Research into the Rhizobium/Leguminosae symbiosis in Latin America

More than 60 institutions and 100 researchers were involved in Rhizobium research in 1978 in Latin America. Half of these researchers were located in Argentina and Brazil. Research activity and the application of research findings vary widely among countries. Problems that plague research include 1) inadequate training of research personnel and insufficient attention paid to the Rhizobium/Legume symbiosis at agriculture schools; 2) poorly-established research priorities that do not sufficiently weigh the immediate needs for the farmers such as the identification of limiting environmental factors (e.g. nutritional deficiencies), techniques for smallscale inoculant production, and quality control of available inoculants; 3) isolation of the researchers and a lack of adequate library support; 4) poorly integrated research teams (e.g. in many institutes researchers are either microbiologists with no agricultural background or agronomists lacking microbiological training); and 5) insufficient dissemination of research findings. Problems with inoculant production and control include 1) a local dependence on national or imported inoculants rather than on locally-selected strains, 2) poor inoculant quality control which results in low inoculation success rates and subsequent discredit to the inoculation practice, and 3) high prices for inoculants. Extension problems include 1) lacking or deficient legume-promotion programs by government agencies, 2) poor contact between research and extension workers, and 3) administrators, leaders, extension workers and agronomists working in the field that lack adequate knowledge of the Rhizobium/Legume symbiosis. Immediate measures to foster extension and legume promotion programs and informal and/or official quality control are needed in Argentina, Uruguay, Brazil, Mexico, and probably Colombia. Countries where combined efforts should primarily be directed toward stimulating research and extension include Peru, Venezuela, Costa Rica, and Chile. In Ecuador, Paraguay, Bolivia, Nicaragua, Honduras, Guatemala, the Dominican Republic and Panama, priority should be given to research. Colombia should also be included in this group as national research institutions need to be strengthened. Table 2 lists these priorities more fully.     

Ultrasonography as a diagnostic tool for Neural Pain in Leprosy

Leprosy is still a prevalent disease in Brazil, representing 93% of all occurrences in the Americas. Leprosy neuropathy is one of the most worrying manifestations of the disease. Acute neuropathy usually occurs during reaction episodes and is called neuritis. Twenty-two leprosy patients were included in this study. These patients had neural pain associated with ulnar sensory neuropathy, with or without adjunct motor involvement. The neurological picture began within thirty days of the clinical evaluation. The patients underwent a nerve conduction study and the demyelinating findings confirmed the diagnosis of neuritis. Ultrasonographic study (US) of the ulnar nerve was performed in all patients by a radiologist who was blinded to the clinical or neurophysiological results. Morphological characteristics of the ulnar nerve were analyzed, such as echogenicity, fascicular pattern, transverse cross-sectional area (CSA), aspect of the epineurium, as well as their anatomical relationships. The volume of selected muscles referring to the ulnar nerve, as well as their echogenicity, was also examined. Based on this analysis, patients with increased ulnar nerve CSA associated with loss of fascicular pattern, epineurium hyperechogenicity and presence of power Doppler flow were classified as neuritis. Therefore, patients initially classified by the clinical-electrophysiological criteria were reclassified by the imaging criteria pre-established in this study as with and without neuritis. Loss of fascicular pattern and flow detection on power Doppler showed to be significant morphological features in the detection of neuritis. In 38.5% of patients without clinical or neurophysiological findings of neuritis, US identified power Doppler flow and loss of fascicular pattern. The US is a method of high resolution and portability, and its low cost means that it could be used as an auxiliary tool in the diagnosis of neuritis and its treatment, especially in basic health units.     

Adenylosuccinate Synthetase and Adenylosuccinate Lyase Deficiencies Trigger Growth and Infectivity Deficits in Leishmania donovani

Leishmania are auxotrophic for purines, and consequently purine acquisition from the host is a requisite nutritional function for the parasite. Both adenylosuccinate synthetase (ADSS) and adenylosuccinate lyase (ASL) have been identified as vital components of purine salvage in Leishmania donovani, and therefore Δadss and Δasl null mutants were constructed to test this hypothesis. Unlike wild type L. donovani, Δadss and Δasl parasites in culture exhibited a profoundly restricted growth phenotype in which the only permissive growth conditions were a 6-aminopurine source in the presence of 2′-deoxycoformycin, an inhibitor of adenine aminohydrolase activity. Although both knock-outs showed a diminished capacity to infect murine peritoneal macrophages, only the Δasl null mutant was profoundly incapacitated in its ability to infect mice. The enormous discrepancy in parasite loads observed in livers and spleens from mice infected with either Δadss or Δasl parasites can be explained by selective accumulation of adenylosuccinate in the Δasl knock-out and consequent starvation for guanylate nucleotides. Genetic complementation of a Δasl lesion in Escherichia coli implied that the L. donovani ASL could also recognize 5-aminoimidazole-(N-succinylocarboxamide) ribotide as a substrate, and purified recombinant ASL displayed an apparent K_m of ∼24 μm for adenylosuccinate. Unlike many components of the purine salvage pathway of L. donovani, both ASL and ADSS are cytosolic enzymes. Overall, these data underscore the paramount importance of ASL to purine salvage by both life cycle stages of L. donovani and authenticate ASL as a potential drug target in Leishmania.     

Liver-targeting of primaquine-(poly-γ-glutamic acid) and its degradation in rat hepatocytes

We have synthesized poly-γ-glutamic acid (PGA) modified with a synthetic trivalent glyco-ligand (TriGalNAc) for the hepatocyte asialoglycoprotein receptor (ASGP-R). We investigated in vivo distribution of unmodified PGA and TriGalNAc-modified PGA (TriGalNAc-PGA) in mice after intravenous injection. Most of unmodified PGA administered was transported to the bladder over 20–80 min, suggesting a rapid excretion of unmodified PGA into urine. In contrast, TriGalNAc-PGA was found exclusively in the liver over the same period of time. We further synthesized TriGalNAc-PGA–primaquine conjugate (TriGalNAc-PGA–PQ), and investigated binding, uptake, and catabolism of the conjugate by rat hepatocytes. Our studies indicated that approximately 250 ng per million cells of the conjugate bound to one million rat hepatocytes at 0 °C, and approximately 2 μg per million cells of the conjugate was taken up over 7 h incubation at 37 °C. Furthermore, our results suggested that TriGalNAc-PGA–PQ was almost completely degraded over 24 h, and small degradation products were secreted into cell culture medium.The results described in this report suggest that the TriGalNAc ligand can serve as an excellent targeting device for delivery of PGA-conjugates to the liver hepatocytes, and rat hepatocytes possess sufficient capacity to digest PGA even modified with other substituents.     

Dynamics of Cattle Production in Brazil

Movement of livestock production within a country or region has implications for genetics, adaptation, well-being, nutrition, and production logistics, particularly in continental-sized countries, such as Brazil. Cattle production in Brazil from 1977 to 2011 was spatialized, and the annual midpoint of production was calculated. Changes in the relative production and acceleration of production were calculated and spatialized using ARCGIS®. Cluster and canonical discriminant analyses were performed to further highlight differences between regions in terms of cattle production. The mean production point has moved from the Center of Minas Gerais State (in the southeast region) to the North of Goiás State (in the Midwest region). This reflects changes in environmental factors, such as pasture type, temperature and humidity. Acceleration in production in the northern region of Brazil has remained strong over the years. More recently, “traditional” cattle-rearing regions, such as the south and southeast, showed a reduction in growth rates as well as a reduction in herd size or internal migration over the period studied. These maps showed that this movement tends to be gradual, with few regions showing high acceleration or deceleration rates.     

Follow-up assessment of patients with Pure Neural Leprosy in a reference center in Rio de Janeiro—Brazil

IntroductionPure Neural Leprosy (PNL) is a rare clinical form of leprosy in which patients do not present with the classical skin lesions but have a high burden of the disability associated with the disease. Clinical characteristics and follow up of patients in PNL are still poorly described in the literature.ObjectiveThis paper aims to describe the clinical, electrophysiological and histopathological characteristics of PNL patients, as well as their evolution after multidrug therapy (MDT).MethodsFifty-two PNL patients were selected. Clinical, nerve conduction studies (NCS), histopathological and anti-PGL-1serology were evaluated. Patients were also assessed monthly during the MDT. At the end of the MDT, all of the patients had a new neurological examination and 44 were submitted to another NCS.ResultsParesthesia was the complaint most frequently reported by patients, and in the neurological examination the most common pattern observed was impairment in sensory and motor examination and a mononeuropathy multiplex. Painful nerve enlargement, a classical symptom of leprosy neuropathy, was observed in a minority of patients and in the motor NCS axonal injuries, alone or in combination with demyelinating features, were the most commonly observed. 88% of the patients did not present any leprosy reaction during MDT. There was no statistically significant difference between the neurological examinations, nor the NCS pattern, performed before and after the MDT.DiscussionThe classical hallmarks of leprosy neuropathy are not always present in PNL making the diagnosis even more challenging. Nerve biopsy is an important tool for PNL diagnosis as it may guide therapeutic decisions. This paper highlights unique characteristics of PNL in the spectrum of leprosy in an attempt to facilitate the diagnosis and management of these patients.     

Comparative genomics of grasses tolerant to aluminum

The family Poaceae includes over 10,000 species, among which are the most economically important cereals: maize, sorghum, rice, wheat, rye, barley, and oat. These cereals are very important components of human and animal food. Although divergence of the members of this family occurred about 40 million years ago, comparative genome analyses demonstrated that gene orders among species of this family remain largely conserved, which can be very useful for understanding their roles and evolution. Even with an intricate evolutionary history in which chromosome fragments, losses and duplications have to be considered at the ploidy level, grasses present a genetic model system for comparative genomics. The availability of mapped molecular markers, rice genome sequences and BAC and EST libraries from several grass species, such as rice, wheat, sorghum, and maize, facilitates biology and phylogeny studies of this group. The value of using information from different species in modern plant genetics is unquestionable, especially in the study of traits such as tolerance to aluminum in soils, which affects plant growth and development. Comparative genomic approaches to aluminum tolerance can identify genomic regions and genes responsible for aluminum tolerance in grasses.     

Saikosaponin-d,a novel SERCA inhibitor,induces autophagic cell death in apoptosis-defective cells

Autophagy is an important cellular process that controls cells in a normal homeostatic state by recycling nutrients to maintain cellular energy levels for cell survival via the turnover of proteins and damaged organelles. However, persistent activation of autophagy can lead to excessive depletion of cellular organelles and essential proteins, leading to caspase-independent autophagic cell death. As such, inducing cell death through this autophagic mechanism could be an alternative approach to the treatment of cancers. Recently, we have identified a novel autophagic inducer, saikosaponin-d (Ssd), from a medicinal plant that induces autophagy in various types of cancer cells through the formation of autophagosomes as measured by GFP-LC3 puncta formation. By computational virtual docking analysis, biochemical assays and advanced live-cell imaging techniques, Ssd was shown to increase cytosolic calcium level via direct inhibition of sarcoplasmic/endoplasmic reticulum Ca²⁺ ATPase pump, leading to autophagy induction through the activation of the Ca²⁺/calmodulin-dependent kinase kinase–AMP-activated protein kinase–mammalian target of rapamycin pathway. In addition, Ssd treatment causes the disruption of calcium homeostasis, which induces endoplasmic reticulum stress as well as the unfolded protein responses pathway. Ssd also proved to be a potent cytotoxic agent in apoptosis-defective or apoptosis-resistant mouse embryonic fibroblast cells, which either lack caspases 3, 7 or 8 or had the Bax-Bak double knockout. These results provide a detailed understanding of the mechanism of action of Ssd, as a novel autophagic inducer, which has the potential of being developed into an anti-cancer agent for targeting apoptosis-resistant cancer cells.     

The cystathionine‐β‐synthase domains on the guanosine 5′’‐monophosphate reductase and inosine 5′‐monophosphate dehydrogenase enzymes from Leishmania regulate enzymatic activity in response to guanylate and adenylate nucleotide levels

The Leishmania guanosine 5′‐monophosphate reductase (GMPR) and inosine 5′‐monophosphate dehydrogenase (IMPDH) are purine metabolic enzymes that function maintaining the cellular adenylate and guanylate nucleotide. Interestingly, both enzymes contain a cystathionine‐β‐synthase domain (CBS). To investigate this metabolic regulation, the Leishmania GMPR was cloned and shown to be sufficient to complement the guaC (GMPR), but not the guaB (IMPDH), mutation in Escherichia coli. Kinetic studies confirmed that the Leishmania GMPR catalyzed a strict NADPH‐dependent reductive deamination of GMP to produce IMP. Addition of GTP or high levels of GMP induced a marked increase in activity without altering the K_m values for the substrates. In contrast, the binding of ATP decreased the GMPR activity and increased the GMP K_m value 10‐fold. These kinetic changes were correlated with changes in the GMPR quaternary structure, induced by the binding of GMP, GTP, or ATP to the GMPR CBS domain. The capacity of these CBS domains to mediate the catalytic activity of the IMPDH and GMPR provides a regulatory mechanism for balancing the intracellular adenylate and guanylate pools.