Hyalomma (Hyalommina) arabica: nymphal and larval identity and spiny mouse hosts in Saudi Arabia (Acarina: Ixodoidea: Ixodidae)

Nymphal and larval Hyalomma (Hyalommina) arabica Pegram, Hoogstraal, and Wassef, 1982, described herein, closely resemble those of Hyalomma (Hyalommina) rhipicephaloides Neumann, differing chiefly in body size (both stages), nymphal basis capituli and scutal proportional dimensions, and distinctness of larval coxal spurs. Females of these species are also structurally similar but males differ in major critical characters. Nymphs and larvae of both species parasitize the spiny mouse (Acomys spp.), but immatures of the third African- Arabian species of this subgenus, Hyalomma (Hyalommina) punt Hoogstraal, Kaiser, and Pedersen, are unknown. Hyalomma (Hyalommina) arabica occurs in valleys and hills of western Saudi Arabia and western Yemen; H. (H.) rhipicephaloides in the Red Sea and Dead Sea areas; and H. (H.) punt in northeastern Somalia and eastern Ethiopia. The ibex (Capra ibex nubiana Cuvier) is probably the original host of adult H. (H.) arabica and H. (H.) rhipicephaloides; the related domestic goat is an important host of adults of the 3 species, which also parasitize domestic sheep. Gazelles are recorded hosts of adults of H. (H.) rhipicephaloides and H. (H.) punt and the latter is also recorded from goats, sheep, camels and cattle.     

Substitution of selenocysteine for cysteine in a reticulocyte lysate protein synthesis system

Selenocysteine occurs in the peptide backbone of several selenoenzymes. The mechanism, of selenocysteine incorporation has not been well characterized. The incorporation of selenocysteine into protein in a rabbit reticulocyte lysate (RRL) was studied at high levels of selenocysteine. [⁷⁵Se]Selenocysteine incorporation was inhibited by cycloheximide and by nuclease treatment. Random RNA copolymers were tested for protein synthesis activity in the messenger RNA-dependent RRL system. Of the active polymers, poly CIU and GU most strongly stimulated the incorporation of selenocysteine. In a series of four polymers with different ratios of U to G, incorporation of selenocysteine and cysteine increased with increasing percentages of U, suggesting that selenocysteine and cysteine responded to the same codon, presumably UGU. Of the 20 protein amino acids, only cysteine and cystine competed with selenocysteine incorporation. Selenocysteine was charged to cysteine-accepting tRNA in RRL. These results show that at supraphysiological concentrations selenocysteine can substitute for cysteine in RRL protein synthesis. Misincorporation of selenocysteine could be important when animal tissues contain high levels of selenium.     

Cryopreservation of non-encapsulated embryogenic tissue of sweet potato (Ipomoea batatas)

Embryogenic tissue of the sweet potato (Ipomoea batatas (L) LAM) genotype TIB 10 was established from in vitro axillary shoot tips on Murashige and Skoog (1962) medium supplemented with 5 M 2,4-dichlorophenoxyacetic acid. Embryogenic aggregates of fresh mass 9.0–12 mg were subjected to a rapid freezing protocol in liquid nitrogen following sucrose preculture and varying degrees of dehydration. Up to 50% of embryogenic explants survived rapid freezing after preculture on 0.4 or 0.7M sucrose only. Dehydration with silica gel to moisture contents in the range 18–41% improved the survival after cryopreservation of embryogenic tissue. Tissue dehydrated for intermediate periods exhibited poor survival. Following freezing, embryogenic tissue appeared to develop normally, retaining its competence to produce mature embryos and plantlets.Abbreviations BA 6-benzyladenine - 2,4-D 2,4-dichlorophenoxyacetic acid - MS Murashige and Skoog (1962) medium     

Successful synthesis of a glial-specific blood–brain barrier shuttle peptide following a fragment condensation approach on a solid-phase resin

Successful manual synthesis of the TD2.2 peptide acting as a blood–brain barrier shuttle was achieved. TD2.2 was successfully synthesised by sequential condensation of four protected peptide fragments on solid-phase settings, after several unsuccessful attempts using the stepwise approach. These fragments were chosen to minimise the number of demanding amino acids (in terms of coupling, Fmoc removal) in each fragment that are expected to hamper the overall synthetic process. Thus, the hydrophobic amino acids as well as Arg(Pbf) were strategically spread over multiple fragments rather than having them congested in one fragment. This study shows how a peptide that shows big challenges in the synthesis using the common stepwise elongation methodology can be synthesised with an acceptable purity. It also emphasises that choosing the right fragment with certain amino acid constituents is key for a successful synthesis. It is worth highlighting that lower amounts of reagents were required to synthesise the final peptide with an identical purity to that obtained by the automatic synthesiser.     

Thermal biology and locomotor performance of the Andean lizard Liolaemus fitzgeraldi (Liolaemidae) in Argentina

Ectotherms thermoregulate to maintain their body temperature within the optimal range needed for performing vital functions. The effect of climate change on lizards has been studied as regards the sensitivity of locomotor performance to environmental temperatures. We studied thermoregulatory efficiency and locomotor performance for Liolaemus fitzgeraldi in the Central Andes of Argentina. We determined body temperature, micro-environmental temperatures and operative temperatures in the field. In the laboratory, we measured preferred temperatures and calculated the index of thermoregulatory efficiency. We estimated the thermal sensitivity of locomotion by measuring sprint speed (initial velocity and long sprint) and endurance at five different body temperatures. Body temperature was not associated with either micro-environmental temperature, nor did it show differences with preferred temperatures. Thermoregulatory efficiency was moderate (0.61). Initial velocity and long sprint trials showed differences at different temperatures; however, endurance did not. Moreover, the optimal temperatures for the performance trials showed no significant differences among themselves. We conclude that Liolaemus fitzgeraldi has thermal sensitivity in locomotor performance with respect to body temperature and that it is an eurythermic lizard that experiences a large variation in body temperature and that has thermal flexibility in the cold.     

Patterns of Integration and Expression of Retroviral, Non-Replicative Vectors in Avian Embryos: Embryo Developmental Stage and Virus Subgroup Envelope Modulate Tissue-Tropism

We previously demonstrated that Avian Leukemia Viruses (ALV) carrying the v-myc gene specifically induce two types of tumors, cardiomyocytic tumors when the virus is injected before embryonic day 3 (E3), skin tumors when the virus is injected at E3 or E5.

Aiming to elucidate the mechanisms which determine this time-dependent change in target, we infected chick and quail embryos at E3 and E5 with replication-deficient, lacZ gene-carrying, ALV-based viruses produced by a packaging cell line. Three constructs driven by 3 different Long Terminal Repeats (LTRs) were tested and yielded similar results. When the constructs were inoculated at E3 and the lacZ gene product revealed 5 days later, around 70% of the embryos carried lacZ+ clones in the heart, around 50% had positive clones in the skin anywhere on the body, while a few embryos displayed clones in internal organs (liver, stomach, lungs). Immunocytological identification of the heart cell type(s) expressing the virus revealed that the only cells infected were cardiomyocytes. When the constructs were inoculated at E5, no lacZ+ clones appeared in the heart but all were located in the cephalic skin. In order to examine the relationship between viral integration and expression, DNA of different organs or tissues from lacZ stained embryos was analyzed by PCR. A tight correlation between integration and expression in the heart and in the skin was revealed in most cases. In contrast, a significant PCR signal was often detected in the liver or the stomach despite weak or absent expression as revealed by lacZ+ clones.

We then investigated the influence of envelope glycoprotein subgroups on the tropism of these constructs. The lacZ vector driven by RAV-2 LTRs was packaged as subgroups A, B or E viral particles. The A subgroup, used in the part of the study described above, infects both chick and quail while the B and E subgroups are specific for chick or quail respectively. These B and E subgroups induced lacZ+ clones in the heart (after E3 injection) while no clones or only a few were detected in the skin either after E3 or E5 injection. The following conclusions can be drawn: 1) cardiomyocytes are at E3 the major target for integration and expression of ALV-derived viruses in vivo; 2) targets change rapidly with embryonic age; and 3) tissue-specific infections depend on the envelope subgroup, thus presumably on the presence of the cognate receptor. This study clearly indicates that E3 inoculation of ALV-based retroviral vectors is a simple and powerful method to transfer gene sequences into cardiomyocytes and epidermal cells.     

In Silico Design of an Oseltamivir Derivative with Increased Affinity against Wild-Type and Mutant Variants of Neuraminidase and Hemagglutinin of Influenza A H1N1 Virus

Antiviral resistance has turned into a world concern nowadays. Influenza A H1N1 emerged as a problem at the world level due to the neuraminidase (NA) mutations. The NA mutants conferred resistance to oseltamivir and zanamivir. Several efforts were conducted to develop better anti-influenza A H1N1 drugs. Our research group combined in silico methods to create a compound derived from oseltamivir to be tested in vitro against influenza A H1N1. Here we show the results of a new compound derived from oseltamivir but with specific chemical modifications, with significant affinity either on NA (in silico and in vitro assays) or HA (in silico) from influenza A H1N1 strain. We include docking and molecular dynamics (MD) simulations of the oseltamivir derivative at the binding site onto NA and HA of influenza A H1N1. Additionally, the biological experimental results show that oseltamivir derivative decreases the lytic-plaque formation on viral susceptibility assays, and it does not show cytotoxicity. Finally, oseltamivir derivative assayed on viral NA showed a concentration-dependent inhibition behavior at nM, depicting a high affinity of the compound for the enzyme, corroborated with the MD simulations results, placing our designed oseltamivir derivative as a potential antiviral against influenza A H1N1.     

Use of histone antibodies for studying chromatin topography and the phosphorylation of chromatin subunits

Polyclonal and monoclonal antibodies specific for histones as well as sera directed against synthetic peptides of histones were used to probe the topography of chromatin subunits. In native chromatin, the regions corresponding to residues 130-135 of H3 and 6-18 of H2B were found to be exposed and able to interact with antibodies whereas the regions 26-35 and 36-43 of H2B and 80-89 and 85-102 of H4 were not. In vitro phosphorylation of H3 and H5 in native chromatin or of H3 in H1/H5-depleted chromatin led to a marked drop in the binding of antibodies specific for residues 130-135 of H3 and 6-18 of H2B. Phosphorylation of H1/H5-depleted chromatin also altered the degree of exposure of certain H2A epitopes but it did not affect the surface accessibility of residues 1-11 of H2B.     

Genetic control of the conversion of dihydroflavonols into flavonols and anthocyanins in flowers of Petunia hybrida

Petunia hybrida mutants, homozygous recessive for one of the genes An1, An2, An6, or An9 do not show anthocyanin synthesis in in vitro complementation experiments per se (see also Kho et al. 1977). Extracts of flowers of these mutants all provoke anthocyanin synthesis in isolated petals of an an3an3 mutant. Mutants homozygous recessive for one of the genes An1, An2, An6, or An9 and homozygous recessive for F1 accumulate dihydroflavonols in comparable amounts. The synthesis of dihydromyricetin is blocked in an1an1 mutants, which indicates a regulating effect of the gene An1 on the gene Hfl. Similar mutants, but dominant for F1, accumulate flavonols (kaempferol and quercetin) instead of dihydroflavonols. Myricetin is accumulated in minor amounts and not at all in an1an1 mutant. Furthermore, the synthesis of this flavonol is not controlled by the gene F1. The synthesis of cyanidin (derivatives) is greatly reduced when flavonols are synthesized (F1 dominant). In mutants dominant for Ht1 and Hf1 and thus able to synthesize cyanidin (derivatives) and delphinidin (derivatives), predominantly delphinidin (derivatives) are synthesized. The results indicate that kaempferol (derivatives), quercetin (derivatives), and delphinidin (derivatives) are the main endproducts of flavonoid biosynthesis in Petunia hybrida.     

cAMP concentration,morphological differentiation and citric acid production in Aspergillus niger

Summary The possibility that adenosine 3,5 monophosphate exerts an effect on citric acid production by Aspergillus niger by influencing pellet morphology has been investigated. The effect of pH and inoculum size on pellet formation, citric acid production, and intracellular and extracellular cAMP levels were studied. High levels of intracellular and extracellular cAMP in the later stages of the fermentation, the period of maximum citric acid formation, were associated with those treatments which gave pellets of intermediate size. The highest cAMP levels were associated with those treatments which gave the highest citric acid titre. It was concluded that high cAMP levels are principally associated with an optimum physiological state for citric acid production and that cAMP levels do not vary directly with pellet size.