UNC-45 is required for NMY-2 contractile function in early embryonic polarity establishment and germline cellularization in C. elegans

The Caenorhabditis elegans UNC-45 protein is required for proper body wall muscle assembly and acts as a molecular co-chaperone for type II myosins. In contrast to other body wall muscle components, UNC-45 is also abundant in the germline and embryo. We show that maternally provided UNC-45 acts with non-muscle myosin II (NMY-2) during embryonic polarity establishment, cytokinesis and germline cellularization. In embryos depleted for UNC-45, myosin contractility is eliminated resulting in embryonic defects in polar body extrusion, cytokinesis and establishment of polarity. Despite a lack of contractility in an unc-45(RNAi) embryo, NMY-2::GFP localizes to the cortex and accumulates at the presumptive cytokinetic furrow indicating that UNC-45 is not required for cortical localization. UNC-45 and NMY-2 are also required for fertility since the lack of either component results in complete sterility due to failed initiation of the cellularization furrows that separate syncytial nuclei into germ cells. In the absence of UNC-45, the actomyosin cytoskeleton does not contract despite non-functional myosin still directly binding actin. UNC-45 has been previously suggested to be required for the folding of the myosin head, and our results refine this hypothesis suggesting that UNC-45 is not required to fold or maintain the actin binding domain but is still required for myosin function.     

RAB2A: a major subacrosomal protein of bovine spermatozoa implicated in acrosomal biogenesis

The perinuclear theca (PT) of mammalian sperm is a unique subcellular structure encapsulating the nucleus. Compositionally, the PT is made up of at least six prominent polypeptides (60, 36, 31, 28, 24, and 15 kDa), of which only two have been sequence identified, as well as many less prominent ones. As an ongoing process in unveiling the protein composition of the PT, we have uncovered the sequence identity of the prominent 24-kDa polypeptide (PT24). Initial N-terminal sequence analysis obtained by Edman degradation suggested that PT24 is a RAB2 protein. This was corroborated by mass spectrometric analyses of trypsin-digested fragments of PT24, identifying RAB2A of the RAB2 subfamily as the best sequence match. Quadrapole/time-of-flight analysis identified 72%% sequence coverage between PT24 and bull, human, mouse, or rabbit RAB2A. Since a genome search only identified two RAB2 subfamily members, RAB2A and RAB2B, the 72%% sequence coverage of PT24 provides assurance that this protein is RAB2A and not a new RAB2 subfamily member. Furthermore, commercial RAB2A antibodies, raised against oligopeptide fragments in the unique C-terminal region of RAB2A, specifically labeled PT24 on Western blot analysis of PT extracts. These anti-RAB2A antibodies, along with immune serum that we raised and affinity purified against isolated PT24, demonstrated at both light and electron microscope levels that RAB2 is associated with the periphery of the growing proacrosomic and acrosomic vesicles in the Golgi and cap phases of spermiogenesis and consequently assembled as part of the PT. This pattern of subacrosomal assembly is reminiscent of the pathway used by SubH2Bv (PT15), another prominent and exclusive subacrosomal protein, indicating a common route for subacrosomal-PT assembly. Traditionally somatic RAB2 proteins are involved in vesicular transport between the endoplasmic reticulum and the cis-side of the Golgi apparatus. Our study suggests an unprecedented direction of RAB2A-mediated vesicular transport in spermatids during acrosomal biogenesis, from the trans-side of the Golgi apparatus to the nuclear envelope.     

Spatially discordant alternans in cardiomyocyte monolayers

Repolarization alternans is a harbinger of sudden cardiac death, particularly when it becomes spatially discordant. Alternans, a beat-to-beat alternation in the action potential duration (APD) and intracellular Ca (Cai), can arise from either tissue heterogeneities or dynamic factors. Distinguishing between these mechanisms in normal cardiac tissue is difficult because of inherent complex three-dimensional tissue heterogeneities. To evaluate repolarization alternans in a simpler two-dimensional cardiac substrate, we optically recorded voltage and/or Cai in monolayers of cultured neonatal rat ventricular myocytes during rapid pacing, before and after exposure to BAY K 8644 to enhance dynamic factors promoting alternans. Under control conditions (n = 37), rapid pacing caused detectable APD alternans in 81% of monolayers, and Cai transient alternans in all monolayers, becoming spatially discordant in 62%. After BAY K 8644 (n = 28), conduction velocity restitution became more prominent, and APD and Cai alternans developed and became spatially discordant in all monolayers, with an increased number of nodal lines separating out-of-phase alternating regions. Nodal lines moved closer to the pacing site with faster pacing rates and changed orientation when the pacing site was moved, as predicted for the dynamically generated, but not heterogeneity-based, alternans. Spatial APD gradients during spatially discordant alternans were sufficiently steep to induce conduction block and reentry. These findings indicate that spatially discordant alternans severe enough to initiate reentry can be readily induced by pacing in two-dimensional cardiac tissue and behaves according to predictions for a predominantly dynamically generated mechanism.     

Alcohol abuse and suicide attempts among youth.

This study uses the Youth Risk Behavior Survey (YRBS) and the National Comorbidity Survey (NCS) to explore the causal relationship between alcohol abuse (binge drinking and clinically defined alcohol use disorders) and suicide attempts among youth. We use an empirical approach that allows one to assess the possible existence and strength of a causal relationship without relying on identifying assumptions. Our results suggest that a causal relationship between binge drinking and suicide attempts is very unlikely. The findings, however, support a causal relationship between clinically defined alcohol use disorders and suicide attempts among girls.     

Diving behaviour of dugongs, Dugong dugon

The diving behaviour of 15 dugongs (Dugong dugon) was documented using time-depth recorders (TDRs), which logged a total of 39,507 dives. The TDRs were deployed on dugongs caught at three study sites in northern Australia: Shark Bay, the Gulf of Carpentaria and Shoalwater Bay. The average time for which the dive data were collected per dugong was 10.4±1.1 (S.E.) days. Overall, these dugongs spent 47% of their daily activities within 1.5 m of the sea surface and 72% less than 3 m from the sea surface. Their mean maximum dive depth was 4.8±0.4 m (S.E.), mean dive duration was 2.7±0.17 min and the number of dives per hour averaged 11.8±1.2. The maximum dive depth recorded was 20.5 m; the maximum dive time in water >1.5 m deep was 12.3 min. The effects of dugong sex, location (study site), time of day and tidal cycle on diving rates (dives per hour), mean maximum dive depths, durations of dives, and time spent ≤1.5 m from the surface were investigated using weighted split-plot analysis of variance. The dugongs exhibited substantial interindividual variation in all dive parameters. The interaction between location and time of day was significant for diving rates, mean maximum dive depths and time spent within 1.5 m of the surface. In all these cases, there was substantial variation among individuals within locations among times of day. Thus, it was the variation among individuals that dominated all other effects. Dives were categorised into five types based on the shape of the time-depth profile. Of these, 67% of dives were interpreted as feeding dives (square and U-shaped), 8% as exploratory dives (V-shaped), 22% as travelling dives (shallow-erratic) and 3% as shallow resting dives. There was systematic variation in the distribution of dive types among the factors examined. Most of this variation was among individuals, but this differed across both time of day and tidal state. Not surprisingly, there was a positive relationship between dive duration and depth and a negative relationship between the number of dives per hour and the time spent within 1.5 m of the surface after a dive.     

Study of amide-proton exchange of Escherichia coli melibiose permease by attenuated total reflection-Fourier transform infrared spectroscopy: evidence of structure modulation by substrate binding.

The accessibility of Escherichia coli melibiose permease to aqueous solvent was studied following hydrogen-deuterium exchange kinetics monitored by attenuated total reflection-Fourier transform infrared spectroscopy under four distinct conditions where MelB forms different complexes with its substrates (H(+), Na(+), melibiose). Analysis of the amide II band upon (2)H(2)O exposure discloses a significant sugar protection of the protein against aqueous solvent, resulting in an 8% less exchange of the corresponding H(+)*melibiose*MelB complex compared with the protein in the absence of sugar. Investigation of the amide I exchange reveals clear substrate effects on beta-sheet accessibility, with the complex H(+)*melibiose*MelB being the most protected state against exchange, followed by Na(+)*melibiose*MelB. Although of smaller magnitude, similar changes in alpha-helices plus non-ordered structures are detected. Finally, no differences are observed when analyzing reverse turn structures. The results suggest that sugar binding induces a remarkable compactness of the carrier's structure, affecting mainly beta-sheet domains of the transporter, which, according to secondary structure predictions, may include cytoplasmic loops 4-5 and 10-11. A possible catalytic role of these two loops in the functioning of MelB is hypothesized.     

The potential of Chrysoperla rufilabris and Doru taeniatum as agents for dispersal of Spodoptera frugiperda nucleopolyhedrovirus in maize

The behaviour of two abundant predators in Mesoamerican maize crops, Chrysoperla rufilabris larvae and Doru taeniatum adults, towards healthy and nucleopolyhedrovirus-infected Spodoptera frugiperda larvae was compared. C. rufilabris did not discriminate between healthy and virus-infected prey, although the mean search time was approximately two times longer towards virus-infected larvae. In contrast, D. taeniatum directed a greater proportion of their attacks towards virus-infected prey but there was no significant difference in the search time. Prey consumption time did not differ significantly for each type of prey by either predator, although prey consumption was much faster in D. taeniatum. Viable virus was detected in D. taeniatum faeces up to 3 d after feeding on infected S. frugiperda larvae, whereas virus was inactivated in the gut of C. rufilabris. Both predators were shown to have acidic guts. A field experiment demonstrated that D. taeniatum that had fed on infected prey could contaminate foliage resulting in the transmission of the disease at a low prevalence (4.7%) to S. frugiperda larvae in a field maize crop.     

A novel artificial habitat collection device for studying resettlement patterns in anguillid glass eels

The number of glass eels Anguilla australis and A. reinhardtii caught in artificial habitat collectors, made from a PVC base and polyethylene split rope fibres, was related to the number of rope fibre tufts attached to each collector rather than collector area directly. Ageing of collectors in situ to promote algal growth enhanced the catch of glass eels. Glass eels entered the collectors at night primarily during the flood tide, and did not move into the collectors during daylight hours. Glass eel abundance increased with increasing distance from the freshwater drain located in the causeway. The artificial habitat collectors are effective for assessing relative numbers of resettling glass eels and may be useful for studying recruitment and settlement patterns of other anguillid eel species, as well as identifying areas and habitats within a catchment that provide important shelter for glass eels. Sampling glass eels can be carried out with maximum effect and minimum effort using compact, aged artificial habitat collectors on the night time flood tide when low tide coincides with dusk.     

Dipsticks for rapid detection of Plasmodium in vectoring Anopheles mosquitoes

Malaria remains the most serious vector-borne disease, affecting some 300-500 million people annually, transmitted by many species of Anopheles mosquitoes (Diptera: Culicidae). Monoclonal antibodies developed against specific circumsporozoite (CS) proteins of the main malaria parasites Plasmodium falciparum and P. vivax have been used previously for enzyme-linked immunosorbent assays (ELISA), widely employed for detection of malaria sporozoites in vector Anopheles for local risk assessment, epidemiological studies and targeting vector control. However, ELISA procedures are relatively slow and impractical for field use. To circumvent this, we developed rapid wicking assays that identify the presence or absence of specific peptide epitopes of CS protein of the most important P. falciparum and two strains (variants 210 and 247) of the more widespread P. vivax. The resulting assay is a rapid, one-step procedure using a 'dipstick' wicking test strip. In laboratory assessment, dipsticks identified 1 ng/ mL of any of these three CS protein antigens, with sensitivity nearly equal to the CS standard ELISA. We have developed and are evaluating a combined panel assay that will be both qualitative and quantitative. This quick and easy dipstick test (VecTest Malaria) offers practical advantages for field workers needing to make rapid surveys of malaria vectors.