Prospecting for scarabid specific Bacillus thuringiensis crystal toxin cry8 gene in sugarcane ecosystem of Tamil Nadu, India

In the present study, we report the occurrence of cry8 positive isolates of Bacillus thuringiensis (Bt) in selected white grub, Holotrichia serrata F. (Coleoptera: Scarabaeidae), endemic soils of sugarcane ecosystem and other places in Tamil Nadu. Out of the 66 soil samples collected and screened for white grub specific Bt, 74 isolates of the bacterium, all containing only spherical crystal toxin, were identified. PCR screening of these isolates with cry8 gene universal primer revealed six isolates to be positive. Further, the amplicon of a 370 bp band, amplified with another set of degenerate primer designed based on the conserved sequence of cry8 genes, was sequenced from four isolates. Multiple sequence alignment revealed the gene sequences to be the same for all the isolates. The present report of the availability of cry8 positive Bt isolates opens the avenue for controlling white grubs through transgenic research

Temperature-dependent development of Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) and its larval parasitoid, Habrobracon hebetor (Say) (Hymenoptera: Braconidae): implications for species interactions

Habrobracon hebetor (Say) is a parasitoid of various Lepidoptera including Helicoverpa armigera (Hübner), a key pest of different crops and vegetables. The development of both H. armigera and H. hebetor were simultaneously evaluated against a wide range of constant temperatures (10, 15, 17.5, 20, 25, 27.5, 30, 35, 37.5 and 40 °C). Helicoverpa armigera completed its development from egg to adult within a temperature range of 17.5–37.5 °C and H. hebetor completed its life cycle from egg to adult within a temperature range of 15–40 °C. Based on the Ikemoto and Takai model the developmental threshold (T o) and thermal constant (K) to complete the immature stages, of H. armigera were calculated as 11.6 °C and 513.6 DD, respectively, and 13 °C and 148 DD, respectively, for H. hebetor. Analytis/Briere-2 and Analytis/Briere-1 were adjudged the best non-linear models for prediction of phenology of H. armigera and H. hebetor, respectively and enabled estimation of the optimum (T opt) and maximum temperature (T max) for development with values of 34.8, 38.7, 36.3, and 43 °C for host and the parasitoid, respectively. Parasitisation by H. hebetor was maximal at 25 °C but occurred even at 40 °C. This study suggests although high temperature is limiting to insects, our estimates of the upper thermal limits for both species are higher than previously estimated. Some biological control of H. armigera by H. hebetor may persist in tropical areas, even with increasing temperatures due to climate change

From microscopic to macroscopic descriptions of cell\ud migration on growing domains

Cell migration and growth are essential components of the development of multicellular organisms. The role of various cues in directing cell migration is widespread, in particular, the role of signals in the environment in the control of cell motility and directional guidance. In many cases, especially in developmental biology, growth of the domain also plays a large role in the distribution of cells and, in some cases, cell or signal distribution may actually drive domain growth. There is a ubiquitous use of partial differential equations (PDEs) for modelling the time evolution of cellular density and environmental cues. In the last twenty years, a lot of attention has been devoted to connecting macroscopic PDEs with more detailed microscopic models of cellular motility, including models of directional sensing and signal transduction pathways. However, domain growth is largely omitted in the literature. In this paper, individual-based models describing cell movement and domain growth are studied, and correspondence with a macroscopic-level PDE describing the evolution of cell density is demonstrated. The individual-based models are formulated in terms of random walkers on a lattice. Domain growth provides an extra mathematical challenge by making the lattice size variable over time. A reaction-diffusion master equation formalism is generalised to the case of growing lattices and used in the derivation of the macroscopic PDEs

Genome-Wide Tissue-Specific Occupancy of the Hox Protein Ultrabithorax and Hox Cofactor Homothorax in Drosophila

The Hox genes are responsible for generating morphological diversity along the anterior-posterior axis during animal development. The Drosophila Hox gene Ultrabithorax (Ubx), for example, is required for specifying the identity of the third thoracic (T3) segment of the adult, which includes the dorsal haltere, an appendage required for flight, and the ventral T3 leg. Ubx mutants show homeotic transformations of the T3 leg towards the identity of the T2 leg and the haltere towards the wing. All Hox genes, including Ubx, encode homeodomain containing transcription factors, raising the question of what target genes Ubx regulates to generate these adult structures. To address this question, we carried out whole genome ChIP-chip studies to identify all of the Ubx bound regions in the haltere and T3 leg imaginal discs, which are the precursors to these adult structures. In addition, we used ChIP-chip to identify the sites bound by the Hox cofactor, Homothorax (Hth). In contrast to previous ChIP-chip studies carried out in Drosophila embryos, these binding studies reveal that there is a remarkable amount of tissue- and transcription factor-specific binding. Analyses of the putative target genes bound and regulated by these factors suggest that Ubx regulates many downstream transcription factors and developmental pathways in the haltere and T3 leg. Finally, we discovered additional DNA sequence motifs that in some cases are specific for individual data sets, arguing that Ubx and/or Hth work together with many regionally expressed transcription factors to execute their functions. Together, these data provide the first whole-genome analysis of the binding sites and target genes regulated by Ubx to specify the morphologies of the adult T3 segment of the fly

Whole genome analysis and functional characterization of a novel Bacillus thuringiensis (Bt 62) isolate against sugarcane white grub Holotrichia serrata (F)

In this study, we report the whole genome assembly of Bt 62, a novel isolate harbouring cry8 holotype gene identified by us earlier. Sequencing was carried out using a combination of Illumina NextSeq 500 and Oxford Nanopore sequencing Technologies (ONT). The final assembled genome was 6.13 Mb comprising a circular chromosome and four plasmids. The bioassay studies against Holotrichia serrata (F.) (Coleoptera: Scarabaeidae), a polyphagous pest infesting sugarcane and other crops, indicated significant toxicity to first instar grubs over untreated larvae achieving a highest mean mortality of 91.11% for various doses tested. In vitro proteolytic assay and histopathological studies of the midgut of infected white grubs revealed proteolytic processing of the protoxin and extensive degeneration of larval midgut epithelial cells. The results demonstrate that this novel isolate could be used as a biopesticide or its crystal toxin genes could be expressed in sugarcane and other crops for resistance against H. serrata

Development and validation of real-time PCR screening methods for detection of cry1A.105 and cry2Ab2 genes in genetically modified organisms

Primers and probes were developed for the element-specific detection of cry1A.105 and cry2Ab2 genes, based on their DNA sequence as present in GM maize MON89034. Cry genes are present in many genetically modified (GM) plants and they are important targets for developing GMO element-specific detection methods. Element-specific methods can be of use to screen for the presence of GMOs in food and feed supply chains. Moreover, a combination of GMO elements may indicate the potential presence of unapproved GMOs (UGMs). Primer-probe combinations were evaluated in terms of specificity, efficiency and limit of detection. Except for specificity, the complete experiment was performed in 9 PCR runs, on 9 different days and by testing 8 DNA concentrations. The results showed a high specificity and efficiency for cry1A.105 and cry2Ab2 detection. The limit of detection was between 0.05 and 0.01 ng DNA per PCR reaction for both assays. These data confirm the applicability of these new primer-probe combinations for element detection that can contribute to the screening for GM and UGM crops in food and feed samples

Caracterização do gene vip3A e toxicidade da proteína Vip3Aa50 à lagarta-do-cartucho e à lagarta-da-soja

O objetivo deste trabalho foi caracterizar o gene vip3A de Bacillus thuringiensis e verificar a toxicidade da proteína Vip3Aa50 a larvas da lagarta-do-cartucho (Spodoptera frugiperda) e da lagarta-da-soja (Anticarsia gemmatalis). O gene vip3A foi amplificado por PCR, com iniciadores específicos, e gerou um fragmento de 2.370 pb. Esse fragmento foi clonado em vetor pGEM-T Easy e, em seguida, sequenciado, subclonado em vetor de expressão pET-28a (+) e inserido em células de Escherichia coli BL21 (DE3). A expressão da proteína Vip3Aa50 foi induzida por isopropil-β-D-1-tiogalactopiranosídeo (IPTG), visualizada em SDS-PAGE e detectada por "Western blot". Os ensaios de toxicidade revelaram alta atividade da proteína Vip3Aa50 contra as larvas neonatas da lagarta-da-soja e da lagarta-do-cartucho, com CL50 de 20,3 e 79,6 ng cm-2, respectivamente. O gene vip3Aa50 é um novo gene da classe vip3A