Severe necrotizing myocarditis caused by serratia marcescens infection in an axolotl (Ambystoma mexicanum)

This report provides the first account of the pathological changes associated with infection by Serratia marcescens in an adult male axolotl. The infection resulted in septicaemia with severe multifocal necrotizing myocarditis. The latter lesion evolved to cardiac rupture, haemopericardium and death resulting from cardiac tamponade. This animal was exposed to higher than usual temperatures (24–25°C) 2 weeks before the onset of disease and this may have resulted in immunocompromise and opportunistic bacterial infection. S. marcescens was isolated from the coelomic and pericardial cavity. Both isolates were identical and were resistant to β-lactam antibiotics, but not to aminoglycosides or fluoroquinolones. The production of red prodigiosin pigment by the bacterium suggested an environmental origin. Overall, the clinical and histopathological presentation suggests that S. marcescens should be included in the list of aetiological agents of the ‘red-leg’/bacterial dermatosepticaemia syndrome of amphibians.     

The use of transgenics in the laboratory axolotl

The ability to generate transgenic animals sparked a wave of research committed to implementing such technology in a wide variety of model organisms. Building a solid base of ubiquitous and tissue-specific reporter lines has set the stage for later interrogations of individual cells or genetic elements. Compared to other widely used model organisms such as mice, zebrafish and fruit flies, there are only a few transgenic lines available in the laboratory axolotl (Ambystoma mexicanum), although their number is steadily expanding. In this review, we discuss a brief history of the transgenic methodologies in axolotl and their advantages and disadvantages. Next, we discuss available transgenic lines and insights we have been able to glean from them. Finally, we list challenges when developing transgenic axolotl, and where further work is needed in order to improve their standing as both a developmental and regenerative model.     

Gene and transgenics nomenclature for the laboratory axolotl—Ambystoma mexicanum

The laboratory axolotl (Ambystoma mexicanum) is widely used in biological research. Recent advancements in genetic and molecular toolkits are greatly accelerating the work using axolotl, especially in the area of tissue regeneration. At this juncture, there is a critical need to establish gene and transgenic nomenclature to ensure uniformity in axolotl research. Here, we propose guidelines for genetic nomenclature when working with the axolotl.     

Histochemical study of the heart of the axolotl (Ambystoma mexicanum)

We have investigated the presence of cells containing monoamines, substance P, and neuron-specific enolase (NSE) in the heart and in the pericardial wall of a urodele amphibian, the axolotl. Fibers containing substance P-like immunoreactivity were present in the heart but not in the pericardial wall. Also present in the heart were small branched cells, which stained metachromatically with toluidine blue. Similar cells were found in the peritoneum and were tentatively identified as mast cells. NSE-immunoreactive fibers were found both in the heart and in the pericardial wall. Small intensely fluorescent (SIF) cells of the pericardial wall contained a high concentration of norepinephrine but no other monoamines, substance P, or NSE. Comparison with data available for the mudpuppy, Necturus maculosus, a closely related amphibian species, suggests that the innervation of the heart in the axolotl is substantially different. © 1992 Wiley-Liss, Inc.     

Eye enucleation and regeneration of neural retina in axolotl larvae (Ambystoma mexicanum)

The eyes of Axolotl larvae were enucleated at stages 30 and 37. Animals with single dorsomedian eyes resulted in the first case (i.e. stage 30). When a piece of pigment epithelium was re-implanted into stage 37 animals at the site of the lesion, limited regeneration was observed when the implant formed a vesicle, but, when the pigment epithelium remained "open" regeneration of the neural retina was extensive. The possible resons for this difference was discussed.     

Regional differences in cell density and cell genesis in the olfactory epithelium of the salamander,Ambystoma tigrinum

Summary The olfactory epithelium undergoes continuous regeneration. The present quantitative study uses tritiated thymidine autoradiography to investigate regional differences in the rate of olfactory epithelial cell genesis in the tiger salamander. There was a significant gradient in the incorporation of thymidine from the posterior to the anterior in the nasal cavity: the posterior epithelium underwent cell genesis much faster than the anterior. Additionally, the posterior epithelium was thinner and contained fewer cells than the anterior, although the proportions of receptor, supporting and basal cells remained about the same throughout the epithelium. After 5 or 20 days most of the labelled cells were found in the basal cell layer, although there were a few labelled supporting cells. This confirms observations in other species that there are two populations of dividing cells in the olfactory epithelium: the basal cells which give rise to receptor cells, and the supporting cells. The gradients in epithelial thickness, receptor cells, and the rate of cell genesis parallel a gradient in responsiveness to odorants observed in electrophysiological studies (Mackay-Sim et al. 1982; Mackay-Sim and Shaman 1984). The significance of these anatomical and physiological gradients is presently unclear.     

PTPRC (CD45) variation and disease association studied using single nucleotide polymorphism tagging

CD45 is a haemopoietic tyrosine phosphatase, crucial for lymphocyte signalling. Two polymorphisms (C77G and A138G), which alter CD45 isoform expression, are associated with autoimmune and infectious diseases. Using HapMap data, we show that there is substantial linkage disequilibrium across the CD45 gene (PTPRC), with similar patterns in different populations. Employing a set of single nucleotide polymorphisms, correlated with a substantial proportion of variation across this gene, we tested for association with type 1 diabetes, Graves' disease in a Japanese population, hepatitis C in UK population and tuberculin response in a Chinese population. A limited number of common haplotypes was found. Most 138G alleles are present on only one haplotype, which is associated with Graves' disease, supporting previous data that A138G is a functionally important CD45 polymorphism.     

A scanning electron microscopy and histological study on the effects of the mutant eyeless (e/e) gene upon the hypothalamus in the Mexican axolotl Ambystoma mexicanum Shaw

A scanning electron microscopy, histological, and immunochemical investigation examined the effects of the mutant gene (e) upon hypothalamic development in the Mexican axolotl. The adult eyeless mutant is sterile. Previous studies indicated that this reproductive defect was due to the mutation's effect upon the hypothalamus. The present study demonstrated the pleiotropic effects of the eyeless gene upon development of the hypothalamus. Scanning electron microscopy studies looked at the early ontogeny of the hypothalamohypophyseal system. The major morphological difference observed in the hypothalamus of normals compared to eyeless mutants was the reduced nature or complete lack of a preoptic recess in eyeless mutants. Early embryonic tissue movements also differed when normal siblings were compared to eyeless mutant axolotls. Histological examination looking for paraldehyde-fuchsin-positive secretory neurons revealed a paired nucleus preopticus in both normals and eyeless mutants, but this region lacked the emanating paraldehyde-fuchsin-positive fiber tracts in eyeless mutants. The neurohypophysis of the eyeless mutants was atrophied and contained far less paraldehyde-fuchsin-positive material when compared to normal axolotls. Immunochemical studies were done to look at the distribution of immunoreactive luteinizing-hormone-releasing hormone (ir-LHRH) in brains of eyed and eyeless mutant axolotls of different stages. This study detected deficiencies in ir-LHRH in the anterior hypothalamus of eyeless mutants. In general in the eyeless mutant axolotl, the observed anterior hypothalamic deficiencies are comparable to those observed in anurans which have had their optic vesicles removed. These observations suggest a possible utility of the eyeless mutant axolotl for studies concerned with endocrine development in the absence of hypothalamic modulation.     

Genome-wide single nucleotide polymorphism array analysis reveals recurrent genomic alterations associated with histopathologic features in intrahepatic cholangiocarcinoma

Recent studies indicate that genomic alterations (GAs) are associated with many human malignancies. Genome-wide analysis of GAs involved in intrahepatic cholangiocarcinoma (ICC) and association with histopathologic features are limited. To help characterize this relatively rare neoplasm, we collected 32 frozen tissue samples of ICC to study GAs and molecular karyotypes by using single-nucleotide polymorphism array. Recurrent GAs occurring in at least 40% of the patients were further correlated with histopathologic features. Gain of 1q21.3-q23.1 and losses of 1p36.33-p35.3 and 3p26.3-p13 were significantly associated with larger tumor size more than 5 cm in diameter; and loss of 4q13.2-q35.2 with tumor multiplicity. Moreover, losses of 1p36.32-p35.3, 3p26.3-p22.2, 4q13.1-q21.23, 4q31.3-q34.3 and 4q34.3-35.2 were inclined to be associated with high histological grade. As to tumor vascular invasion, gain of 1q21.3-q23.1 and losses of 3p22.1-p12.3 and 4q13.2-q35.2 were significantly associated with tumor vascular invasion. Some regions were concurrently associated with multiple histopathologic characteristics, including loss of 4q13.2-q35.2 associated with larger tumor size, high histological grade and vascular invasion; losses of 1p36.33-p35.3 and 3p26.3-p22.2 with larger tumor size and high histological grade; and gain of 1q21.3-q23.1 with larger tumor size and vascular invasion. Our study indicates that complex chromosomal instability is characteristic of ICC. Detecting crucial GAs will enable risk stratification and development of personalized therapies.     

Association of rs1285933 single nucleotide polymorphism in CLEC5A gene with dengue severity and its functional effects

Outbreaks of the Zika, dengue, and chikungunya viruses, especially in the Americas, pose a global threat due to their rapid spread and difficulty controlling the vector. Extreme phenotypes are often observed, from asymptomatic to severe clinical manifestations, which are well-studied in dengue. Host variations are also important contributors to disease outcomes, and many case-control studies have associated single nucleotide polymorphisms (SNPs) with severe dengue. Here, we found that the TC genotype and T-carriers for SNP rs1285933 in the C-type lectin superfamily member 5 (CLEC5A) gene was associated with severe dengue in a Northern Brazilian population (OR = 2.75 and p-value = 0.01, OR = 2.11 and p-value = 0.04, respectively). We also tested the functional effect of the CLEC5A protein and found that it is upregulated on the surface of human monocytes after in vitro dengue infection. CLEC5A was correlated with viral load inside the monocytes (Spearman r = 0.55, p = 0.008) and TNF production in culture supernatants (Spearman r = 0.72, p = 0.03). Analysis of mRNA in blood samples from DENV4-infected patients exhibiting mild symptoms showed that CLEC5A mRNA expression is correlated with TNF (r = 0.67, p = 0.0001) and other immune mediators. Monocytes from rs1285933 TT/TC individuals showed lower CLEC5A expression compared to CC genotypes. However, in these cells, CLEC5A was not correlated with TNF production. In summary, we confirmed that CLEC5A is genetically associated with dengue severity outcome, playing a central role during the immune response triggered by a dengue viral infection, and rs1285933 is a relevant SNP that is able to regulate signaling pathways after interactions between the dengue virus and CLEC5A receptors.