Measurement and Selection of Parasitoid Quality for Mass-Reared Trichogramma minutum Riley Used in Inundative Release

Parasitoid quality, subject to both genetic and environmental influences, is critical to the success of any biological control program, however, its measurement and improvement is poorly understood. In this study, a classic genetic approach is taken to develop two indices, namely a character index and a fitness index, for the measurement and selection of high quality parasitoids used in inundative release. Six life-history traits and corresponding fitness components in 33 inbred strains of Trichogramma minutum were used to generate both genotypic and phenotypic variance-covariance matrices that then allowed for the construction of the indices. Most traits and their fitness components were positively correlated, both phenotypically and genotypically, with lifetime fecundity and the number of female offspring appearing to have an important influence. Selection of the top three strains showed that parasitoid quality could be improved by 36% using the character index and possibly up to 150% using the fitness index. The two indices were linearly correlated suggesting that either could be used to measure quality. The character index is recommended because it requires information on only three life-history traits (fecundity, number of female offspring, and number of male offspring) and has highly correlated responses of fitness components. Our work demonstrates that the best quality T. minutum will be obtained by using the character index to select for inbred strains which have high fecundity and number of female offspring.     

Restriction fragment length polymorphism in amplification products of the bovine butyrophilin gene: assignment of bovine butyronhilin to bovine chromosome 23

A polymorphism was identified in the bovine butyrophilin (BTN) gene by digesting poly-merase chain reaction products with the restriction enzyme Hae III. This polymorphism was segregating in a Holstein-Friesian sire selected as part of an ongoing study directed towards the identification of quantitative trait loci affecting milk composition. Screening of a half-sib family established for the heterozygous sire allowed the localization of BTN to bovine chromosome 23 (BTA23).     

Lysine Succinylation of VBS Contributes to Sclerotia Development and Aflatoxin Biosynthesis in Aspergillus flavus

Aspergillus flavus is a common saprophytic and pathogenic fungus, and its secondary metabolic pathways are one of the most highly characterized owing to its aflatoxin (AF) metabolite affecting global economic crops and human health. Different natural environments can cause significant variations in AF synthesis. Succinylation was recently identified as one of the most critical regulatory post-translational modifications affecting metabolic pathways. It is primarily reported in human cells and bacteria with few studies on fungi. Proteomic quantification of lysine succinylation (Ksuc) exploring its potential involvement in secondary metabolism regulation (including AF production) has not been performed under natural conditions in A. flavus. In this study, a quantification method was performed based on tandem mass tag labeling and antibody-based affinity enrichment of succinylated peptides via high accuracy nano-liquid chromatography with tandem mass spectrometry to explore the succinylation mechanism affecting the pathogenicity of naturally isolated A. flavus strains with varying toxin production. Altogether, 1240 Ksuc sites in 768 proteins were identified with 1103 sites in 685 proteins quantified. Comparing succinylated protein levels between high and low AF-producing A. flavus strains, bioinformatics analysis indicated that most succinylated proteins located in the AF biosynthetic pathway were downregulated, which directly affected AF synthesis. Versicolorin B synthase is a key catalytic enzyme for heterochrome B synthesis during AF synthesis. Site-directed mutagenesis and biochemical studies revealed that versicolorin B synthase succinylation is an important regulatory mechanism affecting sclerotia development and AF biosynthesis in A. flavus. In summary, our quantitative study of the lysine succinylome in high/low AF-producing strains revealed the role of Ksuc in regulating AF biosynthesis. We revealed novel insights into the metabolism of AF biosynthesis using naturally isolated A. flavus strains and identified a rich source of metabolism-related enzymes regulated by succinylation.     

Intrageneric amplification of horse microsatellite markers with emphasis on the Przewalski's horse (E. pmewalskii)

Primer sequences flanking 13 microsatellite loci isolated from the domestic horse (E. caballus) were successfully used to amplify homologous loci in the Przewalski's horse (E. przewalskii). The results demonstrate that the level of polymorphism at all 13 loci in the Przewalski's horse was comparable to that in the domestic horse and the overall exclusion probability in the Przewalski's horse was calculated to be 0.9994. The results suggest that it should be possible to use E. caballus-derived microsatellite markers to provide parentage verification and additional valuable information to the captive management of E. przewalskii. The ability to amplify corresponding loci in the remaining five species of the genus was also confirmed, illustrating the general application of markers isolated from the domestic horse to the evaluation of polymorphism in the other six species of the genus.     

Campylobacter jejuni: Specific oligonucleotides and DNA probes for use in polymerase chain reaction-based diagnosis

Abstract A 1189 base-pair long DNA fragment, VS1, was isolated from a Campylobacter jejuni CIP 70.2 cosmid library and was found to contain regions specific for this bacterial species. For detection and identification of C. jejuni , two oligonucleotides derived from the VS1 sequence were used as primers in polymerase chain reaction test on genomic DNAs from 38 Campylobacter and from 10 non- Campylobacter strains. A specific, 358 base-pair long DNA fragment was amplified only when C. jejuni DNA was used as a target. The detection limit of the amplification reaction was as low as 1.86 fg DNA, which is the equivalent of one C. jejuni genome.     

Detection of Plasmodium sporozoites in mosquitoes by polymerase chain reaction and oligonucleotide rDNA probe, without dissection of the salivary glands

Dried Anopheles gambiae mosquito head + thorax portions, infected with Plasmodium falciparum sporozoites, were processed by the polymerase chain reaction. The PCR product was hybridized to an oligonucleotide probe (known as 114R or AW34) diagnostic for Plasmodium . The detection level by autoradiography was ten sporozoites per mosquito. Head + thorax of mosquitoes that contained mature P.falciparum oocysts, without sporozoites, gave no positive signal, indicating that the test detects only infective mosquitoes. This test can be applied to wild mosquito specimens collected, prepared and processed at different time intervals. The technique is convenient, highly sensitive, and could be used with a non-radioactive detection system and specific probes to differentiate Plasmodium spp.     

Comparative HILIC/ESI-QTOF-MS and HPTLC studies of pyrrolizidine alkaloids in flowers of Tussilago farfara and roots of Arnebia euchroma

The utility of HPTLC and HILIC/ESI-QTOF-MS for the determination of pyrrolizidine alkaloids (PAs) and their N-oxides (PANOs) was compared in the selected plant species: Tussilago farfara L. (TF, flower) and Arnebia euchroma (Royle) I.M. Johnst. (AE, root). HPTLC confirmed the postulated presence of PAs (saturated and unsaturated) or PANOs in the tested extracts. In accordance with previous studies, HILIC/ESI-Q-TOF-MS confirmed the presence of the toxic PA senkirkine and the saturated otonecine-type PAs, tussilagine and isotussilagine in the TF extract and 7-angeloylretronecine and 9-angeloylretronecine in AE extract. Moreover, the following alkaloids were identified in AE root: intermedine, intermedine-N-oxide, leptanthine-N-oxide, echimidine-N-oxide (or their corresponding stereoisomers) and traces of 7-angeloylretronecine and 9-angeloylretronecine-N-oxide. The study demonstrates the HILIC/ESI-Q-TOF-MS method to be a very useful tool for monitoring PAs and PANOs in the test samples, even when not all of the necessary standards are available. Quantitative analysis of senkirkine in TF flower by HILIC/ESI-QTOF-MS featured high resolution, high precision, high mass accuracy, and very high sensitivity with limit-of-detection (LOD) of 27.50 fg/μL and limit-of-quantitation (LOQ) of 91.60 fg/μL. The results from both methods may be used for the development or rejection of European Pharmacopoeia (X) monographs of both investigated species.     

Synthesis of Sphingosine Analogs with Acyclic Terpenoidal Carbon Skeleton

A simple and general synthetic method is described for erythro- and threo-forms of 2-amino-1,3-diols with an acyclic terpenoid-type side chain. ¹³C-NMR data are given for the triacetyl and N-acetyl derivatives of the synthetic erythro- and threo-2-amino-1,3-diols.