Constitutively active myosin light chain kinase alters axon guidance decisions in Drosophila embryos

Conventional myosin II activity provides the motile force for axon outgrowth, but to achieve directional movement during axon pathway formation, myosin activity should be regulated by the attractive and repulsive guidance cues that guide an axon to its target. Here, evidence for this regulation is obtained by using a constitutively active Myosin Light Chain Kinase (ctMLCK) to selectively elevate myosin II activity in Drosophila CNS neurons. Expression of ctMLCK pan-neurally or in primarily pCC/MP2 neurons causes these axons to cross the midline incorrectly. This occurs without altering cell fates and is sensitive to mutations in the regulatory light chains. These results confirm the importance of regulating myosin II activity during axon pathway formation. Mutations in the midline repulsive ligand Slit, or its receptor Roundabout, enhance the number of ctMLCK-induced crossovers, but ctMLCK expression also partially rescues commissure formation in commissureless mutants, where repulsive signals remain high. Overexpression of Frazzled, the receptor for midline attractive Netrins, enhances ctMLCK-dependent crossovers, but crossovers are suppressed when Frazzled activity is reduced by using loss-of-function mutations. These results confirm that proper pathway formation requires careful regulation of MLCK and/or myosin II activity and suggest that regulation occurs in direct response to attractive and repulsive cues.     

Site-specific biosynthetic incorporation of a fluorescent tag into proteins via cysteine-tRNA(Cys)

Site-specific incorporation of biophysical probes into proteins during translation can permit structure/function studies on selected proteins in heterogeneous environments. We report here a procedure for incorporating a fluorescent tag into proteins via Escherichia coli Cys-tRNA(Cys) during in vitro protein synthesis. Naturally occurring Cys-tRNA(Cys) is an attractive vehicle for fluorophore incorporation since it can be readily prepared in quantity and reacted with commercially available fluorophores. Moreover, proteins can often be constructed with a single Cys so that fluorophore incorporation results in a tag at a unique site.     

Is it possible to exclude a diagnosis of myocardial damage within six hours of admission to an emergency department? Diagnostic cohort study

ObjectiveTo assess the clinical efficacy and accuracy of an emergency department based six hour rule-out protocol for myocardial damage.DesignDiagnostic cohort study.SettingEmergency department of an inner city university hospital.Participants383 consecutive patients aged over 25 years with chest pain of less than 12 hours'' duration who were at low to moderate risk of acute myocardial infarction.InterventionSerial measurements of creatine kinase MB mass and continuous ST segment monitoring for six hours with 12 leads.ResultsOutcome of the gold standard test was available for 292 patients. On the diagnostic test for the protocol, 53 patients had positive results and 239 patients had negative results. There were 18 false positive results and one false negative result. Sensitivity was 97.2% (95% confidence interval 95.0% to 99.0%), specificity 93.0% (90.0% to 96.0%), the negative predictive value 99.6%, and the positive predictive value 66.0%. The positive likelihood ratio was 13.9 and the negative likelihood ratio 0.03.ConclusionsThe six hour rule-out protocol for myocardial infarction is accurate and efficacious. It can be used in patients presenting to emergency departments with chest pain indicating a low to moderate risk of myocardial infarction.

What is already known on this topic

Many patients with chest pain in emergency departments indicating a low to moderate risk of myocardial infarction are admitted to rule out myocardial damageSome 6% of those discharged have undiagnosed myocardial damage

What this study adds

An emergency department based chest pain assessment unit protocol to rule out myocardial damage is sensitive enough to allow safe discharge of patients at low to moderate risk of myocardial infarction within six hoursSuch units can also reduce the number of patients admitted unnecessarily     

Interactions among endophytic bacteria and fungi: Effects and potentials

Plants benefit extensively by harbouring endophytic microbes. They promote plant growth and confer enhanced resistance to various pathogens. However, the way the interactions among endophytes influence the plant productivity has not been explained. Present study experimentally showed that endophytes isolated from rice (Oryza sativa) used as the test plant produced two types of interactions; biofilms (bacteria attached to mycelia) and mixed cultures with no such attachments. Acidity, as measured by pH in cultures with biofilms was higher than that of fungi alone, bacteria alone or the mixed cultures. Production of indoleacetic acid like substances (IAAS) of biofilms was higher than that of mixed cultures, fungi or bacteria. Bacteria and fungi produced higher quantities of IAAS than mixed cultures. In mixed cultures, the potential of IAAS production of resident microbes was reduced considerably. There was a negative relationship between IAAS and pH of the biofilms, indicating that IAAS was the main contributor to the acidity. However, such a relationship was not observed in mixed cultures. Microbial acid production is important for suppressing plant pathogens. Thus the biofilm formation in endophytic environment seems to be very important for healthy and improved plant growth. However, it is unlikely that an interaction among endophytes takes place naturally in the endophytic environment, due to physical barriers of plant tissues. Further, critical cell density dependant quorum sensing that leads to biofilm formation may not occur in the endophytic environment as there is a limited space. As suchin vitro production and application of beneficial biofilmed inocula of endophytes are important for improved plant production in any agro-ecosystem. The conventional practice of plant inoculation with monocultures or mixed cultures of effective microbes may not give the highest microbial effect, which may only be achieved by biofilm formation.     

Dairy propionibacteria as direct-fed microbials: in vitro effect on acid metabolism of <Emphasis Type="Italic">Streptococcus bovis</Emphasis> and <Emphasis Type="Italic">Megasphaera elsdenii</Emphasis>

Ruminal acidosis caused by accumulation of lactic acid, a decrease of pH in the rumen and subsequent imbalance of the rumen fermentation process, affects the health and productivity of dairy cows and beef cattle. Direct-fed microbials have potential for use in the control and prevention of ruminal acidosis. This study investigated the interaction between five strains of dairy propionibacteria, Megasphaera elsdenii and Streptococcus bovis in various co-culture combinations in a simulated rumen environment comprising unmodified rumen digesta supplemented with excess glucose. While suppression of lactic acid accumulation by both the dairy propionibacteria and M. elsdenii in the presence of S. bovis in the simulated rumen conditions was evident, propionibacteria were found to be more effective than M. elsdenii in controlling lactic acid levels.     

Nanoparticle-Encapsulated Chlorhexidine against Oral Bacterial Biofilms

Background

Chlorhexidine (CHX) is a widely used antimicrobial agent in dentistry. Herein, we report the synthesis of a novel mesoporous silica nanoparticle-encapsulated pure CHX (Nano-CHX), and its mechanical profile and antimicrobial properties against oral biofilms.

Methodology/Principal Findings

The release of CHX from the Nano-CHX was characterized by UV/visible absorption spectroscopy. The antimicrobial properties of Nano-CHX were evaluated in both planktonic and biofilm modes of representative oral pathogenic bacteria. The Nano-CHX demonstrated potent antibacterial effects on planktonic bacteria and mono-species biofilms at the concentrations of 50–200 µg/mL against Streptococcus mutans, Streptococcus sobrinus, Fusobacterium nucleatum, Aggregatibacter actinomycetemcomitans and Enterococccus faecalis. Moreover, Nano-CHX effectively suppressed multi-species biofilms such as S. mutans, F. nucleatum, A. actinomycetemcomitans and Porphyromonas gingivalis up to 72 h.

Conclusions/Significance

This pioneering study demonstrates the potent antibacterial effects of the Nano-CHX on oral biofilms, and it may be developed as a novel and promising anti-biofilm agent for clinical use.     

Comparative anatomical study of the gracilis and coracobrachialis muscles: implications for facial reanimation

Since the introduction of cross-facial nerve grafting and free vascularized muscle transfer for the treatment of longstanding facial paralysis, substantial progress has been made toward restoration of facial expression that is as normal as possible. Much of the focus has remained on the gracilis as a donor muscle. However, its inherent anatomical characteristics may preclude it from ever being more than simply a mass of contractile tissue in the face. The coracobrachialis muscle, which is the analogue in the arm of the lower limb adductor mass, was proposed as an alternative donor muscle because it was thought that certain features would allow it to improve on the overall results that are currently possible with the gracilis. A comparative anatomical study was conducted to gauge this potential. A total of 133 muscles were analyzed, including 96 dissected specimens, 16 arterial and 14 venous study specimens, and seven neurovascular study specimens. Anatomical parameters were recorded for each muscle and later tabulated. Histological analysis of the nerves to 10 gracilis and 10 coracobrachialis muscles was performed, and the findings were confirmed with intraneural dissection of an additional 20 nerves under an operating microscope. The coracobrachialis was observed to be a practical alternative to the gracilis. Indeed, it has many of the attributes that initially drew attention to the gracilis as a possible donor muscle, including a reliable neurovascular supply, minimal donor-site morbidity, and the option of having two teams operate simultaneously. In addition, it has a size, shape, and form that make it an excellent choice for transfer to the face. It could be easily attached in the face to provide static support as well as animation, because of its long proximal tendon, the thick intermuscular septum along its lateral surface, and, when present, the ligament of Struthers.     

Concordance of genetic and phenotypic characters across a sapsucker hybrid zone

Hybridization has presented a challenge for taxonomists and conservation biologists, since hybridizing forms could be stable evolutionary entities or ephemeral forms that are blending together. However, hybrid zones also provide a unique opportunity for evolutionary biologists who study the interaction between gene flow and reproductive isolation in speciation. Three forms of woodpeckers (sapsuckers; genus Sphyrapicus) in North America that are mostly geographically separated but hybridize with each other where they come into contact present a remarkable system for the study of hybridization. We provide the first comprehensive analysis of phenotypic and genetic variation across a hybrid zone between two of these forms, the red‐breasted Sphyrapicus ruber and yellow‐bellied S. varius sapsuckers. The objective was to infer whether selection maintains the differences between forms. Our analysis of eight morphometric and 20 plumage traits, and two molecular markers showed clear differences between the forms and roughly concordant clinal variation across a narrow hybrid zone. Thirty percent of sampled birds in the hybrid zone had mixed west/east genotypes at the genetic markers examined. The center of the genetic cline was located 20 km west of the crest of the Rocky Mountains. The width of the zone was 122 km, narrower than would be expected under neutral blending given reasonable estimates of the age of the zone and individual dispersal distances. Heterozygote deficit and cytonuclear disequilibrium at the centre of the hybrid zone suggested nonrandom mating or limited hybridization. Given these patterns and lack of evidence for habitat segregation we conclude that this hybrid zone is maintained by selection, most likely in the form of hybrid inferiority. This study provides an illustrative example of extensive hybridization between stable entities, providing additional evidence against the historical practice of treating hybridizing forms as members of the same species.     

The in vitro inhibition effect of 2 nm gold nanoparticles on non-enzymatic glycation of human serum albumin

The reaction of amino groups of protein and the carbonyl groups of reducing sugar molecules, non-enzymatically induce a series of chemical reactions that form a heterogeneous group of compounds known as advanced glycation end products (AGEs). The accumulation of AGEs is associated with various disease conditions that include complications in diabetes, Alzheimer's disease and aging. The current study monitored the extent of non-enzymatic glycation of human serum albumin (HSA) in order to estimate the formation of HSA related AGEs in the presence of 2 nm gold nanoparticles. The rate of glycation was evaluated using several analytical methods. Physiological concentrations of HSA and glyceraldehyde mixtures, incubated with various concentrations of negatively charged 2 nm gold nanoparticles, resulted in a lower reaction rate than mixtures without 2GNP. Moreover, increasing concentrations of gold nanoparticles exhibited a pronounced reduction in AGE formation. High performance liquid chromatography, UV-visible spectroscopy and circular dichroism analytical methods provide reliable techniques for evaluating AGE formation of HSA adducts.