A genetic linkage map of an apple rootstock progeny anchored to the Malus genome sequence

An apple rootstock progeny raised from the cross between the very dwarfing ‘M.27’ and the more vigorous ‘M.116’ (‘M.M.106’ × ‘M.27’) was used for the construction of a linkage map comprising a total of 324 loci: 252 previously mapped SSRs, 71 newly characterised or previously unmapped SSR loci (including 36 amplified by 33 out of the 35 novel markers reported here), and the self-incompatibility locus. The map spanned the 17 linkage groups (LG) expected for apple covering a genetic distance of 1,229.5 cM, an estimated 91% of the Malus genome. Linkage groups were well populated and, although marker density ranged from 2.3 to 6.2 cM/SSR, just 15 gaps of more than 15 cM were observed. Moreover, only 17.5% of markers displayed segregation distortion and, unsurprisingly in a semi-compatible backcross, distortion was particularly pronounced surrounding the self-incompatibility locus (S) at the bottom of LG17. DNA sequences of 273 SSR markers and the S locus, representing a total of 314 loci in this investigation, were used to anchor to the ‘Golden Delicious’ genome sequence. More than 260 of these loci were located on the expected pseudo-chromosome on the ‘Golden Delicious’ genome or on its homeologous pseudo-chromosome. In total, 282.4 Mbp of sequence from 142 genome sequence scaffolds of the Malus genome were anchored to the ‘M.27’ × ‘M.116’ map, providing an interface between the marker data and the underlying genome sequence. This will be exploited for the identification of genes responsible for traits of agronomic importance such as dwarfing and water use efficiency.

Development of a dense SNP-based linkage map of an apple rootstock progeny using the Malus Infinium whole genome genotyping array

Background A whole-genome genotyping array has previously been developed for Malus using SNP data from 28 Malus genotypes. This array offers the prospect of high throughput genotyping and linkage map development for any given Malus progeny. To test the applicability of the array for mapping in diverse Malus genotypes, we applied the array to the construction of a SNPbased linkage map of an apple rootstock progeny. Results Of the 7,867 Malus SNP markers on the array, 1,823 (23.2 %) were heterozygous in one of the two parents of the progeny, 1,007 (12.8 %) were heterozygous in both parental genotypes, whilst just 2.8 % of the 921 Pyrus SNPs were heterozygous. A linkage map spanning 1,282.2 cM was produced comprising 2,272 SNP markers, 306 SSR markers and the S-locus. The length of the M432 linkage map was increased by 52.7 cM with the addition of the SNP markers, whilst marker density increased from 3.8 cM/marker to 0.5 cM/marker. Just three regions in excess of 10 cM remain where no markers were mapped. We compared the positions of the mapped SNP markers on the M432 map with their predicted positions on the ‘Golden Delicious’ genome sequence. A total of 311 markers (13.7 % of all mapped markers) mapped to positions that conflicted with their predicted positions on the ‘Golden Delicious’ pseudo-chromosomes, indicating the presence of paralogous genomic regions or misassignments of genome sequence contigs during the assembly and anchoring of the genome sequence. Conclusions We incorporated data for the 2,272 SNP markers onto the map of the M432 progeny and have presented the most complete and saturated map of the full 17 linkage groups of M. pumila to date. The data were generated rapidly in a high-throughput semi-automated pipeline, permitting significant savings in time and cost over linkage map construction using microsatellites. The application of the array will permit linkage maps to be developed for QTL analyses in a cost-effective manner, and the identification of SNPs that have been assigned erroneous positions on the ‘Golden Delicious’ reference sequence will assist in the continued improvement of the genome sequence assembly for that variety.

Sequence and function of lysosomal and digestive cathepsin D-like proteinases of Musca domestica midgut

Musca domestica larvae display in anterior and middle midgut contents, a proteolytic activity with pH optimum of 3.0-3.5 and kinetic properties like cathepsin D. Three cDNAs coding for preprocathepsin D-like proteinases (ppCAD 1, ppCAD 2, ppCAD 3) were cloned from a M. domestica midgut cDNA library. The coded protein sequences included the signal peptide, propeptide and mature enzyme that has all conserved catalytic and substrate binding residues found in bovine lysosomal cathepsin D. Nevertheless, ppCAD 2 and ppCAD 3 lack the characteristic proline loop and glycosylation sites. A comparison among the sequences of cathepsin D-like enzymes from some vertebrates and those found in M. domestica and in the genomes of Aedes aegypti, Drosophila melanogaster, Tribolium castaneum, and Bombyx mori showed that only flies have enzymes lacking the proline loop (as defined by the motif: DxPxPx(G/A)P), thus resembling vertebrate pepsin. ppCAD 3 should correspond to the digestive cathepsin D-like proteinase (CAD) found in enzyme assays because: (1) it seems to be the most expressed CAD, based on the frequency of ESTs found. (2) The mRNA for CAD 3 is expressed only in the anterior and proximal middle midgut. (3) Recombinant procathepsin D-like proteinase (pCAD 3), after auto-activation has a pH optimum of 2.5-3.0 that is close to the luminal pH of M. domestica midgut. (4) Immunoblots of proteins from different tissues revealed with anti-pCAD 3 serum were positive only in samples of anterior and middle midgut tissue and contents. (5) CAD 3 is localized with immunogold inside secretory vesicles and around microvilli in anterior and middle midguit cells. The data support the view that on adapting to deal with a bacteria-rich food in an acid midgut region, M. domestica digestive CAD resulted from the same archetypical gene as the intracellular cathepsin D, paralleling what happened with vertebrates. The lack of the proline loop may be somehow associated with the extracellular role of both pepsin and digestive CAD 3. (C) 2009 Elsevier Ltd. All rights reserved.

Role of the triad N46, S106 and T107 and the surface charges in the determination of the acidic pH optimum of digestive lysozymes from Musca domestica

Structures of digestive lysozymes 1 and 2 from housefly (MdL1 and MdL2) show that S106-T107 delimit a polar pocket around E32 (catalytic acid/base) and N46 contributes to the positioning of 050 (catalytic nucleophile), whereas those residues are replaced by V109-A110 and D48 in the non-digestive lysozyme from hen egg-white (HEWL). Further analyses revealed that MdL1 and MdL2 surfaces are less positively charged than HEWL surface. To verify the relevance of these differences to the acidic pH optimum of digestive lysozymes it was determined that pKas of the catalytic residues of the triple mutant MdL2 (N46D-S106V-T107A) are similar to HEWL pKas and higher than those for MdL2. In agreement, triple mutant MdL2 and HEWL exhibits the same pH optimum upon methylumbelliferylchitotrioside. In addition to that, the introduction of six basic residues on MdL1 surface increased by 1 unit the pH optimum for the activity upon bacterial walls. Thus, the acidic pH optimum for MdL2 and MdL1 activities upon methylumbelliferylchitotrioside is determined by the presence of N46, S106 and T107 in the environment of their catalytic residues, which favors pKas reduction. Conversely, acidic pH optimum upon bacterial walls is determined by a low concentration of positive charges on the MdL2 and MdL1 surfaces. (C) 2010 Elsevier Inc. All rights reserved.

Efeitos de espécies de timbós (Derris spp.: Fabaceae) em populações de Musca domestica L.

A partir de ensaios com dosagens crescentes, foi avaliado o efeito do pó da raiz de duas espécies de timbó (Derris urucu e D. nicou) sobre populações de larvas de duas linhagens de Musca domestica L., provenientes de duas localidades do Estado de São Paulo, Jaboticabal (Jab) e Brodósqui (Bro). Para obtenção das doses letais foram utilizados ajustes de regressão de acordo com o modelo logístico. D. urucu foi mais eficiente que D. nicou no controle das duas linhagens, sendo necessário mais que o dobro da quantidade de D. nicou para se obter os mesmos efeitos causados com D. urucu. Foi demonstrada a existência de especificidade de ação dos timbós nas linhagens de moscas. D, urucu foi mais eficiente no controle da linhagem Bro, enquanto que D. nicou controlou maior número de indivíduos da linhagem Jab.

Avaliação dos níveis séricos de triiodotironina (T3) e tiroxina (T4) durante o ciclo estral em marrãs (Sus scrofa domestica - Linnaeus, 1758)

No presente trabalho, foram utilizadas 21 fêmeas suínas, virgens, sexualmente aptas, criadas e mantidas sob condições industriais, para observação dos perfis hormonais séricos de triiodotironina e tiroxina, durante o ciclo estral. As colheitas de sangue foram efetuadas sempre no mesmo intervalo, entre 8 e 10 horas. Cada animal foi submetido a 14 punções venosas, distribuídas nos dias zero, 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 21, 22 e 23 do ciclo estral. Consideramos o dia zero como o primeiro dia da fase estral, e o 23º dia como o primeiro do estro subseqüente. Os ensaios para dosagens hormonais foram executados utilizando-se a técnica de radioimunoensaio (RIE) em fase sólida e para isso foi empregado conjunto de reagentes comerciais (Coat-A-Count®). Para o hormônio triiodotironina foram observados valores altos no 1º e 23º dia e valor mínimo no 12º dia do ciclo estral. O hormônio tiroxina apresentou valores que oscilaram entre 2,42 e 3,74 mig/dl.

Avaliação dos níveis séricos de 17-alfa-OH progesterona e androstenediona durante o ciclo estral em marrãs (Sus Scrofa Domestica - Linnaeus, 1758)

No presente trabalho, foram utilizadas 21 fêmeas suínas, virgens, sexualmente aptas, criadas e mantidas sob condições industriais, para observação dos perfis hormonais séricos de 17-alfa-OH progesterona e androstenediona, durante o ciclo estral. As colheitas de sangue foram efetuadas sempre no mesmo intervalo, entre 8 e 10 horas. Cada animal foi submetido a 14 punções venosas, distribuídas nos dias zero, 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 21, 22 e 23 do ciclo estral. Considerou-se o dia zero como o primeiro dia da fase estral, e o 23º dia como o primeiro do estro subseqüente. Os ensaios para dosagens hormonais foram executados utilizando-se a técnica de radioimunoensaio (RIE) em fase sólida e para isso foi empregado conjunto de reagentes comerciais (Coat-A-Count®). Para o hormônio 17-alfa-OH progesterona, foram encontrados valores médios que variaram entre 0,18 e 2,7 ng/ml e para o hormônio androstenediona esses valores oscilaram entre 0,08 e 0,24 ng/ml.

Evaluation of thyroxine (T4) and progesterone-17-alpha-OH circadian rhythm in swine females (Sus scrofa domestica - Linnaeus, 1758)

No presente trabalho foram utilizadas quatro fêmeas suínas, adultas, mestiças, não-gestantes e sem sinais clínicos de estro, criadas e mantidas sob condições industriais de criação. Objetivou-se avaliar a ocorrência de ritmicidade biológica circadiana para tiroxina e 17-alfa -OH progesterona. Os ensaios para dosagens hormonais foram executados utilizando-se a técnica de radioimunoensaio (RIE) em fase sólida e para isso foi empregado conjunto de reagentes comerciais (COAT-A-COUNT R). As análises séricas de tiroxina mostraram valores mais elevados ao redor das 15 horas, decrescendo a partir dai até atingir níveis menores no intervalo da zero às 4 horas. Quanto a 17-alfa -OH progesterona, observaram-se níveis mais elevados por volta das 3 horas, decrescendo gradativamente ao longo do dia, até atingir menor concentração no intervalo das 12 às 15 horas.

The effect of refuge on Dermestes ater (Coleoptera : Dermestidae) predation on Musca domestica (Diptera : Muscidae): Refuge for prey or the predator?

It is believed that habitat heterogeneity can change the extent of predator-prey interactions. Therefore, in this study we examined the effect of habitat heterogeneity (characterized here as an addition of refuge) on D. ater predation on M. domestica. Predation of D. ater on M. domestica larvae was carried out in experimental habitats with and without refuge, and examined at different prey densities. The number of prey eaten by beetles over 24 h of predator-prey interaction was recorded, and we investigated the strength of interaction between prey and predator in both experimental habitats by determining predator functional response. The mean number of prey eaten by beetles in the presence of refuge was significantly higher than in the absence of refuge. Females had greater weight gains than males. Logistic regression analyses revealed the type II functional response for both experimental habitats, even though data did not fit well into the random predator model. Results suggest that the addition of refuge in fact enhanced predation, as prey consumption increased in the presence of refuge. Predators kept in the presence of refuge also consumed more prey at high prey densities. Thus, we concluded that the addition of refuge was an important component mediating D. ater-M. domestica population interactions. Refuge actually acted as a refuge for predators from prey, since prey behaviors detrimental to predators were reduced in this case.

Prey choice by Nesticodes rumpes (Araneae, Theridiidae) on Musca domestica (Diptera, Muscidae) and Dermestes ater (Coleoptera, Dermestidae)

Nesticodes rufipes is widely distributed in tropical and subtropical regions, being strongly associated with humans. However, few behavioral and ecological studies have investigated interspecific interactions between these spiders and insects of medical and veterinary importance. Here, we have investigated prey choice by N. rufipes when two different prey species, Musca domestica and Dermestes ater, were offered simultaneously. We also quantified the capture of these prey types by this predator in a poultry house and analyzed the association between prey-choice with physical characteristics of the prey. Finally, we discuss whether there is an antagonistic intraguild interaction in such a system composed of N. rufipes (top predator), D. ater (predator of larvae of M. domestica and prey of N. rufipes) and M. domestica (N. rufipes' prey). We found that Musca domestica were more abundant than D. ater in N. rufipes webs in the poultry house. Spiders given a choice of adults of M. domestica plus adults of D. ater, and also on adults plus larvae of M. domestica, preyed more on adult flies than on the other prey types. This preference was probably associated with the lesser mass and shorter lengths of adult flies. Our experiments demonstrated that the predation impact of N. rufipes on D. ater is low when compared to M. domestica. This result provides evidence that an antagonistic interaction between these predators does not occur, suggesting that they are in fact acting either synergistically or additively on M. domestica prey.

Population dynamics of Musca domestica (Diptera: Muscidae): experimental and theoretical studies at different temperatures

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Stochastic dynamics of Musca domestica (Diptera: Muscidae) and Chrysomya putoria (Diptera: Calliphoridae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

STRUCTURE OF THE RETE TESTIS OF THE CAT (FELIS-DOMESTICA, L)

The rete testis of the cat consists of 3 parts: a septal or interlobular part; a mediastinal part and a tunical part. The septal part contains the septal or transitory tubuli recti and the tubuli recti. The transitory tubules are formed as a confluence of the seminiferous tubules at the apex of the testicular lobules and the tubuli recti. The mediastinal rete is formed of long, straight channels which increase in size and become more irregular and anastomotic below the tunica albuginea at the cranial extremity of the testis. The end is characterized as the tunical part of the rete testis and communicates with the extratesticular rete testis. The channels all parts of the rete are lined by simple cuboidal or columnar epithelium. These epithelial channels are supported by a connective tissue containing smooth muscle cells. The framework tissue of the rete is more conspicuous at the cranial extremity of the testis, with a mio-connective matrix organization.