RESUMO
Projection neurons are the commonest neuronal type in the mammalian forebrain and their individual characterization is a crucial step to understand how neural circuitry operates. These cells have an axon whose arborizations extend over long distances, branching in complex patterns and/or in multiple brain regions. Axon length is a principal estimate of the functional impact of the neuron, as it directly correlates with the number of synapses formed by the axon in its target regions; however, its measurement by direct 3D axonal tracing is a slow and labor-intensive method. On the contrary, axon length estimations have been recently proposed as an effective and accessible alternative, allowing a fast approach to the functional significance of the single neuron. Here, we analyze the accuracy and efficiency of the most used length estimation tools-design-based stereology by virtual planes or spheres, and mathematical correction of the 2D projected-axon length-in contrast with direct measurement, to quantify individual axon length. To this end, we computationally simulated each tool, applied them over a dataset of 951 3D-reconstructed axons (from NeuroMorpho.org), and compared the generated length values with their 3D reconstruction counterparts. The evaluated reliability of each axon length estimation method was then balanced with the required human effort, experience and know-how, and economic affordability. Subsequently, computational results were contrasted with measurements performed on actual brain tissue sections. We show that the plane-based stereological method balances acceptable errors (~5%) with robustness to biases, whereas the projection-based method, despite its accuracy, is prone to inherent biases when implemented in the laboratory. This work, therefore, aims to provide a constructive benchmark to help guide the selection of the most efficient method for measuring specific axonal morphologies according to the particular circumstances of the conducted research.
Assuntos
Axônios/fisiologia , Biologia Computacional/métodos , Imageamento Tridimensional/métodos , Neurônios/citologia , Animais , Benchmarking , Bases de Dados Factuais , Camundongos , TomografiaRESUMO
Haemosporidians are a diverse group of vector-borne parasitic protozoa that includes the agents of human malaria; however, most of the described species are found in birds and reptiles. Although our understanding of these parasites' diversity has expanded by analyses of their mitochondrial genes, there is limited information on these genes' evolutionary rates. Here, 114 mitochondrial genomes (mtDNA) were studied from species belonging to four genera: Leucocytozoon, Haemoproteus, Hepatocystis, and Plasmodium. Contrary to previous assertions, the mtDNA is phylogenetically informative. The inferred phylogeny showed that, like the genus Plasmodium, the Leucocytozoon and Haemoproteus genera are not monophyletic groups. Although sensitive to the assumptions of the molecular dating method used, the estimated times indicate that the diversification of the avian haemosporidian subgenera/genera took place after the Cretaceous-Paleogene boundary following the radiation of modern birds. Furthermore, parasite clade differences in mtDNA substitution rates and strength of negative selection were detected. These differences may affect the biological interpretation of mtDNA gene lineages used as a proxy to species in ecological and parasitological investigations. Given that the mitochondria are critically important in the parasite life cycle stages that take place in the vector and that the transmission of parasites belonging to particular clades has been linked to specific insect families/subfamilies, this study suggests that differences in vectors have affected the mode of evolution of haemosporidian mtDNA genes. The observed patterns also suggest that the radiation of haemosporidian parasites may be the result of community-level evolutionary processes between their vertebrate and invertebrate hosts.
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Evolução Biológica , Genoma Mitocondrial , Genoma de Protozoário , Haemosporida/genética , Seleção GenéticaRESUMO
Helicobacter presence and viability in waters is not well characterized. The identification of natural reservoirs and infection sources may provide novel insights into its waterborne transmission. The goal of this study was to investigated the occurrence of Helicobacter spp. in natural freshwaters from Roraima Tepui, a little studied and unique ecosystem of the Guayana Shield. Freshwaters collected from two localities at Roraima Tepui were cultured in HP selective broth and agar for Helicobacter pylori and analysed by fluorescent in situ hybridization (FISH), specific PCR assays, 16S rRNA gene sequencing and phylogenetic analysis. The presence of other bacteria in freshwater enrichments was determined using clone library sequencing of the 16S rRNA gene and phylogenetic inferences. Helicobacter spp. were detected by semi-nested PCR and FISH in freshwater enrichments from both sites. Coccoid viable but nonculturable (VBNC) cells were evidenced using 16S rRNA gene Helicobacter species and H. pylori-specific probes. Partial 16S rRNA gene sequences of two HP enrichments showed high similarity to H. pylori and Helicobacter nemestrinae (99-100 %). Other bacteria such as Serratia, Aquitalea, Chromobacterium, Mycobacterium, Acinetobacter, Curvibacter and Dysgonomonas were also detected using complete 16S rRNA gene sequences, with Serratia, Aquitalea and Chromobacterium the most common genera (40.9, 18.2 and 15.2 %, respectively). This is the first time that Helicobacter spp. have been reported in freshwaters of a tepui ecosystem. Our results contribute to the current knowledge of these bacteria in the aquatic environment and expand their known/potential sites outside the human host.
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Água Doce/microbiologia , Helicobacter/classificação , Helicobacter/isolamento & purificação , Sequência de Bases , DNA Bacteriano/genética , Ecossistema , Helicobacter/genética , Infecções por Helicobacter/microbiologia , Humanos , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , América do Sul , Recursos HídricosRESUMO
Proteus mirabilis is a human pathogen able to form biofilms on the surface of urinary catheters. Little is known about P. mirabilis biofilms on natural or industrial surfaces and the potential consequences for these settings. The main aim of this work was to assess and compare the adhesion and biofilm formation of P. mirabilis strains from different origins on chitin and stainless steel surfaces within 4 to 96 h. Using environmental scanning electron microscopy, the biofilms of a clinical strain grown on chitin at 4 h showed greater adhesion, aggregation, thickness, and extracellular matrix production than those grown on stainless steel, whereas biofilms of an environmental strain had less aggregation on both surfaces. Biofilms of both P. mirabilis strains developed different structures on chitin, such as pillars, mushrooms, channels, and crystalline-like precipitates between 24 and 96 h, in contrast with flat-layer biofilms produced on stainless steel. Significant differences (p < 0.05) were found in the frequency of pillars and channels. Images of transmission electron microscopy demonstrated abundant fimbriae in 100 % of cells from both strains, which could be related to surface adherence and biofilm formation. This represents the first study of P. mirabilis showing adhesion, biofilm formation, and development of different structures on surfaces found outside the human host.
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Two main neuronal pathways connect facial whiskers to the somatosensory cortex in rodents: (i) the lemniscal pathway, which originates in the brainstem principal trigeminal nucleus and is relayed in the ventroposterior thalamic nucleus and (ii) the paralemniscal pathway, originating in the spinal trigeminal nucleus and relayed in the posterior thalamic nucleus. While lemniscal neurons are readily activated by whisker contacts, the contribution of paralemniscal neurons to perception is less clear. Here, we functionally investigated these pathways by manipulating input from the whisker pad in freely moving mice. We report that while lemniscal neurons readily respond to neonatal infraorbital nerve sectioning or whisker contacts in vivo, paralemniscal neurons do not detectably respond to these environmental changes. However, the paralemniscal pathway is specifically activated upon noxious stimulation of the whisker pad. These findings reveal a nociceptive function for paralemniscal neurons in vivo that may critically inform context-specific behaviour during environmental exploration.
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Nociceptividade/fisiologia , Núcleo Espinal do Trigêmeo/metabolismo , Animais , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas c-fos/metabolismo , Núcleo Espinal do Trigêmeo/fisiologia , Vibrissas/inervaçãoRESUMO
Current mesoscale connectivity atlases provide limited information about the organization of thalamocortical projections in the mouse brain. Labeling the projections of spatially restricted neuron populations in thalamus can provide a functionally relevant level of connectomic analysis, but these need to be integrated within the same common reference space. Here, we present a pipeline for the segmentation, registration, integration and analysis of multiple tract-tracing experiments. The key difference with other workflows is that the data is transformed to fit the reference template. As a test-case, we investigated the axonal projections and intranuclear arrangement of seven neuronal populations of the ventral posteromedial nucleus of the thalamus (VPM), which we labeled with an anterograde tracer. Their soma positions corresponded, from dorsal to ventral, to cortical representations of the whiskers, nose and mouth. They strongly targeted layer 4, with the majority exclusively targeting one cortical area and the ones in ventrolateral VPM branching to multiple somatosensory areas. We found that our experiments were more topographically precise than similar experiments from the Allen Institute and projections to the primary somatosensory area were in agreement with single-neuron morphological reconstructions from publicly available databases. This pilot study sets the basis for a shared virtual connectivity atlas that could be enriched with additional data for studying the topographical organization of different thalamic nuclei. The pipeline is accessible with only minimal programming skills via a Jupyter Notebook, and offers multiple visualization tools such as cortical flatmaps, subcortical plots and 3D renderings and can be used with custom anatomical delineations.
Assuntos
Neurônios , Tálamo , Camundongos , Animais , Vias Neurais/fisiologia , Projetos Piloto , Tálamo/anatomia & histologia , Neurônios/fisiologia , AxôniosRESUMO
The nuclei of the human amygdaloid complex can be distinguished from each other on the basis of their cytoarchitecture, chemistry and connections, all of which process the information needed for the different functions (ranging from attention to memory and emotion) of the amygdala. This complex receives dopaminergic input that exerts modulatory effects over its intrinsic network and is critical for reward-related learning and fear conditioning. To determine the specific distribution of the dopaminergic input through the different nuclei and nuclear subdivisions of this structure we used stereological tools to quantify the fibers containing the dopamine transporter (used to signal the dopaminergic phenotype) in post-mortem samples from control individuals. Dopaminergic axons targeted every nucleus of the amygdaloid complex, and the density of dopamine transporter-containing axons varied considerably among its nuclear groups. The central group showed the greatest density of dopamine transporter-positive fibers, more than double the density of the basolateral group, the second most densely innervated structure. The dopamine transporter-positive innervation is very scant in the corticomedial group. The density of dopamine transporter-positive fibers did not vary among the nuclei of the basolateral group - i.e. basal, lateral and accessory basal nuclei - although there were significant density gradients among the subdivisions of these nuclei. These detailed quantitative data on dopamine transporter-positive innervation in the human amygdaloid complex can offer a useful reference in future studies aimed at analysing putative dysfunctions of this system in diseases involving brain dopamine, such as certain anxiety disorders, Parkinson's disease and schizophrenia.
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Tonsila do Cerebelo/metabolismo , Axônios/metabolismo , Proteínas da Membrana Plasmática de Transporte de Dopamina/metabolismo , Idoso , Tonsila do Cerebelo/citologia , Proteínas da Membrana Plasmática de Transporte de Dopamina/genética , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Especificidade de ÓrgãosRESUMO
Background: The Horned Screamer (Anhima cornuta) is an herbivorous bird that inhabits wetlands of the South American tropical region. We hypothesize that due to its herbivorous niche, its digestive tract compartments may have bacteria specialized in fermenting complex plant carbohydrates. To test this hypothesis, we compared the bacterial communities along the gastrointestinal tract (GIT) of a Horned Screamer captured in Venezuela. Methods: Samples were taken from tissues and content of the proventriculus and the small intestine (considered for this study as upper GIT), and the large intestine and cecum (lower GIT). The bacterial community was characterized by sequencing the V4 region of the 16S rRNA gene. Bioinformatic analysis was performed using QIIME, QIITA and Microbiome Analyst. The association between microbial taxonomy and function was analyzed using their Greengenes OTU IDs and a custom KEGG BRITE hierarchical tree and visualized with BURRITO. Results: The Screamer's gastrointestinal microbiota was composed by seven phyla being Firmicutes and Bacteroidetes the most predominant. The dominant taxa in the upper GIT were Helicobacter, Vibrio, Enterobacter, Acinetobacter and Staphylococcus. The dominant taxa in the lower GIT were Oribacterium, Blautia, Roseburia, Ruminococcus, Desulfovibrio, Intestinimonas, Marvinbryantia and Parabacteroides. Complete degradation of cellulose to the end-products acetate, propanoate, butanoate and acetoacetate was found in the upper and lower GIT without significant differences. Conclusion: Our study confirmed changes in bacterial community composition throughout the GIT of the Horned Screamer primarily associated with the production of metabolic end-products of carbohydrate digestion essential for the fermentation of the herbivorous diet.
Assuntos
Anseriformes , Microbioma Gastrointestinal , Animais , RNA Ribossômico 16S/genética , Trato Gastrointestinal/microbiologia , Bactérias , Microbioma Gastrointestinal/genética , Bacteroidetes/genética , Aves/genética , Anseriformes/genéticaRESUMO
Characterizing the connectomic and morphological diversity of thalamic neurons is key for better understanding how the thalamus relays sensory inputs to the cortex. The recent public release of complete single-neuron morphological reconstructions enables the analysis of previously inaccessible connectivity patterns from individual neurons. Here we focus on the Ventral Posteromedial (VPM) nucleus and characterize the full diversity of 257 VPM neurons, obtained by combining data from the MouseLight and Braintell projects. Neurons were clustered according to their most dominantly targeted cortical area and further subdivided by their jointly targeted areas. We obtained a 2D embedding of morphological diversity using the dissimilarity between all pairs of axonal trees. The curved shape of the embedding allowed us to characterize neurons by a 1-dimensional coordinate. The coordinate values were aligned both with the progression of soma position along the dorsal-ventral and lateral-medial axes and with that of axonal terminals along the posterior-anterior and medial-lateral axes, as well as with an increase in the number of branching points, distance from soma and branching width. Taken together, we have developed a novel workflow for linking three challenging aspects of connectomics, namely the topography, higher order connectivity patterns and morphological diversity, with VPM as a test-case. The workflow is linked to a unified access portal that contains the morphologies and integrated with 2D cortical flatmap and subcortical visualization tools. The workflow and resulting processed data have been made available in Python, and can thus be used for modeling and experimentally validating new hypotheses on thalamocortical connectivity.
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Thalamoreticular circuitry plays a key role in arousal, attention, cognition, and sleep spindles, and is linked to several brain disorders. A detailed computational model of mouse somatosensory thalamus and thalamic reticular nucleus has been developed to capture the properties of over 14,000 neurons connected by 6 million synapses. The model recreates the biological connectivity of these neurons, and simulations of the model reproduce multiple experimental findings in different brain states. The model shows that inhibitory rebound produces frequency-selective enhancement of thalamic responses during wakefulness. We find that thalamic interactions are responsible for the characteristic waxing and waning of spindle oscillations. In addition, we find that changes in thalamic excitability control spindle frequency and their incidence. The model is made openly available to provide a new tool for studying the function and dysfunction of the thalamoreticular circuitry in various brain states.
Assuntos
Tálamo , Vigília , Camundongos , Animais , Tálamo/fisiologia , Sono/fisiologia , Núcleos Talâmicos/fisiologia , Percepção , Córtex Cerebral/fisiologiaRESUMO
The primate amygdaloid complex (AC) contains projection neurons as well as subsets of interneurons (IN), many of which express calcium-binding proteins, that through their local circuits control the activity of the projection neurons. The inhibitory parvalbumin (PV) and calretinin (CR)-positive (+) AC IN have a crucial role in the appearance of synchronized oscillations in local ensembles of projection neurons that mediate the consolidation and recall of fear memories. The GABAergic transmission of these subsets of IN is modulated by dopamine. To expand the knowledge regarding the cellular composition and distribution of IN in the human AC, we focused on two non-overlapping populations: the PV+ and CR+. We have analyzed the distribution of these IN throughout the AC from subjects without any neurological or psychiatric disorders and estimated their absolute number and density using stereological methods. We have also provided percentages of the IN with respect to the total AC neurons. The CRâ¯+â¯IN were distributed throughout the AC, whereas the PV+ were only present in the basolateral nuclear group. The quantity of CRâ¯+â¯IN was four times higher than that of PV+ and the percentages varied from less than 1% for PVâ¯+â¯IN to 6-20% for CR+. The differences in quantity and distribution of CR+ and PVâ¯+â¯IN could be related to their differential inhibitory properties and to the intrinsic and extrinsic connections of every amygdaloid region.
Assuntos
Interneurônios , Parvalbuminas , Tonsila do Cerebelo/metabolismo , Calbindina 2/metabolismo , Humanos , Interneurônios/metabolismo , Neurônios/metabolismo , Parvalbuminas/metabolismo , Proteína G de Ligação ao Cálcio S100/metabolismoRESUMO
The molecular mechanism of Helicobacter pylori resistance to tetracycline involves mutations in the primary binding site of the ribosome. A resistance or reduced susceptibility to tetracycline could be the result of single, double or triple mutations in the 16S rRNA gene of H. pylori. We investigated if the genotype was correlated to tetracycline resistance as determined phenotypically in vitro for 96 H. pylori isolates in the gastroesophageal mucosa of Venezuelan individual hosts. E-test for antimicrobial susceptibility test and real-time PCR for the detection of 16S rRNA gene mutations were performed in 96 H. pylori isolates (48 obtained from antrum, and 48 from oesophagus) from eight dyspeptic patients. In the gastric mucosa, 38 isolates were identified sensitive and 10 resistant to tetracycline by E-test, whereas 44 sensitive and 4 resistant isolates were found in the oesophagus. Real-time PCR detection of the 16S rRNA gene exhibited mutants with a single base-pair substitution (AGA926GGA) in six antrum isolates and seven oesophagus isolates, whereas only three harboured a low level of tetracycline resistance in vitro. Our results indicate that real-time PCR detection of 16S rRNA is a reliable method to classify among tetracycline-resistant genotypes and useful in patients who have experienced a first-line treatment failure with triple therapy.
Assuntos
Infecções por Helicobacter/microbiologia , Helicobacter pylori/efeitos dos fármacos , Helicobacter pylori/genética , RNA Ribossômico 16S/genética , Resistência a Tetraciclina/genética , Tetraciclina/farmacologia , Antibacterianos/farmacologia , Mucosa Esofágica/microbiologia , Mucosa Gástrica/microbiologia , Genótipo , Humanos , Testes de Sensibilidade Microbiana , Mutação , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Rodents extract information about nearby objects from the movement of their whiskers through dynamic computations that are carried out by a network of forebrain structures that includes the thalamus and the primary sensory (S1BF) and motor (M1wk) whisker cortices. The posterior nucleus (Po), a higher order thalamic nucleus, is a key hub of this network, receiving cortical and brainstem sensory inputs and innervating both motor and sensory whisker-related cortical areas. In a recent study in rats, we showed that Po inputs differently impact sensory processing in S1BF and M1wk. Here, in C57BL/6 mice, we measured Po synaptic bouton layer distribution and size, compared cortical unit response latencies to "in vivo" Po activation, and pharmacologically examined the glutamatergic receptor mechanisms involved. We found that, in S1BF, a large majority (56%) of Po axon varicosities are located in layer (L)5a and only 12% in L2-L4, whereas in M1wk this proportion is inverted to 18% and 55%, respectively. Light and electron microscopic measurements showed that Po synaptic boutons in M1wk layers 3-4 are significantly larger (~ 50%) than those in S1BF L5a. Electrical Po stimulation elicits different area-specific response patterns. In S1BF, responses show weak or no facilitation, and involve both ionotropic and metabotropic glutamate receptors, whereas in M1wk, unit responses exhibit facilitation to repetitive stimulation and involve ionotropic NMDA glutamate receptors. Because of the different laminar distribution of axon terminals, synaptic bouton size and receptor mechanisms, the impact of Po signals on M1wk and S1BF, although simultaneous, is likely to be markedly different.
Assuntos
Axônios/fisiologia , Axônios/ultraestrutura , Córtex Motor/fisiologia , Núcleos Posteriores do Tálamo/fisiologia , Córtex Somatossensorial/fisiologia , Sinapses/fisiologia , Sinapses/ultraestrutura , Animais , Estimulação Elétrica , Masculino , Camundongos Endogâmicos C57BL , Córtex Motor/ultraestrutura , Vias Neurais/fisiologia , Vias Neurais/ultraestrutura , Núcleos Posteriores do Tálamo/ultraestrutura , Receptores de Glutamato Metabotrópico/fisiologia , Receptores de N-Metil-D-Aspartato/fisiologia , Córtex Somatossensorial/ultraestrutura , Vibrissas/fisiologiaRESUMO
The amygdaloid complex (AC) is a heterogeneous aggregate of nuclei located in the rostromedial region of the temporal lobe. In addition to being partly connected among themselves, the AC nuclei are strongly interconnected with the cerebral cortex, striatum, basal forebrain, hypothalamus and brainstem. Animal and human functional studies have established that the AC is a central hub of the neuronal networks supporting emotional responsivity, particularly its negative/aversive components. Dysfunction of AC circuits in humans has been implicated in anxiety, depression, schizophrenia and bipolar disorder. The small New-World marmoset monkey (Callithrix jacchus) has recently become a key model for neuroscience research. However, the nuclear and fiber tract organization of marmoset AC has not been examined in detail. Thus, the extent to which it can be compared to the AC of Old-World (human and macaque) primates is yet unclear. Here, using Nissl and acetylcholinesterase (AChE) histochemical stains as a reference, we analyzed the cytoarchitecture and nuclear parcellation of the marmoset AC. In addition, given the increasing relevance of tractographic localization for high-resolution in vivo imaging studies in non-human primates, we also identified the myelin fiber tracts present within and around the AC as revealed by the Gallyas method. The present study provides a detailed atlas of marmoset AC. Moreover, it reveals that, despite phylogenetic distance and brain size differences, every nucleus and myelinated axon bundle described in human and macaque studies can be confidently recognized in marmosets.
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The evolution of flight in birds involves (i) decoupling of the primitive mode of quadrupedal locomotor coordination, with a new synchronized flapping motion of the wings while conserving alternating leg movements, and (ii) reduction of wing digits and loss of functional claws. Our observations show that hoatzin nestlings move with alternated walking coordination of the four limbs using the mobile claws on their wings to anchor themselves to the substrate. When swimming, hoatzin nestlings use a coordinated motion of the four limbs involving synchronous or alternated movements of the wings, indicating a versatile motor pattern. Last, the proportions of claws and phalanges in juvenile hoatzin are radically divergent from those in adults, yet strikingly similar to those of Archaeopteryx. The locomotor plasticity observed in the hoatzin suggests that transitional forms that retained claws on the wings could have also used them for locomotion.
Assuntos
Aves/crescimento & desenvolvimento , Aves/fisiologia , Voo Animal , Aprendizagem , Animais , Fenômenos Biomecânicos , Aves/embriologia , Extremidades , Marcha , Destreza Motora , Natação , Asas de Animais , Microtomografia por Raio-XRESUMO
The hoatzin is unique among known avian species because of the fermentative function of its enlarged crop. A small-bodied flying foregut fermenter is a paradox, and this bird provides an interesting model to examine how diet selection and the gut microbiota contribute to maximizing digestive efficiency. Therefore, we characterized the bacterial population in the crop of six adult hoatzins captured from the wild. A total of 1,235 16S rRNA gene sequences were grouped into 580 phylotypes (67% of the pooled species richness sampled, based on Good's coverage estimator, with C(ACE) and Chao1 estimates of 1,709 and 1,795 species-level [99% identity] operational taxonomic units, respectively). Members of 9 of the approximately 75 known phyla in Bacteria were identified in this gut habitat; the Firmicutes were dominant (67% of sequences, belonging to the classes Clostridia, Mollicutes, and Bacilli), followed by the Bacteroidetes (30%, mostly in the order Bacteroidales), Proteobacteria (1.8%), and Lentisphaerae, Verrucomicrobia, TM7, Spirochaetes, Actinobacteria, and Aminanaerobia (all <0.1%). The novelty in this ecosystem is great; 94% of the phylotypes were unclassified at the "species" level and thus likely include novel cellulolytic lineages.
Assuntos
Bactérias/classificação , Bactérias/isolamento & purificação , Biodiversidade , Aves/microbiologia , Papo das Aves/microbiologia , Animais , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Dados de Sequência Molecular , Filogenia , RNA Bacteriano/genética , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Homologia de Sequência do Ácido NucleicoRESUMO
A key parameter to constrain predictive, bottom-up circuit models of a given brain domain is the number and position of the neuronal populations involved. These include not only the neurons whose bodies reside within the domain, but also the neurons in distant regions that innervate the domain. The mouse visual cortex receives its main subcortical input from the dorsal lateral geniculate nucleus (dLGN) and the lateral posterior (LP) complex of the thalamus. The latter consists of three different nuclei: lateral posterior lateral (LPL), lateral posterior medial rostral (LPMR), and lateral posterior medial caudal (LPMC), each exhibiting specific patterns of connections with the various visual cortical areas. Here, we have determined the number of thalamocortical projection neurons and interneurons in the LP complex and dLGN of the adult C57BL/6 male mouse. We combined Nissl staining and histochemical and immunolabeling methods for consistently delineating nuclei borders, and applied unbiased stereological cell counting methods. Thalamic interneurons were identified using GABA immunolabeling. The C57BL/6 dLGN contains â¼21,200 neurons, while LP complex contains â¼31,000 total neurons. The dLGN and LP are the only nuclei of the mouse dorsal thalamus containing substantial numbers GABA-immunoreactive interneurons. These interneurons, however, are scarcer than previously estimated; they are 5.6% of dLGN neurons and just 1.9% of the LP neurons. It can be thus inferred that the dLGN contains â¼20,000 and the LP complex â¼30,400 thalamocortical projection neurons (â¼12,000 in LPL, 15,200 in LPMR, and 4,200 in LPMC). The present dataset is relevant for constraining models of mouse visual thalamocortical circuits, as well as for quantitative comparisons between genetically modified mouse strains, or across species.
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The treatment of Helicobacter pylori infection is complicated by antibiotic resistance. A high levofloxacin (LVX) resistance rate was previously demonstrated in H. pylori isolates from gastric mucosa (40%) and esophagus (19%) in individual hosts of a Venezuelan population. We aimed to assess the molecular mechanisms of LVX resistance and susceptibility in isolates from the gastroesophageal mucosa, by studying point mutations in the quinolone resistance-determining region of gyrA and gyrB genes. Sequencing of gyrA and gyrB genes (N = 120) helped to identify point mutations in 60 isolates (30 from antrum and 30 from esophagus) of five dyspeptic patients. Double (Asn87Thr and Asp91Asn) and single (Asn87Ile or Asn87Thr) mutations in the gyrA gene were identified in the esophageal mucosa. These mutations have been commonly found in the stomach. Occurrence of a single (Asn87Ile) mutation was associated with high resistance (minimum inhibitory concentration ≥ 32 µg/mL) to LVX. Only a single (Ser479Gly) mutation was found in the gyrB gene in both mucosae. One patient presented isolates with no mutations in the two genes studied. Isolates with the same mutation pattern in individual hosts revealed identical genetic profiles for these genes, confirming that isolates identified in the esophageal mucosa come from isolates colonizing the stomach. Helicobacter pylori resistance to LVX in the esophagus is related to double- and single-point mutations in gyrA and gyrB genes, such as those found in the stomach. Levofloxacin should be applied with caution, because its antibiotic effect on H. pylori is decreasing in Latin America, perhaps owing to high prescription rates.
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Antibacterianos/farmacologia , DNA Girase/genética , Farmacorresistência Bacteriana/genética , Infecções por Helicobacter/microbiologia , Helicobacter pylori/genética , Levofloxacino/farmacologia , Substituição de Aminoácidos , Mucosa Esofágica/microbiologia , Mucosa Gástrica/microbiologia , Gastroenterologia , Infecções por Helicobacter/epidemiologia , Helicobacter pylori/efeitos dos fármacos , Helicobacter pylori/isolamento & purificação , Humanos , Testes de Sensibilidade Microbiana , Mutação Puntual , Quinolinas/farmacologia , Venezuela/epidemiologiaRESUMO
Helicobacter species can colonize digestive tract of animals and humans and have been associated with gastrointestinal diseases; however, this genus has not been studied in crocodiles. Our objective was to detect by PCR Helicobacter genus and Helicobacter pylori in oral and cloacal swabs from Orinoco crocodiles of two wild (Cojedes River System and Capanaparo River) and two captive breeding centers (CBCs; Masaguaral Ranch and UNELLEZ) populations. Bacterial DNA was found in 100% of oral samples (10 wild and 10 captives), and in the 95% of cloacal samples (10 wild and 9 captives). In wild populations, Helicobacter spp. was not detected, whereas in CBCs, Helicobacter was detected in 10% of the oral samples, and 66.7% of cloacal samples. H. pylori was detected in two Orinoco crocodiles. Two cloacal non-pylori Helicobacter amplicons were sequenced, showing low similarity (≤97%) to Helicobacter sequences reported. This is the first report of Helicobacter species, including H. pylori in Crocodylus intermedius from CBCs.
Assuntos
Jacarés e Crocodilos/microbiologia , Cloaca/microbiologia , Helicobacter/isolamento & purificação , Boca/microbiologia , Criação de Animais Domésticos , Animais , Feminino , Masculino , VenezuelaRESUMO
The fecal contamination of raw seafood by indicators and opportunistic pathogenic microorganisms represents a public health concern. The objective of this study was to investigate the presence of enteric bacteria colonizing oysters collected from a Venezuelan touristic area. Oyster samples were collected at the northwestern coast of Venezuela and local salinity, pH, temperature, and dissolved oxygen of seawater were recorded. Total and fecal coliforms were measured for the assessment of the microbiological quality of water and oysters, using the Multiple Tube Fermentation technique. Analyses were made using cultures and 16S rRNA gene sequencing. Diverse enrichment and selective culture methods were used to isolate enteric bacteria. We obtained pure cultures of Gram-negative straight rods with fimbriae from Isognomon alatus and Crassostrea rhizophorae. Our results show that P. mirabilis was predominant under our culture conditions. We confirmed the identity of the cultures by biochemical tests, 16S rRNA gene sequencing, and data analysis. Other enterobacteria such as Escherichia coli, Morganella morganii and Klebsiella pneumoniae were also isolated from seawater and oysters. The presence of pathogenic bacteria in oysters could have serious epidemiological implications and a potential human health risk associated with consumption of raw seafood.