteogenic media were also able to support differentiation of hMSCs towards an osteogenic lineage as shown by alkaline phosphatase and Runx2 gene expression.
* Molly is Professor of Biomedical Materials and Regenerative Medicine and the Research Director for Biomedical Material Sciences in the Institute of Biomedical Engineering at Imperial College. She is featured in a recent article in The Lancet and also as one of British Vogue Magazine’s Wonder Women.
Gentilini, C., et al. (2012) - Functionalized Poly(γ-Glutamic Acid) Fibrous Scaffolds for Tissue Engineering. Adv. Health. Mat. 1: 308-315. doi: 10.1002/adhm.201200036
Canonical Wnt/β-catenin signaling has been suggested to promote self-renewal of pluripotent mouse and human ES cells. In this paper from California State University, SB-216763, a GSK3 inhibitor, could maintain mouse ES cells in a pluripotent state without exogenous leukemia inhibitory factor (LIF) when cultured on mouse embryonic fibroblasts.
Kirby, L.A. et al. (2012) - Glycogen Synthase Kinase 3 (GSK3) Inhibitor, SB-216763, Promotes Pluripotency in Mouse Embryonic Stem Cells. PLoS ONE 7(6): e39329.
Further SensiMix references
Lima, M.J. et al. (2012) - Pancreatic Transcription Factors Containing Protein Transduction Domains Drive Mouse Embryonic Stem Cells towards Endocrine Pancreas. PLoS ONE 7(5): e36481.
Fouraschen, S.M., et al. (2012) - Secreted Factors of Human Liver-Derived Mesenchymal Stem Cells Promote Liver Regeneration Early After Partial Hepatectomy. PLoS ONE 7(5): e36481.
The transplantation of neural crest derived stem cells (NCSCs) is a potent alternative for the treatment of Hirschsprung’s disease (HSCR). Cells to be transplanted require an appropriate microenvironment to survive and differentiate. Researchers from Heidelberg University investigated the quality of this microenvironment, by studying the effects of HSCR-smooth-muscle-protein extracts upon NCSCs in vitro.
Hagl, C.L. et al. (2012) - Smooth muscle proteins from Hirschsprung's disease facilitates stem cell differentiation. Pediatr Surg Int. 28(2):135-142.
Lu, Z. et al. (2012) - Short-Term Exposure to Tumor Necrosis Factor-Alpha Enables Human Osteoblasts to Direct Adipose Tissue-Derived Mesenchymal Stem Cells into Osteogenic Differentiation. Stem Cells Dev. 21(13): 2420-2429.
Lu, Z. et al. (2012) - Bone Biomimetic Microenvironment Induces Osteogenic Differentiation of Adipose Tissue-derived Mesenchymal Stem Cells. Nanomedicine: Nanotech. Biol. Med. 8(4):507-515.
Autism spectrum disorders (ASDs), or pervasive developmental disorders (PDDs), are a category of complex developmental brain disorders that appear in early childhood, usually before the age of three. ASDs cause difficulties in social, communication and behavioural skills, but affected individuals can manifest a wide range of symptoms, from very mild to severe - hence the term "spectrum".
One in 110 children in the US has an ASD, with four times as many boys than girls being affected. In the last decade, diagnoses have increased tenfold.
ASDs are currently defined as five disorders:
- Autistic disorder (also called “classic autism”)
- Asperger syndrome
- Pervasive developmental disorder not otherwise specified (PDD-NOS)
- Rett disorder
- Childhood disintegrative disorder (CDD).
Copy number variants (CNVs) – a form of structural variation - are deletions or duplications of genomic segments ranging from several thousand to a few million base pairs. A number of population-based studies have demonstrated that CNVs can affect as much as 12% of the human genome. Large scale studies have gone on to show the importance of CNVs in determining human phenotypic variation and disease susceptibility.
There is strong evidence that rare CNVs play a role in susceptibility to ASDs. A large study published in Nature by the Autism Genome Project (AGP), an international consortium of scientists from more than 60 institutions in 12 countries, compared the DNA of almost 1000 children with ASDs and healthy children, using 1 million single-nucleotide polymorphism (SNP) arrays. The study showed that rare CNVs, possibly acting in tandem, play a significant role in the genetic aetiology of this condition. The rare variants were identified in less than 1% of the normal population but strikingly, occurred almost 20% more frequently in ASD children.
Further recent research has focused on how CNVs mediate a phenotypic effect by altering gene expression levels, which is the focus of the study by Luo and colleagues published in the June 2012 edition of the American Journal of Human Genetics. A second paper from Holt and colleagues published in the May 2012 edition of the European Journal of Human Genetics investigates an alternative mechanism whereby CNVs combine the 5' and 3' ends of two genes, creating a novel 'fusion gene'.
This pair of papers sheds light on some of the genomic imbalances that could contribute to ASD pathogenesis. Bioline offers products ideal for use in genome-wide studies of structural variants and disease susceptibility.
Luo and colleagues used genome-wide transcriptome profiling to evaluate the functional consequences of rare structural CNVs in ASD. The UCLA team along with collaborators from Yale, Wellcome Trust Sanger Institute, Carnegie Mellon, and University of Pittsburgh identified a number of interesting candidate ASD loci at 12p11.22, 15q23, 1p34.3, 3q27, and 3p26.2. For example, the 3p26.2 loci, found deleted, harbours three genes: ITPR1, SETMAR and SUMF1, all of which are down-regulated. Although none of these genes has been previously associated with autism, they are all functionally linked to the nervous system.
This study provides evidence that pathogenic structural variants have a functional impact via transcriptome alterations in ASDs at a genome-wide level. The authors also demonstrate the usefulness of integrating gene expression and mutation data to prioritize candidate genes disrupted by potentially deleterious alterations.
Luo R., et al. Am. J. Hum. Gen. 91(1): 38–55 (2012) – Genome-wide Transcriptome Profiling Reveals the Functional Impact of Rare De Novo and Recurrent CNVs in Autism Spectrum Disorders
Fusion-proteins play an important role in cancer genetics, as exemplified by BCR-ABL in leukemia and TMPRSS2-ERG in prostate cancers. A team from the Wellcome Trust Centre for Human Genetics, University of Oxford addressed the question of how common CNV-induced fusion transcripts are and whether they may play a role in ASD susceptibility. Firstly, using the published Illumina 1 million SNP array data from the AGP consortium, they performed bioinformatic analysis of existing CNV calls. They then validated CNVs using qPCR.
No evidence was found that fusion-gene generating CNVs led to ASD susceptibility. However, the discovery of a MAPKAPK5-ACAD10 transcript with an estimated frequency of ~1/200 suggests that gain-of-function mechanisms should be considered in future studies of genomic imbalance and disease susceptibility.
Holt, R., et al. Eur. J. Hum. Gen. doi: 10.1038/ejhg.2012.73 (2012) - CNVs Leading to Fusion Transcripts in Individuals with Autism Spectrum Disorder
Focus on Breast Cancer
For years scientists have been treating breast cancer as a single disease. However, a new landmark study published in Nature has reclassified breast cancer into ten separate sub-diseases based on their genetic fingerprint. The culmination of decades of research, the study is the largest global study of breast cancer tissue ever performed.
The team, led by the British Columbia Cancer Center in Canada and the Cambridge Cancer Research Institute in the UK, used genome-wide microarrays to analyze the DNA and RNA of 2,000 tumor samples taken from women diagnosed with breast cancer. This huge pool of genetic information (copy number variants, SNPs and gene expression data), as well as survival data, allowed researchers to spot new and previously unacknowledged patterns for ten subtly different cancers that have, historically, been considered as one.
The challenge now is to understand the genetic drivers behind these newly discovered breast cancer variants and to develop new targeted therapies in the future. It could also lead to women with the best prognosis being spared side-effects of chemotherapy. The classification system will likely also form the basis for newer and better ways to diagnose and manage the disease.
Bioline offers a number of reagents that have helped further the study of cancers and, more specifically, breast cancer. So this edition of Bioline Scholar Monthly focuses on the use of Bioline reagents and kits in the field of breast cancer research.
In a diverse cohort of breast cancer patients with a 1–5 year tumor relapse versus those with up to 7 years relapse-free survival, RNA was extracted and subjected to microarray and real-time RT-PCR analysis. Among the 299 genes, five genes which included B cell response genes were found to predict with >85% accuracy relapse-free survival. Real-time RT-PCR confirmed the 5-gene prognostic signature that was distinct from an FDA-cleared 70-gene signature of MammaPrint panel and from the Oncotype DX recurrence score assay panel.
Ascierto, L. M., et al. Breast Can. Res. Treat. 131(3):871-880 (2012) - A signature of immune function genes associated with recurrence-free survival in breast cancer patients.
MicroRNAs (miRNAs) are noncoding RNAs that function as key posttranscriptional regulators of gene expression. This paper found that BRCA1 recognizes the RNA secondary structure and directly binds with primary transcripts of miRNAs via a DNA-binding domain. The findings indicate novel functions of BRCA1 in miRNA biogenesis, which may be linked to its tumor suppressor mechanism and maintenance of genomic stability.
Kawai S. and Amano A. J. Cell Biol. 197 (2):201-208 (2012) - BRCA1 regulates microRNA biogenesis via the DROSHA microprocessor complex.
The bioactive lipid sphingosine 1-phosphate (S1P) uses sphingosine 1-phosphate receptor 4 (S1P4) and human epidermal growth factor receptor 2 (HER2) to stimulate the extracellular signal regulated protein kinase 1/2 (ERK-1/2) pathway in MDA-MB-453 cells. The magnitude of the signaling gain on the ERK-1/2 pathway produced in response to S1P can be increased by HER2 in MDA-MB-453 cells. The linkage of S1P with an oncogene suggests that S1P and specifically S1P4 may have an important role in breast cancer progression.
Long J. S., et al. J. Biol. Chem. 285:35957-35966 (2010) - Sphingosine 1-Phosphate Receptor 4 Uses HER2 (ERBB2) to Regulate Extracellular Signal Regulated Kinase-1/2 in MDA-MB-453 Breast Cancer Cells.
CD44, the transmembrane receptor for hyaluronan, is implicated in tumor cell invasion and metastasis. The expression of CD44 and its variants is associated with poor prognosis in breast cancer. This paper investigated the effect of silibinin (a polyphenolic flavonolignan of the herbal plant of Silybum marianum, milk thistle) on the epidermal growth factor (EGF) ligand-induced CD44 expression in human breast cancer cells. The results suggest that silibinin prevents the EGFR signaling pathway and may be used as an effective drug for the inhibition of metastasis of human breast cancer.
Kim S., et al. Anticancer Res. 31(11): 3767-3773 (2011) - Silibinin Suppresses EGFR Ligand-induced CD44 Expression through Inhibition of EGFR Activity in Breast Cancer Cells.
Human papillomavirus (HPV) and Epstein Barr virus (EBV) have been found in breast carcinomas around the world. In this study, fifty-five BCs from Chile were analyzed for HPV and EBV presence. In addition, HPV- 16 viral load/physical status and E6/E7 expressions were determined. The results suggest that it is unlikely that HPV and/or EBV play a direct role in the etiology of breast carcinomas.
Aguayo F., et al. Infectious Agents and Cancer 6:7 (2011) - Human papillomavirus and Epstein-Barr virus infections in breast cancer from chile.
This study suggests that melatonin may play a role in the desmoplastic reaction in breast cancer through a down regulatory action on the expression of antiadipogenic cytokines, which decrease the levels of these cytokines. Lower levels of cytokines stimulate the differentiation of fibroblasts and decrease both aromatase activity and expression, thereby reducing the number of estrogen-producing cells proximal to malignant cells.
Alonso-González C., et al. J. Pineal Res. 52(3): 282–290, (2012) - Melatonin interferes in the desmoplastic reaction in breast cancer by regulating cytokine production.
Melatonin reduces the development of breast cancer interfering with oestrogen-signalling pathways, and also inhibits aromatase activity and expression. This study shows that melatonin inhibits aromatase activity and expression by regulating the gene expression of specific aromatase promoter regions. A possible mechanism for these effects would be the regulation by melatonin of intracellular cAMP levels, mediated by an inhibition of cyclooxygenase activity and expression.
Martínez-Campa C., et al. British J. Can. 101: 1613–1619 (2009) - Melatonin inhibits aromatase promoter expression by regulating cyclooxygenases expression and activity in breast cancer cells.
Bisphenol A (BPA) has long been suspected to promote carcinogenesis, but the high doses of BPA used in many studies generated conflicting results. This paper shows that BPA at environmentally relevant doses reduces the efficacy of chemotherapeutic agents. These data provide considerable support to the accumulating evidence that BPA is hazardous to human health.
LaPensee E. W., et al. Environ Health Perspect. 117(2): 175–180 (2009) - Bisphenol A at Low Nanomolar Doses Confers Chemoresistance in Estrogen Receptor-a–Positive and –Negative Breast Cancer Cells.
This paper indicates that decreased CDH1, CDH13 and TIMP3 with increased CD44 gene expression levels can be used as an indicator for invasion in both ER-positive and ER-negative breast tumors. In double-negative tumor tissues, CD44 can be considered a marker for aggressive properties. However, additional assays in a larger series of patients with long follow up will be necessary to confirm these results of gene expressions in ER-positive and ER-negative tumors and their relationship with HER2 and ESR1.
Celebiler A., et al. Can. Sci. 100: 2341–2345. (2009) - Predicting invasive phenotype with CDH1, CDH13, CD44, and TIMP3 gene expression in primary breast cancer.
It's time for the second in our series of Bioline Scholar Monthly compilations. This month we're focusing on the many uses of Bioline polymerases in forensic science.
Identification of a report's species is one of the basic analyses in forensics. However, due to the nature of the sampling environment, DNA samples often contain PCR-inhibitory substances which may generate blank or incomplete DNA profiles. The common approach to overcoming PCR inhibition is extensive DNA purification, but this can increase the risk of DNA loss. In some cases, isolation of single cells using laser-capture microdissection can be used, but again this reduces the amount of DNA available.
DNA polymerases that can improve the quality of forensic DNA analysis and efficiently circumvent PCR inhibition, without any additional sample preparation, are therefore advantageous, as are polymerases that result in high yields.
Bioline’s DNA polymerases are very robust and have been carefully designed to overcome these problems. BIO-X-ACT™ Short in particular is specifically designed for difficult/problematic PCR applications that require high processivity and fidelity, applications that would normally fail with other DNA polymerase. MangoTaq™ has also been designed for problematic and ancient DNA, whereas IMMOLASE™ and BIOTAQ™ are high yield for small sample sizes. Together with our new MyTaq™, MyFi™ and RANGER Bioline has polymerases to meet all the high-fidelity requirements of forensic science.
So, without further ado, here's...
Bioline Scholar Monthly: February 2012
Loop-mediated isothermal amplification (LAMP) is an uncomplicated, quick and relatively inexpensive diagnostic tool. In Barkway and colleagues procedure, BIO-X-ACT Short DNA Polymerase was initially used to successfully verify the LAMP primer pair for Eimeria species specificity using PCR.
Barkway, C. P., et al. BMC Veterinary Research 2011, 7:67 (2011) – Loop-mediated isothermal amplification (LAMP) assays for the species-specific detection of Eimeria that infect chickens
In a comparison of nine DNA polymerases, the DNA detection limit was lowest with BIO-X-ACT Short, providing the highest number of improved DNA profiles, using real crime scene saliva samples.
Hedman, J., et al. BioTechniques 47, 951-958 (2009) – Improved forensic DNA analysis through the use of alternative DNA polymerases and statistical modeling of DNA profiles
The identification via DNA analysis is reliably and reproducibly possible from well preserved and semi-burnt bones.
Schwark, T., et al Forensic Sci. Int.: Gene. 5(5), 393-399 (2011) – Reliable genetic identification of burnt human remains.
Dried herbarium specimens may be invaluable to understand long-term changes at sites with a history of cyanobacterial blooms.
Metcalf, J.S., et al Harmful Algae 15 47–52 - (2012) – Analysis of microcystins and microcystin genes in 60–170-year-old dried herbarium specimens of cyanobacteria
Out of 19 polymerases, the best performance was exhibited by the Mango-Taq DNA polymerase, which was the only polymerase which was able to amplify the ~620 bp amplification product from the 102 year old sample.
Telle, S. & Marco Thines, M. PLoS ONE 3(10):doi:10.1371/journal.pone.0003584 - (2008) – Amplification of cox2 (~620 bp) from 2 mg of Up to 129 Years Old Herbarium Specimens, Comparing 19 Extraction Methods and 15 Polymerases
In Australia and globally, Sarcophagidae flies remain unexploited as indicators of post-mortem interval in forensic investigations. A molecular identification method involving DNA ‘barcoding’ of the mitochondrial COI gene from 16 species of Australian Sarcophagidae was successfully developed. The authors conclude analysis of sarcophagids in forensic entomology should increase and their value as tools in criminal investigations realised.
Meiklejohn, K. A., et al. Int. J. Legal Med. 125(1), 27-32 - (2011) – DNA-based identification of forensically important Australian Sarcophagidae (Diptera)
Microsatellite markers were developed for the medicinal plant Tripterygium (Celastraceae) to assess its population structure and to facilitate source tracking of plant materials used for medicinal extracts.
Novy, A. & Jones, K. C. Am. J. Bot. 98(10) e280-e281 (2011) – Characterization of polymorphic microsatellites for Tripterygium (Celastraceae), a monospecific genus of medicinal importance
Lindgren funnel traps baited with aggregation pheromones are effective tools for monitoring flight activity in the red flour beetle (Tribolium castaneum) and lesser grain borer (Rhyzopertha dominica). But the samples are affected by the preservation method.
Stevens, M. M,. et al. J. Stored Products Res. 47(2), 69–75 - (2011) – Maintaining DNA quality in stored-grain beetles caught in Lindgren funnel traps