two-algorithms-for-high-throughput-multi-parametric-quantification

Two Algorithms for High-throughput and Multi-parametric Quantification of <em>Drosophila</em> Neuromuscular Junction Morphology

Synaptic morphology is tightly related to synaptic efficacy, and in many cases morphological synapse defects ultimately lead to synaptic malfunction. The ...

Multidrug-resistant tuberculosis (MDR-TB) is threatening control of TB in many parts of the world. As a result of limited treatment options, patients have a poor prognosis and low chances of cure. This situation can be exacerbated by HIV epidemics. In some cases, the risk exists of a real shift from susceptible to resistant strains. Despite its relevance, currently there are more contradictions and confusion surrounding MDR-TB than hard evidence. No randomized controlled trials have been performed and published evidence is limited. Rather than just the selection of expensive drugs, MDR-TB management requires well-structured programmes with a comprehensive approach, which involve the actions of a wide range of participants. Even with current investments in research and development, new drugs and vaccines will take many years to be applied in low and middle income countries. The most successful results will depend on the optimization of existing tools. The majority of the patients, even those with extensive patterns of bacilli resistance, have a possibility of cure if current clinical knowledge and effective logistics are applied. This paper is a critical review of current best practice regarding the diagnosis and treatment of MDR-TB.

Management of multidrug-resistant tuberculosis: an update.

Despite killing nearly 2 million people every year, TB is arguably the most neglected disease in terms of the funding and research it receives. In many ways, multidrug-resistant TB is a result of this disregard. In high-burden countries, not many improvements have been implemented in the diagnosis and treatment tools of TB since the 1960s. Following this period, fluoroquinolones, developed for other infections, are the only highly active new drugs against TB. Multidrug- and extensively drug-resistant TB is booming worldwide as a result of insufficient control measures. Furthermore, the prevalence of this disease is substantially facilitated by the HIV epidemic. After a deadly and well-reported extensively drug-resistant TB outbreak occurred in HIV-infected patients in South Africa, the threat of an untreatable TB epidemic is receiving increased attention internationally. Nevertheless, drug-resistant management has lacked research and funding over several decades and we are now faced with many controversial issues and little research-based evidence. There are no clinical trials comparing different treatments, and current management is based on personal experiences and agreement between experts. The major challenge for the next few years is to improve the evidence base in order to develop more rational recommendations that adequately address the current problem and avoid making a bad situation worse.

MDR-/XDR-TB management: what it was, current standards and what is ahead.

It is becoming increasingly clear that the activation of specific terminal differentiation genes during neural development is critically dependent upon the establishment of unique combinatorial transcription factor codes within distinct neural cell subtypes. However, it is still unclear to which extent these codes are shared by lineage-unrelated neurons expressing the same terminal differentiation genes. Additionally, it is not known if the activation of a specific terminal differentiation gene is restricted to cells born at a particular developmental time point. Here, we utilize the terminal differentiation gene FMRFa which is expressed by the Ap4 and SE2 neurons in the Drosophila ventral nerve cord, to explore these issues in depth. We find that the Ap4 and SE2 neurons are generated by different neural progenitors and use different combinatorial codes to activate FMRFa expression. Additionally, we find that the Ap4 and SE2 neurons are generated in different temporal gene expression windows. Extending the investigation to include a second Drosophila terminal differentiation gene, Leucokinin, we find similar results, suggesting that neurons generated by different progenitors might commonly use different transcription factor codes to activate the same terminal differentiation gene. Furthermore, these results imply that the activation of a particular terminal differentiation gene in temporally unrestricted.

Lineage-unrelated neurons generated in different temporal windows and expressing different combinatorial codes can converge in the activation of the same terminal differentiation gene.

Multidrug-resistant tuberculosis (MDR-TB), defined as being resistant to at least rifampicin and isoniazid, has an increasing burden and threatens TB control. Diagnosis is limited and usually delayed while treatment is long lasting, toxic and poorly effective. MDR-TB management in scarce-resource settings is demanding however it is feasible and extremely necessary. In these settings, cure rates do not usually exceed 60-70% and MDR-TB management is novel for many TB programs. In this challenging scenario, both clinical and programmatic errors are likely to occur. The majority of these errors may be prevented or alleviated with appropriate and timely training in addition to uninterrupted procurement of high-quality drugs, updated national guidelines and laws and an overall improvement in management capacities. While new tools for diagnosis and shorter and less toxic treatment are not available in developing countries, MDR-TB management will remain complex in scarce resource settings. Focusing special attention on the common errors in diagnosis, regimen design and especially treatment delivery may benefit patients and programs with current outdated tools. The present article is a compilation of typical errors repeatedly observed by the authors in a wide range of countries during technical assistant missions and trainings.

Common errors in multidrug-resistant tuberculosis management.

The emergence of multidrug-resistant (MDR) and extensively drug-resistant (XDR) tuberculosis (TB) substantially challenges TB control, especially in the European Region of the World Health Organization, where the highest prevalence of MDR/XDR cases is reported. The current management of patients with MDR/XDR-TB is extremely complex for medical, social and public health systems. The treatment with currently available anti-TB therapies to achieve relapse-free cure is long and undermined by a high frequency of adverse drug events, suboptimal treatment adherence, high costs and low treatment success rates. Availability of optimal management for patients with MDR/XDR-TB is limited even in the European Region. In the absence of a preventive vaccine, more effective diagnostic tools and novel therapeutic interventions the control of MDR/XDR-TB will be extremely difficult. Despite recent scientific advances in MDR/XDR-TB care, decisions for the management of patients with MDR/XDR-TB and their contacts often rely on expert opinions, rather than on clinical evidence. This document summarises the current knowledge on the prevention, diagnosis and treatment of adults and children with MDR/XDR-TB and their contacts, and provides expert consensus recommendations on questions where scientific evidence is still lacking.

Management of patients with multidrug-resistant/extensively drug-resistant tuberculosis in Europe: a TBNET consensus statement.

Studies in the Drosophila embryonic NB4-2 lineage have suggested that the transcription factor Klumpfuss (Klu) functions within embryonic neuroblast lineages to differentiate between the identities of two adjacent ganglion mother cells (GMCs). However, because of the limited lineage markers available, these observations have been made only for the NB4-2 lineage. Recent findings have placed this transcription factor in the vanguard of Drosophila neural stem cell biology by demonstrating that Klu is necessary for larval neuroblast growth and self-renewal. Here, we have studied the role of klu in an incipient model in order to address basic mechanisms of neural specification: the Va system. None of the previously reported roles of Klu satisfactorily explain our observations. Unexpectedly, in this lineage, klu is necessary for differentiating between the fates of the two neurons born from a unique GMC; klu mutants produce two B-type cells, rather than one B-type (Notch-OFF) and one A-type (Notch-ON) cell. Additionally, our results demonstrate that Klu operates in the GMC and/or in the newly born neuron, but not in the neuroblast. Unlike in larval neuroblasts in which Klu is an executor of Notch signaling, we have found that Klu does not lie downstream of the Notch pathway in this cell division context.

A new role of Klumpfuss in establishing cell fate during the GMC asymmetric cell division.

The changing landscape in drug resistant-tuberculosis: an analysis of recent advances.

The morphology of synapses is of central interest in neuroscience because of the intimate relation with synaptic efficacy. Two decades of gene manipulation studies in different animal models have revealed a repertoire of molecules that contribute to synapse development. However, since such studies often assessed only one, or at best a few, morphological features at a given synapse, it remained unaddressed how different structural aspects relate to one another. Furthermore, such focused and sometimes only qualitative approaches likely left many of the more subtle players unnoticed. Here, we present the image analysis algorithm 'Drosophila_NMJ_Morphometrics', available as a Fiji-compatible macro, for quantitative, accurate and objective synapse morphometry of the Drosophila larval neuromuscular junction (NMJ), a well-established glutamatergic model synapse. We developed this methodology for semi-automated multiparametric analyses of NMJ terminals immunolabeled for the commonly used markers Dlg1 and Brp and showed that it also works for Hrp, Csp and Syt. We demonstrate that gender, genetic background and identity of abdominal body segment consistently and significantly contribute to variability in our data, suggesting that controlling for these parameters is important to minimize variability in quantitative analyses. Correlation and principal component analyses (PCA) were performed to investigate which morphometric parameters are inter-dependent and which ones are regulated rather independently. Based on nine acquired parameters, we identified five morphometric groups: NMJ size, geometry, muscle size, number of NMJ islands and number of active zones. Based on our finding that the parameters of the first two principal components hardly correlated with each other, we suggest that different molecular processes underlie these two morphometric groups. Our study sets the stage for systems morphometry approaches at the well-studied Drosophila NMJ.

Article	Year
Management of multidrug-resistant tuberculosis: an update. Therapeutic advances in respiratory disease\| PubMed ID: 20388724	2010
MDR-/XDR-TB management: what it was, current standards and what is ahead. Expert review of respiratory medicine\| PubMed ID: 20477307	2009
Lineage-unrelated neurons generated in different temporal windows and expressing different combinatorial codes can converge in the activation of the same terminal differentiation gene. Mechanisms of development\| PubMed ID: 20732418	2010
Common errors in multidrug-resistant tuberculosis management. Expert review of respiratory medicine\| PubMed ID: 24329041	2014
Management of patients with multidrug-resistant/extensively drug-resistant tuberculosis in Europe: a TBNET consensus statement. The European respiratory journal\| PubMed ID: 24659544	2014
A new role of Klumpfuss in establishing cell fate during the GMC asymmetric cell division. Cell and tissue research\| PubMed ID: 25129108	2014
The changing landscape in drug resistant-tuberculosis: an analysis of recent advances. Expert review of respiratory medicine\| PubMed ID: 26954921	2016
A New Fiji-Based Algorithm That Systematically Quantifies Nine Synaptic Parameters Provides Insights into Drosophila NMJ Morphometry. PLoS computational biology\| PubMed ID: 26998933	2016

Ignacio Monedero

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