Dynamics of natural bovine herpesvirus-1 (BoHV-1) infection in a dairy herd

In this study, natural cycling of BoHV-1 infection was investigated in two groups of dairy cattle containing 2120 head. Group 1 comprised 127 animals and they were monitored for BoHV-1 infection virologically and serologically in six consecutive sampling periods. It consisted of naive heifers between 6 and 8 months of age, while in group 2, age, sex and the BoHV-1 serostatus of the animals were disregarded. The animals in group 1 were found to have seroconverted at the second sampling. Results of the serological study showed slight antibody response after natural BoHV-1 infection in the herd and neutralizing titres fell below protective levels in the 6–8 months after the peak. During the 2-year study period, one recurrence was detected after primary infection. Virus isolation studies revealed a cytopathic effect indicative of BoHV-1 in two nasal swabs taken during the fifth sampling period from animals with mild upper respiratory tract symptoms. As the study was carried out under natural conditions, it is not known whether the viruses isolated were from recurrences or re-infections. Data from cross-neutralization tests with herd isolates showed higher antibody response than those with the reference virus. The dynamics of BoHV-1 in both groups were found to be statistically similar.     

Measurement of the attention time in the horse

A study carried out on 49 horses showed that it is possible to measure the attention time by operant conditioning. After teaching horses an instrumental task using a signal, we were then able to test their attention time by asking them to prolong it increasingly while setting success and failure criteria. Two tests were performed 3 weeks apart. The 2nd test was feasible without relearning, a proof of memory, and was repeatable, a proof of consistency in the attention time. A significant difference was observed between the 3 age groups. Young horses often performed very well during the 1st test but their attention dropped in the 2nd test while older horses were more stable with respect to attention and even increased it slightly. The study shows that there are individual differences but it was not possible to prove a significant influence of breed, gender and paternal influence. Consequently, learning appears to be one of the most interesting approaches for evaluating the attention of horses and for observing their behaviour.     

Validation of endogenous reference genes for expression profiling of RAW264.7 cells infected with Mycobacterium avium subsp. paratuberculosis by quantitative PCR

Reference genes are frequently used to normalize between different biological samples the levels of mRNA measured using quantitative PCR (qPCR). The expression level of many commonly used reference genes has been shown to vary between tissues or cells, or following exposure to various treatments including infection with microbes. The selection of an appropriate reference gene for an individual experiment is therefore a crucial step in the process of accurately determining changes in gene expression. For this purpose, we analyzed the expression of nine commonly used reference genes in a murine macrophage cell line, RAW264.7, for their potential use in the analysis of differential gene expression by quantitative polymerase chain reaction (qPCR) following experimental infection with Mycobacterium avium subsp. paratuberculosis. Only one of nine putative reference genes tested, casc3a, was found to be suitable, and combinations of two or more reference genes were disadvantageous. Based on data from the study, we recommend an approach for selection of reference genes, conducting assays with technical replicates in duplicate rather than triplicate, determining decision-limit quality control criteria for technical replicates and assessing the significance of gene expression fold differences using DeltaDeltaC(t) based on knowledge of the variation in the reference gene.     

Functional genomics of microspore embryogenesis

Isolated plant microspores, when stressed and cultured in vitro, can be diverted from their normal gametophytic pathway towards sporophytic development, with the formation of haploid embryos and ultimately doubled-haploid plants. This process is called androgenesis or microspore embryogenesis, and is widely used in plant breeding programmes to generate homozygous lines for breeding purposes. Protocols for the induction of microspore embryogenesis and the subsequent regeneration of doubled haploid (DH) plants have been successfully developed for more than 200 species. These practical advances stand in stark contrast to our knowledge of the underlying molecular genetic mechanism controlling this process. The majority of information regarding the genetic and molecular control of the developmental switch from gametophytic to sporophytic development has been garnered from four intensely studied (crop) plants comprising two dicotyledonous species, rapeseed (Brassica napus) and tobacco (Nicotiana tabacum), and two monocotyledonous species, wheat (Triticum aestivum) and barley (Hordeum vulgare). In these species the efficiency of microspore embryogenesis is very high and reproducible, making them suitable models for molecular studies. In the past, molecular studies on microspore embryogenesis have focussed mainly on the identification of genes that are differentially expressed during this developmental transition and/or early in embryo development, and have identified a number of genes whose expression marks or predicts the developmental fate of stressed microspores. More recently, functional genomics approaches have been used to obtain a broad overview of the molecular processes that take place during the establishment of microspore embryogenesis. In this review we summarise accumulated molecular data obtained in rapeseed, tobacco, wheat and barley on embryogenic induction of microspores and define common aspects involved in the androgenic switch.     

A 192bp allele at the <Emphasis Type="Italic">Xgwm261</Emphasis> locus is not always associated with the <Emphasis Type="Italic">Rht8</Emphasis> dwarfing gene in wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.)

The height-reducing gene Rht8 was introduced into Italian wheats by breeder Nazareno Strampelli from the Japanese landrace Akakomugi, and has been widely used in wheats adapted to southern and eastern European conditions. Following identification of a close linkage to Rht8, microsatellite marker Gwm261 has been used extensively to screen large numbers of diverse international germplasm. A 192bp allele at this locus has been taken as “diagnostic” for Rht8 and used to infer the international distribution of Rht8. In this paper, we report several instances of cultivars and mapping populations that vary for the presence of the 192bp allele at the Xgwm261 locus (Xgwm261 ₁₉₂ ), but with no associated reduction in height, suggesting a lack of association with Rht8. For instance, in the population derived from a cross between Sunco (Rht-B1b, Xgwm261 ₁₆₅ ) and Tasman (Rht-D1b, Xgwm261 ₁₉₂ ), there were significant height differences associated with the segregation of Rht-B1b and Rht-D1b, but no height differences between Xgwm261 genotypes. Similar results were obtained in a population derived from the cross between Molineux (Rht-B1b, Xgwm261 ₁₉₂ ) and Trident (Rht-D1b Xgwm261 ₂₀₈ ). In contrast, the cross between Trident and Chuanmai 18 (Xgwm261 ₁₉₂ ) gave significant height effects at both the Rht-D1 and Xgwm261 loci, with no epistatic interaction between loci. Chuanmai 18 is closely related to the Strampelli wheat Mara (ancestrally derived from Akakomugi) and is therefore likely to carry Rht8. The old Japanese cultivar Norin 10, used by Norman Borlaug to introduce Rht-B1b and Rht-D1b into Mexican wheats, also has a 192bp allele at the Xgwm261 locus, and the sequence of the amplified product is identical to that of Akakomugi. We suggest that the widespread use of Norin 10-derived germplasm during and after the Green Revolution introduced a second haplotype into international germplasm, in which Xgwm261 ₁₉₂ has no association with Rht8. Therefore, the presence of Xgwm261 ₁₉₂ is only indicative of Rht8 in wheat cultivars that have inherited this allele from Akakomugi or a Strampelli wheat ancestor.     

Integration of transgenes into sexual polyploidization schemes for potato (Solanum tuberosum L.)

Most potato transgenic research has focused on development of resistance to pathogens and modification of potato physiology. Many transgenes, particularly those conferring pathogen resistance, could substantially lower potato production costs in developing countries. However, transgenes have not been reported in sexually propagated 4x-2x potato hybrids commonly grown in developing countries. Two transgenes,the Bacillus thuringiensis cry3Aa endotoxin protein gene and the PVY°coat protein gene, were engineered intodiploid and tetraploid potato using Agrobacterium tumefaciens-mediated transformation. Cry3Aa was produced at high levels in several lines while the PVY° coat protein was not expressed. Diplandroid and tetraploid genotypes were crossed to produce transgenic 4x-2x hybrids. Genetic transformation had no discernable effect on fertility ofthe primary transformants, germination of4x-2x seed derived from the transformants and agronomic performance(tuber set, average tuber weight and total tuber yield) of the 4x-2xhybrids. The transgenic 4x-2xhybrids produced non-viable pollen and could only be crossed as female parents. Results suggest that transgenes, such ascry3Aa, could be expressed in 4x-2x hybrids to lower costs of production with no significant effect on plant phenotype. This revised version was published online in July 2006 with corrections to the Cover Date.     

Genetic variation in bitter pit and fruit calcium concentrations within a diverse apple germplasm collection

Summary Genetic variability in the apple [Malus] fruit disorder bitter pit and fruit calcium concentrations ([Ca]) was determined in 25 seedling families at each of two sites from 1999 to 2000 and again for one site, in 2001. Most trees were free of pit or had low pit incidence, which could be approximated by an over-dispersed binomial distribution. A genetic component to external and internal pit incidence was found across both sites and all years. This effect was irrespective of the use of several cropping factors, average fruit weight or fruit mineral concentrations as covariates in the analyses. Year and/or site always had substantial effects on pit incidence. Interactions of genetic effects with site or year were also detected with large differences in mean pit incidence between sites or years for some families but not others. No relationship was found between mean family pit incidence and mean family [Ca] or mean family harvest date. However fruit [Ca] and harvest date were usually important predictors of within-family variation in pit incidence. For any one seedling, family had the largest effect on bitter pit incidence followed by site and [Ca] and the smallest effect was that of harvest date. [Ca] showed a strong genetic component and estimated family means of [Ca] were consistent at different sites and years. These results suggest that susceptibility of apple genotypes to bitter pit in breeding programmes may be best assessed through screening across multiple sites and seasons. Fruit [Ca] might be useful as an indirect selection index for bitter pit within, but not among families.     

Production and use of maize seed for sowing in Bolívar,Ecuador

In the province of Bolívar in the Andean region 212 farmers, six salesmen and 14 technicians, involved in maize production, were interviewed. The majority of the farmers interviewed were small-holders; 64% of the farmers had a farm size of less than 4.5 ha. Maize is the most important crop and is often grown in association with beans. Most maize, 96%, is soft maize meant for human consumption. The cultivars grown are open pollinated ones. One cultivar, ‘Guagal’, was grown extensively. A few others, among which an improved version of Guagal were of some importance. In the production of maize the farmers consider damage due to insects, wind (lodging), diseases, drought and hail, the low prices for their surplus maize and the high costs of fertilizers, chemicals, sowing seed and labour as the most important constraints. The seed for sowing the next crop mainly came from seed kept from the last harvest. Farmers indicated that they select from the harvested ears the healthy looking larger ones. Kernel type also was a selection criterion. Selection for plant type in the field was rarely done. Of the farmers 71% stored the maize as kernels while 29% stored the seed kept on the ears. The storage was predominantly done somewhere in the house in plastic (56%) or cloth bags (14%), in a pile (13%), just on the floor (4%) or in cans (4%). Some farmers still had seed left from the last sowing and were willing to give a sample for testing. From 32 such samples the quality was assessed by INIAP, the Ecuadorean governmental research and breeding organisation. The samples were consistently of a low quality. Especially the vigour of the seedlings was poor and most of the seeds were infected or contaminated by fungi. In the region a few salesmen offer maize seed for sale of cultivar Guagal at prices affordable by the farmer. Of improved cultivars the prices were higher than most farmers are willing to pay. The technicians considered good seed quality and good maintenance of the cultivars effective methods in obtaining better yields. Many farmers do not have access to seed of INIAP and are not familiar with the improved cultivars.