The gene encoding juvenile hormone response (Krüppel homolog1, Kr-hl) in Coccinella septempunctata was investigated by cloning and analysing expression profiles in different developmental stages and tissues by quantitative real-time polymerase chain reaction (PCR). C. septempunctata Kr-hl (CsKr-hl) encoded a 1338 bp open reading frame (ORF) with a predicted protein product of 445 amino acids; the latter showed high similarity to orthologs in other species and contained eight highly-conserved Zn-finger motifs for DNA-binding. CsKr-hl was expressed in different developmental stages of C. septempunctata. The expression levels of CsKr-hl in eggs, 2nd, 3rd, 4th instar larvae, and pupa were 3.31, 2.30, 7.09, 0.58, and 7.48 times the number of 1st instar larvae, respectively. CsKr-hl expression levels in female adults gradually increased at 25–30 days and were significantly higher than expression at 1–20 days. CsKr-hl expression in 20–30 days-old male adults was significantly higher than males aged 1–15 days. CsKr-hl expression levels in heads of male and female adults were significantly higher than expression levels in the thorax, adipose, and reproductive system. Interestingly, CsKr-hl expression levels in the adipose and reproductive system of female adults were significantly higher than in adult male corresponding organs, which suggest that CsKr-hl plays an important role in regulating reproductive development in C. septempunctata.

Fat metabolism is an important and complex biochemical reaction in vivo and is regulated by many factors. Recently, the findings on high expression of fibroblast growth factor-16 (FGF16) in brown adipose tissue have led to an interest in exploring its role in lipogenesis and lipid metabolism. The study cloned the goat’s FGF16 gene 624 bp long, including the complete open reading frame that encodes 207 amino acids. We found that FGF16 expression is highest in goat kidneys and hearts, followed by subcutaneous fat and triceps. Moreover, the expression of FGF16 reached its peak on the 2nd day of adipocyte differentiation (P < 0.01) and then decreased significantly. We used overexpression and interference to study the function of FGF16 gene in goat intramuscular preadipocytes. Silencing of FGF16 decreased adipocytes lipid droplet aggregation and triglyceride synthesis. This is in contrast to the situation where FGF16 is overexpressed. Furthermore, knockdown of FGF16 also caused down-regulated expression of genes associated with adipocyte differentiation including CCAAT enhancer-binding protein beta (P < 0.01), fatty acid-binding protein-2 (P < 0.01) and sterol regulatory element binding protein-1 (P < 0.05), but the preadipocyte factor-1 was up-regulated. At the same time, the genes adipose triglyceride lipase (P < 0.01) and hormone-sensitive lipase (P < 0.05) associated with triglyceride breakdown were highly expressed. Next, we locked the fibroblast growth factor receptor-4 (FGFR4) through the protein interaction network and interfering with FGF16 to significantly reduce FGFR4 expression. It was found that the expression profile of FGFR4 in adipocyte differentiation was highly similar to that of FGF16. Overexpression and interference methods confirmed that FGFR4 and FGF16 have the same promoting function in adipocyte differentiation. Finally, using co-transfection technology, pc-FGF16 and siRNA-FGFR4, siRNA2-FGF16 and siRNA-FGFR4 were combined to treat adipocytes separately. It was found that in the case of overexpression of FGF16, cell lipid secretion and triglyceride synthesis showed a trend of first increase and then decrease with increasing interference concentration. In the case of interference with FGF16, lipid secretion and triglyceride synthesis showed a downward trend with the increase of interference concentration. These findings illustrated that FGF16 mediates adipocyte differentiation via receptor FGFR4 expression and contributed to further study of the functional role of FGF16 in goat fat formation.

It is known that a countable $\omega $ -categorical structure interprets all finite structures primitively positively if and only if its polymorphism clone maps to the clone of projections on a two-element set via a continuous clone homomorphism. We investigate the relationship between the existence of a clone homomorphism to the projection clone, and the existence of such a homomorphism which is continuous and thus meets the above criterion.

Five presumed biotypes were identified among field bindweed (Convolvulus arvensis L. ♯3 CONAR) clones collected from a field population near Lafayette, IN. Consistent variations in leaf morphology, floral characteristics, and accumulation of shoot and root biomass were found between biotypes when grown in a controlled environment. The biotypes also differed in their flowering capacity. The earliest flowering biotype formed flowers 23 days before the latest and produced 19 times more flowers per plant, which indicated further differences in seed production potential between biotypes. Pollination studies helped to differentiate biotypes within the population and showed that the presumed biotypes were self-incompatible. Vegetative reproduction potential of the biotypes varied from 1.8 to 74.5% in the number of root buds that developed into shoots. The variability in growth and reproduction observed between these field bindweed biotypes may explain the survival and adaptability of a population of this weed as environmental conditions and control practices change.

Although apomixis is the most common form of parthenogenesis in diplodiploid arthropods, it is uncommon in the haplodiploid insect order Hymenoptera. We found a new type of spontaneous apomixis in the Hymenoptera, completely lacking meiosis and the expulsion of polar bodies in egg maturation division, on the thelytokous strain of a parasitoid wasp Meteorus pulchricornis (Wesmael) (Braconidae, Euphorinae) on pest lepidopteran larvae Spodoptera litura (Fabricius) (Noctuidae). The absence of the meiotic process was consistent with a non-segregation pattern in the offspring of heterozygous females, and no positive evidence was obtained for the induction of thelytoky by any bacterial symbionts. We discuss the conditions that enable the occurrence of such rare cases of apomictic thelytoky in the Hymenoptera, suggesting the significance of fixed heterosis caused by hybridization or polyploidization, symbiosis with bacterial agents, and occasional sex. Our finding will encourage further genetic studies on parasitoid wasps to use asexual lines more wisely for biological control.

Clones have been in the forefront of biological interest for many years. Even so, open discussions continue to surround the concept of clonality, which has been recently much debated in the scientific literature, both in terms of philosophical meaning as well as empirical determination. Philosophically, the clone is the horizontally produced lineage from a single fertlized egg (e.g. mammals by division of the fertilized egg and representing a single generation) or vertically produced offspring (e.g. aphids representing different successive generations) from a single asexual stem mother (originally for a particular lineage, following hatching of the overwintering sexual egg in the spring); empirically, the aspect of genetic fidelity is also considered important, so-called clones being assumed to have an identical genome among clone mates. In reality of course, such members of a clonal lineage must differ at various regions of the genome, since mutation is a fundamental property of the DNA itself. Yet few studies have so far set out to show this empirically in eukaryotic organisms, which indulge in periods of asexual reproduction, sometimes, as in aphids, over many generations. In the present study, we have investigated asexual lineages of the grain aphid, Sitobion avenae (F.), a global pest of cereals, over five successive generations employing AFLP-PCR molecular techniques. Our main interest was to see how much variation was present in the early generations and if this variation was transmitted through the asexual lineages. By monitoring AFLP-based polymorphisms, we show that, in this aphid species, of a total of 110 individuals from two lineages tested (termed SA and SB), random mutations (band deletions, more rarely additions) were apparent from the third generation onwards, and although some mutations were found to be transmitted transgenerationally, others were rarely transmitted through the particular lineages they were detected in. Using Arlequin v. 2.0, average gene diversity within the lineages was found to be 0.024 ± 0.013 and 0.031 ± 0.016 for SA and SB, respectively. It was also found from the rearing of the lineages that one lineage, SA, was more fecund than the other lineage, SB, over the five generations (N = 818 vs. N = 358 total stem mothers plus nymphs for the two lineages, respectively).

Walking speeds were calculated for nine clones of the peach potato aphid Myzus persicae collected from three countries along a latitudinal cline of its European distribution from Sweden to Spain (Sweden, UK and Spain), and the effects of collection origin and intra and intergenerational acclimation were investigated. Walking speeds declined with decreasing temperature, with maximum performance at temperatures closest to acclimation temperature (fastest median walking speed of 5.8 cm min–1 was recorded for clone UK 3, collected from the UK, at 25°C after acclimating to 25°C for one generation). Following acclimation at both 20°C and 25°C, walking ceased (as indicated by median walking speeds of 0.0 cm min–1) at temperatures as high as 7.5°C and 12.5°C. However, acclimation at 10°C enabled mobility to occur to temperatures as low as 0°C. There was no relationship between mobility and latitude of collection, suggesting that large scale mixing of aphids may occur across Europe. However, clonal variation was suggested, with clone UK 3 outperforming the majority of other clones across all temperatures at which mobility was maintained following acclimation at 10°C for one and three generations and at 25°C for one generation. The Scandinavian clones consistently outperformed their temperate and Mediterranean counterparts at the majority of temperatures following acclimation for three generations at 25°C.

In Europe, aphids contribute significantly to the so-called ‘aerial plankton’ during the spring to autumn months (growing season), although individual flight behaviour has been found, especially from molecular ecological studies, to be species-specific in terms of migratory range (ambit). Many of these species individuals may be assumed to be clonal in origin, that is, derived from a single asexual foundress. We are presently studying two specialist aphid species on Tansy, Tanacetum vulgare L. from samples collected in Jena, Germany – Macrosiphoniella tanacetaria (Kaltenbach) and Metopeurum fuscoviride Stroyan, using microsatellite markers. On plotting the number of sets of different multilocus genotypes or MLGs (i.e. multiple clonal repeats: 1, 2, 3 copies, etc.), against the frequency of their occurrence, a negative exponential relationship was found, with populations of both species consisting mostly of single (i.e. unique) or low number repeats rather than larger multiple copy (clonal) MLG repeats. To test this further, microsatellite data collected from a previous study on M. tanacetaria in Jena in the year 2000 and on samples of the Grain aphid, Sitobion avenae (F.), collected in the UK in 1997/8, the latter both in the field and from 12.2 m high suction traps, were examined in the same way. Again, similar relations were found, with most MLGs occurring as unique or low copy number repeats. The data are briefly discussed in the light of our evidence, as well as that of other similar studies on other aphid species, relating aphid molecular genetic data to aphid life cycle, behaviour and ecology.

Nine clones were derived from a drug-resistant Trypanosoma congolense stock (IL 2856) and characterized in mice for their sensitivity to isometamidium chloride and diminazene aceturate. All clones were derived from the stock without drug selection and expressed high levels of resistance to isometamidium chloride (50% curative dose [CD50] values ranging from 1·5 to 5·1 mg/kg) and intermediate to high levels of resistance to diminazene aceturate (CD50 values ranging from 5·1 to 21·0 mg/kg). By contrast, the isometamidium chloride and diminazene aceturate CD50 values for a drug-sensitive clone, T. congolense IL 1180, were 0·018 mg/kg and 2·3 mg/kg, respectively. For both drugs, there appeared to be significantly different levels in expression of drug resistance amongst the 9 clones derived from IL 2856. Isoenzyme analysis of 7 enzymes showed that all 9 clones expressed the same electrophoretic variants. Thus, all 9 clones were identical for these phenotypic markers. The clone which expressed the highest level of resistance to isometamidium in mice (IL 3270) was transmitted to Boran cattle via the bite of infected Glossina morsitans centralis. IL 3270 produced an infection rate in tsetse of 5·0%. The resulting infections in cattle were shown to be resistant to intramuscular treatment with 2·0 mg/kg isometamidium chloride and 14·0 mg/kg diminazene aceturate. This contrasts with doses of 0·25 mg/kg isometamidium chloride or 3·5 mg/kg diminazene aceturate which are deemed sufficient to cure fully sensitive infections. Finally, 9 clones (subclones) were derived from IL 3270 and characterized in mice for their sensitivity to isometamidium chloride. Seven of the subclones expressed a significantly lower level of resistance to isometamidium than the parental clone and amongst the subclones there was significant variation in resistance. Thus, expression of a high level of resistance to isometamidium appears to be unstable in the rodent host and at least a component of the genetic determinant(s) for this drug-resistant phenotype is (are) likely to be unstable.

The occurrence of multiple-clone Plasmodium falciparum haploid blood-stage infections is a pre-requisite for cross-fertilization and genetic exchange at the diploid stage in the mosquito. Using monoclonal antibodies against 3 polymorphic blood-stage antigens, a method of two-colour differential immunofluorescence allowed the resolution of between 1 and 4 clones/isolate. A mean of 2 P. falciparum clones was detected in the blood of malaria patients in The Gambia. The mean number of clones/patient showed no correlation with age, parasitaemia, or disease severity. There was a slight difference in mean number between sample periods, probably reflecting temporal differences in transmission intensity. A statistical analysis of 2-locus genetic diversity of clones within isolates concludes that not all multiple-clone infections result from superinfection, but that some are due to single multiple-clone inoculations.

The bovine interferon-tau (bIFN-τ) gene, with signal sequence, was obtained through polymerase chain reaction (PCR) from bovine early embryos and subcloned into a pGEM-T vector. After being verified, the fragments, with or without signal sequence, were inserted into the expression vector pET-30a(+). Two recombinant plasmids were induced to express the recombinant proteins by isopropyl β-d-1-thiogalactopyranoside. The results showed that the bIFN-τ gene could be obtained from five bovine blastocysts by PCR without extraction of genomic DNA. It had 99% homology with nucleotides and 97% with amino acids in the GenBank sequence (accession number: XM_593584). The products of recombinant bIFN-τ, minus signal sequence, expressed in pET-30a(+) were analysed by SDS-PAGE. A new 20 kDa protein was detected and its molecular weight was as expected. The antiviral activity of recombinant bIFN-τ was 1×104 IU/mg using a standard cytopathic reduction assay. Marked morphological changes were induced by recombinant bIFN-τ in bovine endometrial epithelial cells. The cell volume was larger than that of controls and a lot of vesicles appeared in the cytoplasm after 24 h culture in the presence of 2.9 μg/ml recombinant bIFN-τ. In conclusion, a purified recombinant, biologically active bIFN-τ was obtained in this experiment.