Separation of precultured pollen from Nicotiana tabacum by aqueous polymer two-phase partition

Pollen isolated from cold treated and precultivated anthers of tobacco (Nicotiana tabacum L. var. Wisconsin 38) were separated into different fractions with counter-current distribution using an aqueous Dextran-polyethylene glycol two-phase system. It was possible to distinguish among eight pollen classes differing in developmental stage and in partitioning. A part of each fraction was cultivated for analysis of embryo formation. This was highest in a fraction with an intermediate to high partition in the phase system. Enriched in this fraction were also pollen that were fairly well stained with acetocarmine, contained several nuclei and had a relatively low mitochondrial activity. The enrichment of embryogenic pollen offers several advantages especially to physiological studies on embryogenesis.     

A method for computerized modification of certain natural animal sounds for communication study purposes

A new method for computerized modification of sound signals is presented. With digital signal processing in the time domain it is possible to alter the amplitude, the frequency and the time scale of natural sounds independently. The method can be applied to natural sounds with reasonably pure tonal quality.     

Quenching of the amidolytic activity of one-chain tissue-type plasminogen activator by mutation of lysine-416

In contrast to most other serine proteases, tissue-type plasminogen activator (t-PA) possesses enzymatic activity as the one-chain zymogen form. The hypothesis that lysine residues 277 or 416 may be involved in stabilization of an active conformation of one-chain t-PA via salt-bridge formation with aspartic acid residue 477 was tested by site-directed mutagenesis. Four recombinant t-PA mutants were constructed. The amidolytic activities of these analogues were compared to that of authentic t-PA. Substitution of arginine-275 provided an analogue [( R275G]t-PA) resistant to plasmin cleavage. The amidolytic activity of [R275G]t-PA was comparable to that of authentic one-chain t-PA, and so was the activity of [R275L,K277L]t-PA, in which additional substitution of lysine residue 277 was carried out. This suggested that its presence was nonessential for obtaining one-chain t-PA activity. In contrast, substitution of lysine residue 416 to obtain [K416S]t-PA and [K416S,H417T]t-PA resulted in substantial quenching of amidolytic one-chain activity. As expected, the amidolytic activities of the two-chain forms were less affected by the substitution. Involvement of lysine residue 416 in one-chain t-PA activity was also indicated by decreased activities of [K416S]t-PA and [K416S,H417T]t-PA with plasminogen as the substrate. The one-chain activity of the lysine residue 416 substitution analogues was partially restored in the presence of fibrin. This could indicate that strong ligands such as fibrin might provide an alternative stabilization of the active conformation of one-chain t-PA.     

Primary mesodermal dysgenesis of the cornea (Peters' anomaly) in two brothers

Summary In this report two brothers with a severe form of Peters' anomaly, i.e., primary dysgenesis mesodermalis of the cornea, are described. The thirddegree consanguinity of the parents is an additional argument for the autosomal recessive inheritance of this condition.     

Malaria Burden and Artemisinin Resistance in the Mobile and Migrant Population on the Thai–Myanmar Border, 1999–2011: An Observational Study

Background

The Shoklo Malaria Research Unit has been working on the Thai–Myanmar border for 25 y providing early diagnosis and treatment (EDT) of malaria. Transmission of Plasmodium falciparum has declined, but resistance to artesunate has emerged. We expanded malaria activities through EDT and evaluated the impact over a 12-y period.

Methods and Findings

Between 1 October 1999 and 30 September 2011, the Shoklo Malaria Research Unit increased the number of cross-border (Myanmar side) health facilities from two to 11 and recorded the number of malaria consultations. Changes in malaria incidence were estimated from a cohort of pregnant women, and prevalence from cross-sectional surveys. In vivo and in vitro antimalarial drug efficacy were monitored. Over this period, the number of malaria cases detected increased initially, but then declined rapidly. In children under 5 y, the percentage of consultations due to malaria declined from 78% (95% CI 76–80) (1,048/1,344 consultations) to 7% (95% CI 6.2–7.1) (767/11,542 consultations), p<0.001. The ratio of P. falciparum/P. vivax declined from 1.4 (95% CI 1.3–1.4) to 0.7 (95% CI 0.7–0.8). The case fatality rate was low (39/75,126; 0.05% [95% CI 0.04–0.07]). The incidence of malaria declined from 1.1 to 0.1 episodes per pregnant women-year. The cumulative proportion of P. falciparum decreased significantly from 24.3% (95% CI 21.0–28.0) (143/588 pregnant women) to 3.4% (95% CI 2.8–4.3) (76/2,207 pregnant women), p<0.001. The in vivo efficacy of mefloquine-artesunate declined steadily, with a sharp drop in 2011 (day-42 PCR-adjusted cure rate 42% [95% CI 20–62]). The proportion of patients still slide positive for malaria at day 3 rose from 0% in 2000 to reach 28% (95% CI 13–45) (8/29 patients) in 2011.

Conclusions

Despite the emergence of resistance to artesunate in P. falciparum, the strategy of EDT with artemisinin-based combination treatments has been associated with a reduction in malaria in the migrant population living on the Thai–Myanmar border. Although limited by its observational nature, this study provides useful data on malaria burden in a strategically crucial geographical area. Alternative fixed combination treatments are needed urgently to replace the failing first-line regimen of mefloquine and artesunate. Please see later in the article for the Editors'' Summary     

Molecular Dynamics Study of Intrinsic Stability in Six RNA Terminal Loop Motifs

Abstract

Single stranded RNA molecules can assume a wide range of tertiary structures beyond the canonical A-form double helix. Certain sequences, termed motifs, are more common than a random distribution would suggest. The existence of such motifs can be rationalized in structural terms. In this study, we have investigated the intrinsic structural stability of RNA terminal loop motifs using multiple MD simulations in explicit water. Representative loops were chosen from the major tetraloop motifs, including also the U-turn motif. Not all loops retain their folded starting structure, but lowering the temperature to 277 K, or adding adjacent base pairs from the stem to which the motif is attached, helps stabilizing the folded loop structure.     

Dawn song of male blue tits as a predictor of competitiveness in midmorning singing interactions

During the dawn chorus, territorial male songbirds vocalise intensively within signalling range of several conspecific males and can therefore be considered members of a busy communication network. The more or less continuous singing over a long period of time under standardised stimulus conditions makes the dawn song a potentially important information source both for simple receivers and for eavesdroppers. Male blue tits (Parus caeruleus) vary in features of their dawn song, e.g. older males sing longer strophes, and females choose males that sing longer strophes as extra-pair partners. However, so far, dawn song in the blue tit has been investigated separately from other singing behaviour of the same males. In this study, we investigate aspects of blue tit male quality, reflected in dawn song characteristics, and their predictive value for how males behave during singing interactions later in the morning. We acted as simple receivers by recording the singing activity of one male at a time at dawn and compared features of its dawn song, such as onset before sunrise, repertoire size, mean bout length, strophe length and percentage performance time to responses of the same male to a territory intrusion simulated by playback of synthesised songs later during the same morning. We assume that an aggressive response towards an intruder will involve a fast approach to the loudspeaker broadcasting strophes of blue tit song, searching for the intruder (flying around), and a high amount of counter singing and overlapping of the intruders songs. Aspects of vigour of response to the simulated intrusion could be predicted from all five investigated dawn song parameters as well as male age. This is, to our knowledge, the first indication that a simple receiver could extract reliable information from a males dawn singing behaviour about its competitiveness later in the day.Communicated by P.K. McGregor