Phenotype Classification of Zebrafish Embryos by Supervised Learning

Zebrafish is increasingly used to assess biological properties of chemical substances and thus is becoming a specific tool for toxicological and pharmacological studies. The effects of chemical substances on embryo survival and development are generally evaluated manually through microscopic observation by an expert and documented by several typical photographs. Here, we present a methodology to automatically classify brightfield images of wildtype zebrafish embryos according to their defects by using an image analysis approach based on supervised machine learning. We show that, compared to manual classification, automatic classification results in 90 to 100% agreement with consensus voting of biological experts in nine out of eleven considered defects in 3 days old zebrafish larvae. Automation of the analysis and classification of zebrafish embryo pictures reduces the workload and time required for the biological expert and increases the reproducibility and objectivity of this classification.     

Centrally Administered Ghrelin Acutely Influences Food Choice in Rodents

We sought to determine whether the orexigenic hormone, ghrelin, is involved in the intrinsic regulation of food choice in rats. Ghrelin would seem suited to serve such a role given that it signals hunger information from the stomach to brain areas important for feeding control, including the hypothalamus and reward system (e.g. ventral tegmental area, VTA). Thus, in rats offered a choice of palatable foods (sucrose pellets and lard) superimposed on regular chow for 2 weeks, we explored whether acute central delivery of ghrelin (intracerebroventricular (ICV) or intra-VTA) is able to redirect their dietary choice. The major unexpected finding is that, in rats with high baseline lard intake, acute ICV ghrelin injection increased their chow intake over 3-fold, relative to vehicle-injected controls, measured at both 3 hr and 6 hr after injection. Similar effects were observed when ghrelin was delivered to the VTA, thereby identifying the VTA as a likely contributing neurobiological substrate for these effects. We also explored food choice after an overnight fast, when endogenous ghrelin levels are elevated, and found similar effects of dietary choice to those described for ghrelin. These effects of fasting on food choice were suppressed in models of suppressed ghrelin signaling (i.e. peripheral injection of a ghrelin receptor antagonist to rats and ghrelin receptor (GHSR) knock-out mice), implicating a role for endogenous ghrelin in the changes in food choice that occur after an overnight fast. Thus, in line with its role as a gut-brain hunger hormone, ghrelin appears to be able to acutely alter food choice, with notable effects to promote “healthy” chow intake, and identify the VTA as a likely contributing neurobiological substrate for these effects.     

Chromosome studies in the red howler monkey, Alouatta seniculus stramineus (Platyrrhini, Primates): description of an X1X2Y1Y2/X1X1X2X2 sex-chromosome system and karyological comparisons with other subspecies

In the red howler monkey, Alouatta seniculus stramineus (2n = 47, 48, or 49), variations in diploid chromosome number are due to different numbers of microchromosomes. Males exhibit a Y;autosome translocation involving the short arm of an individual biarmed autosome. Consequently, the sex-chromosome constitution in the male is X1X2Y1Y2, with X1 representing the original X chromosome, X2 the biarmed autosome (No. 7), Y1 the Y;7p translocation product, and Y2 the acrocentric homolog of 7q. In the first meiotic division, a quadrivalent with a chain configuration can be observed in spermatocytes. Females have an X1X1X2X2 sex-chromosome constitution. Chromosome heteromorphisms were observed in pair 13, due to a pericentric inversion, and pair 19, due to the presence of constitutive heterochromatin. Microchromosomes, which varied in number between individuals, were also heterochromatic. NOR-staining was observed at two separate sites on a single chromosome pair (No. 10). A comparison of A.s. stramineus with A.s. macconnelli shows that these two subspecies have identical diploid chromosome numbers (47, 48, or 49), again due to a varying number of microchromosomes, and that they share a similar sex-chromosome constitution. Their karyotypes, however, are not identical, but can be derived from each other by a reciprocal translocation. Further comparisons with other A. seniculus subspecies reported in the literature indicate that this taxon is not karyologically uniform and that substantial chromosome shuffling has occurred between populations that have been considered to be subspecies by taxonomic criteria based on their morphometric attributes.     

Comparative mineral and hormonal analyses of wild type and TLS somaclonal variant derived from oil palm (Elaeis guineensis Jacq. var. tenera) tissue culture

Comparative mineral and hormonal analyses were made on tissue culture derived truncated leaf syndrome and wild type oil palm seedlings. Mineral analysis confirmed that Boron, Zinc and chlorophyll levels were significantly lower in truncated leaf syndrome leaves than those of wild type. Hormonal analysis also revealed various cytokinin derivatives such as trans-zeatin, trans-zeatin riboside, trans-zeatin O-glucoside and trans-zeatin riboside 5??mono phosphate were significantly higher in truncated leaf syndrome leaves compared to wild type leaves. Brassinolide level was also significantly higher in truncated leaf syndrome leaves than those of the wild type. These observations suggest that the truncated leaf syndrome abnormality could be associated to high cytokinin and brassinosteroid production which affects the uptake of Boron and Zinc.     

A New Real-Time PCR for the Detection of Plasmodium ovale wallikeri

It has been proposed that ovale malaria in humans is caused by two closely related but distinct species of malaria parasites: P. ovale curtisi and P. ovale wallikeri. We have extended and optimized a Real-time PCR assay targeting the parasite’s small subunit ribosomal RNA (ssrRNA) gene to detect both these species. When the assay was applied to 31 archival blood samples from patients diagnosed with P. ovale, it was found that the infection in 20 was due to P. ovale curtisi and in the remaining 11 to P. ovale wallikeri. Thus, this assay provides a useful tool that can be applied to epidemiological investigations of the two newly recognized distinct P. ovale species, that might reveal if these species also differ in their clinical manifestation, drugs susceptibility and relapse periodicity. The results presented confirm that P. ovale wallikeri is not confined to Southeast Asia, since the majority of the patients analyzed in this study had acquired their P. ovale infection in African countries, mostly situated in West Africa.     

DnaB proteolysis in vivo regulates oligomerization and its localization at oriC in Bacillus subtilis

Initiation of bacterial DNA replication at oriC is mediated by primosomal proteins that act cooperatively to melt an AT-rich region where the replicative helicase is loaded prior to the assembly of the replication fork. In Bacillus subtilis, the dnaD, dnaB and dnaI genes are essential for initiation of DNA replication. We established that their mRNAs are maintained in fast growing asynchronous cultures. DnaB is truncated at its C-terminus in a growth phase-dependent manner. Proteolysis is confined to cytosolic, not to membrane-associated DnaB, and affects oligomerization. Truncated DnaB is depleted at the oriC relative to the native protein. We propose that DNA-induced oligomerization is essential for its action at oriC and proteolysis regulates its localization at oriC. We show that DnaB has two separate ssDNA-binding sites one located within residues 1–300 and another between residues 365–428, and a dsDNA-binding site within residues 365–428. Tetramerization of DnaB is mediated within residues 1–300, and DNA-dependent oligomerization within residues 365–428. Finally, we show that association of DnaB with the oriC is asymmetric and extensive. It encompasses an area from the middle of dnaA to the end of yaaA that includes the AT-rich region melted during the initiation stage of DNA replication.