In situ measurement of cell stiffness of Arabidopsis roots growing on a glass micropillar support by atomic force microscopy

Atomic force microscopy (AFM) can measure the mechanical properties of plant tissue at the cellular level, but for in situ observations, the sample must be held in place on a rigid support and it is difficult to obtain accurate data for living plants without inhibiting their growth. To investigate the dynamics of root cell stiffness during seedling growth, we circumvented these problems by using an array of glass micropillars as a support to hold an Arabidopsis thaliana root for AFM measurements without inhibiting root growth. The root elongated in the gaps between the pillars and was supported by the pillars. The AFM cantilever could contact the root for repeated measurements over the course of root growth. The elasticity of the root epidermal cells was used as an index of the stiffness. By contrast, we were not able to reliably observe roots on a smooth glass substrate because it was difficult to retain contact between the root and the cantilever without the support of the pillars. Using adhesive to fix the root on the smooth glass plane overcame this issue, but prevented root growth. The glass micropillar support allowed reproducible measurement of the spatial and temporal changes in root cell elasticity, making it possible to perform detailed AFM observations of the dynamics of root cell stiffness.     

Development of simple sequence repeat markers and construction of a high-density linkage map of Capsicum annuum

To facilitate marker-assisted breeding and genetic analyses of pepper (Capsicum annuum), we developed non-redundant 2- or 3-base simple sequence repeat (SSR) markers from enriched C. annuum genomic libraries and from C. annuum cDNA sequences in public databases. The SSR-enriched libraries were constructed using combinations of three restriction enzymes (AluI, HaeIII, and RsaI) and two biotinylated oligonucleotides [b(GA)₁₅ and b(CA)₁₅]. Ultimately, we obtained 1,736 genomic SSR markers and 1,344 cDNA-derived SSR markers from 6,528 clones and 13,003 sequences, respectively. We mapped 597 markers, including 265 of the newly developed SSR markers, onto a linkage map by using doubled-haploid (DH) lines derived from an intraspecific cross of two pure lines of C. annuum (K9-11 × MZC-180). The map, designated as the KL-DH map, consisted of 12 linkage groups. The map covered a genetic distance of 2,028 cM, and the average distance between markers was less than 4 cM. The frame structure of the KL-DH map was compared with the published standard conserved ortholog set II (COSII) map, which was derived from an interspecific F₂ population (C. frutescens × C. annuum), by using tomato (Solanum lycopersicum) chromosomal sequences to bridge the two maps. The intraspecific KL-DH map constructed in this study and the interspecific COSII map were similar in map length and marker distribution, suggesting that the KL-DH map covers nearly the whole genome of C. annuum.     

Evidence for Finely-Regulated Asynchronous Growth of Toxoplasma gondii Cysts Based on Data-Driven Model Selection

Toxoplasma gondii establishes a chronic infection by forming cysts preferentially in the brain. This chronic infection is one of the most common parasitic infections in humans and can be reactivated to develop life-threatening toxoplasmic encephalitis in immunocompromised patients. Host-pathogen interactions during the chronic infection include growth of the cysts and their removal by both natural rupture and elimination by the immune system. Analyzing these interactions is important for understanding the pathogenesis of this common infection. We developed a differential equation framework of cyst growth and employed Akaike Information Criteria (AIC) to determine the growth and removal functions that best describe the distribution of cyst sizes measured from the brains of chronically infected mice. The AIC strongly support models in which T. gondii cysts grow at a constant rate such that the per capita growth rate of the parasite is inversely proportional to the number of parasites within a cyst, suggesting finely-regulated asynchronous replication of the parasites. Our analyses were also able to reject the models where cyst removal rate increases linearly or quadratically in association with increase in cyst size. The modeling and analysis framework may provide a useful tool for understanding the pathogenesis of infections with other cyst producing parasites.     

Different patterns of allopreening in the same-sex and opposite-sex interactions of juvenile large-billed crows (Corvus macrorhynchos)

Allogrooming, where an individual grooms another, has been extensively studied in various social animals to understand its role in the evolution of cooperation/prosociality. In existing studies in mammals, allogrooming has been suggested to exhibit not only a hygiene but also a social function. Allopreening, a topic of increasing interest in mammals but recently also in birds, has been studied mostly with mature animals. However, in some species immature individuals also show allopreening and its function remains poorly understood. Crows, Corvus spp., are an ideal model to study this phenomenon, because juveniles form year-round aggregates during their long juvenile stage (e.g., throughout 3–4 years). Here, we investigated the function of allopreening in juvenile groups of wild-caught large-billed crows (C. macrorhynchos). Allopreening frequency and duration for three groups of wild-caught juveniles were analysed to determine whether there was a symmetrical (i.e., reciprocal) or asymmetrical allopreening pattern, and if sex composition of the dyad and/or relative dominance of donor and recipient had an effect. We found that both the frequency and duration of male allopreening correlated with frequency of aggression. Allopreening between both males and females occurred unidirectionally from dominants to subordinates but not in the opposite direction. On the contrary, allopreening between a male and a female was found to be reciprocated, though the absolute frequency and duration were both greater in males than in females. These results suggest that the social function of allopreening in juvenile crows differs depending on the sex composition of the dyad, functioning as a dominance signal for same-sex dyads, and serving a social bonding function for opposite-sex dyads. These findings may reflect the potentially crucial roles of allopreening in within-sex competition and opposite-sex attraction during the 3 year-long juvenile stage affecting future mate choice in lifelong monogamy.     

Effect of Dietary Composition on Lipids in Serum and in Liver of Rats Fed a Cystine-excess Diet

The effects of dietary composition on lipids in serum and in liver of rats fed with a cystine- excess diet were investigated.

When starch was used as the carbohydrate source, the addition of excess-cystine caused an increase in serum cholesterol and phospholipids, and hepatomegaly. Phospholipids in serum of rats fed with a cystine-excess diet containing 5% corn oil were higher than those with a cystine-excess diet that was low in corn oil (0.1 %). The addition of konjac mannan and pectin prevented hypercholesterolemia, and the rise in phospholipids in serum was prevented by the addition of konjac mannan.

Liver cholesterol (mg/liver/100 g of body wt.) increased in rats fed with a cystine-excess diet.

The addition of excess cystine to a diet containing sucrose as the carbohydrate source resulted in a marked increase of cholesterol in serum and liver, and a decrease of serum triglycerides.

The replacement of starch by sucrose in the cystine-excess diet increased liver cholesterol.

Lipids, cholesterol, phospholipids and triglycerides in the liver, but not phospholipids, when expressed as mg per g of liver for rats fed with the diets containing sucrose, increased when compared to those for rats fed with the diets containing starch. In contrast, serum triglycerides increased.