The relationship between the distribution of genetically distinct inbred lines and upper lethal temperature in Lasaea rubra

Lasaea rubra is an inbreeding bivalve species, living at most heights on rocky shores. Freshly collected animals from different shore heights showed significantly different upper median lethal temperatures (MLTs), with upper shore animals having higher MLTs than lower shore specimens. Experiments with animals acclimated for at least one month to a single temperature (15°C) demonstrated that these differences in upper MLT were unaffected by thermal acclimation. Electrophoretic investigation showed that the differences in thermal response had a genetic basis. Homogeneous populations of the high-water inbred line (‘Inbred line A’) had a higher MLT than homogeneous populations of ‘Inbred line C’ which was found on the middle and lower shore. No differences were detected between the MLTs of separate populations of Inbred lines A or C. A third inbred line (‘Inbred line B’) was found on the middle shore, but no homogeneous populations were found. However, indirect evidence suggests that Inbred line B has a thermal response intermediate between those of Inbred lines A and C. Study of populations made up of mixtures of inbred lines confirmed the relationship between upper MLTs and genetic composition of the population.

Localization of polymorphic N-acetyltransferase (NAT2) in tissues of inbred mice

Like humans, mice exhibit polymorphism in the N-acetylation of aromatic amines, many of which are toxic and/or carcinogenic. Mice have three N-acetyltransferase (Nat) genes, Nat1, Nat2 and Nat3, and Nat2 is known to be polymorphic. There is a dramatic difference in the acetylation of NAT2 substrates by blood from fast (C57BL/6J) compared with slow acetylator (A/J) mice. However, the acetylation of these substrates by liver cytosols from the two strains is very similar. In order to determine whether the expression of the NAT2 protein corresponded with the activities measured, a polyclonal antipeptide antisera was raised against the C-terminal decapeptide of NAT2 and characterized using recombinant murine NAT2 antigen. Enzyme-linked immunosorbent assays (ELISAs) demonstrated that the anti-NAT2 antiserum bound in a concentration-dependent fashion to recombinant NAT2. Immunochemical analysis of mouse liver cytosols from C57BL/6J or A/J livers indicated that the level of NAT2 protein expressed in the two strains was similar. Immunohistochemical staining of C57BL/6J liver with anti-NAT2 antiserum showed that NAT2 was expressed in hepatocytes throughout the liver although the intensity of staining in the perivenous (centrilobular) region was higher than that in the periportal region. NAT2 was also detected in epithelial cells in the lung, kidney, bladder, small intestine and skin as well as in erythrocytes and lymphocytes in the spleen and hair follicles and sebaceous glands in the skin. Characterization of the distribution of NAT2 will be of value in elucidating the role of polymorphic N-acetylation in protecting the organism from environmental insults as well as in endogenous metabolism.

The role of Rhipicephalus sanguineus sensu lato saliva in the dissemination of Rickettsia conorii in C3H/HeJ mice.

Animal models have been developed for the study of rickettsial pathogenesis. However, to understand what occurs during the natural route of rickettsial transmission via the tick bite, the role of tick saliva should be considered in these models. To address this, we analysed the role of tick saliva in the transmission of Rickettsia conorii (Rickettsiales: Rickettsiaceae) in a murine host by intradermally (i.d.) inoculating two groups of susceptible C3H/HeJ mice with this Rickettsia, and infesting one group with nymphal Rhipicephalus sanguineus sensu lato (Ixodida: Ixodidae) ticks. Quantification of bacterial loads and mRNA levels of interleukin-1β (IL-1β), IL-10 and NF-κB was performed in C3H/HeJ lung samples by real-time quantitative polymerase chain reaction (PCR) and real-time reverse transcriptase PCR, respectively. Lung histology was examined to evaluate the pathological manifestations of infection. No statistically significant difference in bacterial load in the lungs of mice was observed between these two groups; however, a statistically significant difference was observed in levels of IL-1β and NF-κB, both of which were higher in the group inoculated with rickettsiae but not infected with ticks. Lung histology in both groups of animals revealed infiltration of inflammatory cells. Overall, this study showed that i.d. inoculation of R. conorii caused infection in the lungs of C3H/HeJ mice and tick saliva inhibited proinflammatory effects.

Cardiovascular response to beta-adrenergic blockade or activation in 23 inbred mouse strains.

We report the characterisation of 27 cardiovascular-related traits in 23 inbred mouse strains. Mice were phenotyped either in response to chronic administration of a single dose of the beta-adrenergic receptor blocker atenolol or under a low and a high dose of the beta-agonist isoproterenol and compared to baseline condition. The robustness of our data is supported by high trait heritabilities (typically H(2)>0.7) and significant correlations of trait values measured in baseline condition with independent multistrain datasets of the Mouse Phenome Database. We then focused on the drug-, dose-, and strain-specific responses to beta-stimulation and beta-blockade of a selection of traits including heart rate, systolic blood pressure, cardiac weight indices, ECG parameters and body weight. Because of the wealth of data accumulated, we applied integrative analyses such as comprehensive bi-clustering to investigate the structure of the response across the different phenotypes, strains and experimental conditions. Information extracted from these analyses is discussed in terms of novelty and biological implications. For example, we observe that traits related to ventricular weight in most strains respond only to the high dose of isoproterenol, while heart rate and atrial weight are already affected by the low dose. Finally, we observe little concordance between strain similarity based on the phenotypes and genotypic relatedness computed from genomic SNP profiles. This indicates that cardiovascular phenotypes are unlikely to segregate according to global phylogeny, but rather be governed by smaller, local differences in the genetic architecture of the various strains.

Anatomic correlates of strial presbycusis in recombinant inbred mouse strains

This project attempts to identify anatomic features that predict the range of the ‘normal’ endocochlear potential in young inbred mice. Cochlear lateral wall histologic metrics were compared in recombinant inbred (RI) mouse strains formed from BALB/c and C57BL/6 mice.

A maize bacterial artificial chromosome (BAC) library from the European flint inbred line F2

We report here the construction and characterisation of a BAC library from the maize flint inbred line F2, widely used in European maize breeding programs. The library contains 86,858 clones with an average insert size of approximately 90 kb, giving approximately 3.2-times genome coverage. High-efficiency BAC cloning was achieved through the use of a single size selection for the high-molecular-weight genomic DNA, and co-transformation of the ligation with yeast tRNA to optimise transformation efficiency. Characterisation of the library showed that less than 0.5% of the clones contained no inserts, while 5.52% of clones consisted of chloroplast DNA. The library was gridded onto 29 nylon filters in a double-spotted 8 × 8 array, and screened by hybridisation with a number of single-copy and gene-family probes. A 3-dimensional DNA pooling scheme was used to allow rapid PCR screening of the library based on primer pairs from simple sequence repeat (SSR) and expressed sequence tag (EST) markers. Positive clones were obtained in all hybridisation and PCR screens carried out so far. Six BAC clones, which hybridised to a portion of the cloned Rp1-D rust resistance gene, were further characterised and found to form contigs covering most of this complex resistance locus.

Differential effect of inbred mouse strain (C57BL/6, DBA/2, 129T2) on insulin secretory function in response to a high fat diet

The increasing production of genetically-modified mouse models has necessitated studies to determine the inherent physiological characteristics of commonly used mouse strains. In this study we examined insulin secretory function in response to an intravenous bolus of glucose or glucose plus arginine in anesthetized C57BL/6, DBA/2 and 129T2 mice fed either a control or high fat diet for 6 weeks. The results show that 129T2 mice had higher fasting plasma glucose levels and lower fasting plasma insulin levels compared with C57BL/6 and DBA/2 mice regardless of diet. Furthermore, 129T2 mice were glucose intolerant and secreted significantly less insulin in response to glucose and glucose plus arginine irrespective of diet compared with the other two strains of mice. DBA/2 mice hypersecreted insulin in response to glucose and glucose plus arginine compared with C57BL/6 and 129T2 mice. Moreover while first phase insulin secretion was appropriately increased in response to the high fat diet in C57BL/6 and 129T2 mice, this was not the case for DBA/2 mice. Mean islet area was decreased in response to a high fat diet in DBA/2 mice, while there was no dietary effect on the other two strains. This study highlights the inherent genetic differences that exist among seemingly normal strains of mice that are commonly used to make transgenic and knockout mice. Understanding these differences will provide researchers with the information to choose the appropriate genetic background on which to express their particular genetic alteration.

EFFECT OF MACROPHAGE BLOCKADE ON THE RESISTANCE OF INBRED MICE TO PARACOCCIDIOIDES-BRASILIENSIS INFECTION

The effect of macrophage blockade on the natural resistance and on the adaptative immune response of susceptible (B10.D2/oSn) and resistant (A/Sn) mice to Paracoccidioides brasiliensis infection was investigated. B10.D2/oSn and A/Sn mice previously injected with colloidal carbon were infected ip with yeast cells to determine the 50% lethal dose, and to evaluate the anatomy and histopathology, macrophage activation, antibody production and DTH reactions. Macrophage blockade rendered both resistant and susceptible mice considerably more susceptible to infection, as evidenced by increased mortality and many disseminated lesions. P. brasiliensis infection and/or carbon treatment increased the ability of macrophages from resistant mice to spread up to 25 days after treatment. In susceptible mice the enhanced spreading capacity induced by carbon treatment was impaired at ail assayed periods except at 1 week after infection. Macrophage blockade enhanced DTH reactions in resistant mice, but did not alter these reactions in susceptible mice, which remained anergic. To the contrary, macrophage blockade enhanced specific antibody production by susceptible mice, but did nor affect the low levels produced by resistant mice. The effect of macrophage blockade confirms the natural tendency of resistant animals to mount DTH reactions in the course of the disease and the preferential antibody response developed by susceptible mice after P. brasiliensis infection. on the whole, macrophage functions appear to play a fundamental role in the natural and acquired resistance mechanisms to P. brasiliensis infection.

Karyotype variability in tropical maize sister inbred lines and hybrids compared with KYS standard line

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Experimental infection of inbred BALB/c and A/J mice with Massachusetts and Brazilian strains of infectious bronchitis virus (IBV)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Description of 90 inbred lines of castor plant (Ricinus communis L.)

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)