Genetic diversity of laboratory strains and implications for research: The case of Aedes aegypti.

The yellow fever mosquito (Aedes aegypti), is the primary vector of dengue, Zika, and chikungunya fever, among other arboviral diseases. It is also a popular laboratory model in vector biology due to its ease of rearing and manipulation in the lab. Established laboratory strains have been used worldwide in thousands of studies for decades. Laboratory evolution of reference strains and contamination among strains are potential severe problems that could dramatically change experimental outcomes and thus is a concern in vector biology. We analyzed laboratory and field colonies of Ae. aegypti and an Ae. aegypti-derived cell line (Aag2) using 12 microsatellites and ~20,000 SNPs to determine the extent of divergence among laboratory strains and relationships to their wild relatives. We found that 1) laboratory populations are less genetically variable than their field counterparts; 2) colonies bearing the same name obtained from different laboratories may be highly divergent; 3) present genetic composition of the LVP strain used as the genome reference is incompatible with its presumed origin; 4) we document changes in two wild caught colonies over ~16 generations of colonization; and 5) the Aag2 Ae. aegypti cell line has experienced minimal genetic changes within and across laboratories. These results illustrate the degree of variability within and among strains of Ae. aegypti, with implications for cross-study comparisons, and highlight the need of a common mosquito repository and the implementation of strain validation tools.

Fully Automated Pipetting Sorting System for Different Morphological Phenotypes of Zebrafish Embryos.

Systems biology methods, such as transcriptomics and metabolomics, require large numbers of small model organisms, such as zebrafish embryos. Manual separation of mutant embryos from wild-type embryos is a tedious and time-consuming task that is prone to errors, especially if there are variable phenotypes of a mutant. Here we describe a zebrafish embryo sorting system with two cameras and image processing based on template-matching algorithms. In order to evaluate the system, zebrafish rx3 mutants that lack eyes due to a patterning defect in brain development were separated from their wild-type siblings. These mutants show glucocorticoid deficiency due to pituitary defects and serve as a model for human secondary adrenal insufficiencies. We show that the variable phenotypes of the mutant embryos can be safely distinguished from phenotypic wild-type zebrafish embryos and sorted from one petri dish into another petri dish or into a 96-well microtiter plate. On average, classification of a zebrafish embryo takes approximately 1 s, with a sensitivity and specificity of 87% to 95%, respectively. Other morphological phenotypes may be classified and sorted using similar techniques.

Choosing the right species in research.

When designing animal studies, investigators must choose a species that is appropriate for the research. In this paper, the author examines various criteria that can be used to guide this selection. He discusses the concepts of phylogenetic group and sentience and finds them not to be useful in the selection of appropriate species in biomedical research. He identifies other criteria that are more useful as justifications for species selection, including susceptibility to a targeted disease process, tendency to engage in a targeted behavior, suitable size for the experimental techniques to be used, presence of a large body of data relevant to the study, species specificity (the species itself is the target of the research), intergenerational interval, similarity to humans, contractual specification and existing guidelines. He proposes that investigators should use these justifications, and perhaps others, to choose the most scientifically appropriate species for animal studies.

Use of nonhuman primates in research in North America.

In North America, the biomedical research community faces social and economic challenges to nonhuman primate (NHP) importation that could reduce the number of NHP available for research needs. The effect of such limitations on specific biomedical research topics is unknown. The Association of Primate Veterinarians (APV), with assistance from the Centers for Disease Control and Prevention, developed a survey regarding biomedical research involving NHP in the United States and Canada. The survey sought to determine the number and species of NHP maintained at APV members' facilities, current uses of NHP to identify the types of biomedical research that rely on imported animals, and members' perceived trends in NHP research. Of the 149 members contacted, 33 (22%) replied, representing diverse facility sizes and types. Cynomolgus and rhesus macaques were the most common species housed at responding institutions and comprised the majority of newly acquired and imported NHP. The most common uses for NHP included pharmaceutical research and development and neuroscience, neurology, or neuromuscular disease research. Preclinical safety testing and cancer research projects usually involved imported NHP, whereas research on aging or degenerative disease, reproduction or reproductive disease, and organ or tissue transplantation typically used domestic-bred NHP. The current results improve our understanding of the research uses for imported NHP in North America and may facilitate estimating the potential effect of any future changes in NHP accessibility for research purposes. Ensuring that sufficient NHP are available for critical biomedical research remains a pressing concern for the biomedical research community in North America.

Development of a Nasonia vitripennis outbred laboratory population for genetic analysis.

The parasitoid wasp genus Nasonia has rapidly become a genetic model system for developmental and evolutionary biology. The release of its genome sequence led to the development of high-resolution genomic tools, for both interspecific and intraspecific research, which has resulted in great advances in understanding Nasonia biology. To further advance the utility of Nasonia vitripennis as a genetic model system and to be able to fully exploit the advantages of its fully sequenced and annotated genome, we developed a genetically variable and well-characterized experimental population. In this study, we describe the establishment of the genetically diverse HVRx laboratory population from strains collected from the field in the Netherlands. We established a maintenance method that retains genetic variation over generations of culturing in the laboratory. As a characterization of its genetic composition, we provide data on the standing genetic variation and estimate the effective population size (N(e)) by microsatellite analysis. A genome-wide description of polymorphism is provided through pooled resequencing, which yielded 417,331 high-quality SNPs spanning all five Nasonia chromosomes. The HVRx population and its characterization are freely available as a community resource for investigators seeking to elucidate the genetic basis of complex trait variation using the Nasonia model system.

Establishment and characterisation of salmon louse (Lepeophtheirus salmonis (Krøyer 1837)) laboratory strains.

The salmon louse (Lepeophtheirus salmonis (Krøyer 1837)) is an ectoparasitic copepod which represents a major pathogen of wild and farmed salmonid fishes in the marine environment. In order to facilitate research on this ecologically and economically important parasite, a hatchery and culturing system permitting the closure of the life-cycle of L. salmonis in the laboratory was developed. Here, the hatchery system, breeding practices, and selected louse strains that have been maintained in culture in the period 2002-2009 are presented. The hatchery and culture protocol gave rise to predictable hatching of larvae and infections of host fish, permitting the cultivation of specific strains of L. salmonis for 22 generations. Both in- and out-bred lice and mutant colour strains have been established, and some of these strains were characterised by microsatellite DNA markers confirming their pedigree. No evidence of inbreeding depression, fitness or morphological changes was observed in any of the strains cultured. It is suggested that the culturing system, and the strains produced represent a significant resource for future research on this parasite.

Variability of diurnality in laboratory rodents.

The locomotor activity rhythms of domestic mice, laboratory rats, Syrian hamsters, Siberian hamsters, Mongolian gerbils, degus, and Nile grass rats were compared. Running-wheel activity was monitored under a light-dark cycle with 12 h of light and 12 h of darkness per day. Nile grass rats were found to be reliably diurnal, whereas laboratory rats, Siberian hamsters, domestic mice, and Syrian hamsters were reliably nocturnal. Both diurnal and nocturnal subgroups were observed in Mongolian gerbils and degus. A downward gradient of diurnality was observed from Mongolian gerbils classified as diurnal, degus classified as diurnal, gerbils classified as nocturnal, and degus classified as nocturnal. Nocturnal degus remained nocturnal when tested with an infrared motion detector without running wheels. Thus, although the diurnal-nocturnal dichotomy could be applied to some of the species, it was not appropriate for others. The dichotomy may reflect researchers' needs for systematization more than a natural distinction between species. Through mechanisms as yet poorly understood, the balance between entraining and masking processes seems to generate a gradient of temporal niches that runs from predominantly diurnal species to predominantly nocturnal species with many chronotypes in between, including species that exhibit wide intra-species gradients of temporal niche.

Ethics for research using animals in Italy.

The purpose of this report is to describe the legislative and ethical context in Italy regarding technology and the need for innovative research versus responsibility toward the living world. Questions regarding responsible behaviour in the use of animals for research are examined, and special attention is focused on the implementation of the Three Rs concept (reduction, refinement and replacement) and on considerations of Catholic dogma.

Analysis of pancreatic polypeptide cDNA from the house musk shrew, Suncus murinus, suggests a phylogenetically closer relationship with humans than for other small laboratory animal species.

Pancreatic polypeptide was isolated and sequenced from endocrine cells of the pancreas from an insectivore, the house musk shrew, Suncus murinus. The primary sequence was APLEPAYPGD(10)NATPEQMAQY(20)AAELRKYINM(30)VTRPRYamide. This is the first polypeptide hormone to be characterised from this species and is typical of the primary sequences of pancreatic polypeptide of other animals, being a C-terminal-amidated peptide with 36 residues. Comparison with several vertebrate sequences shows that it has more in common with the human form than do the forms from common laboratory animals such as rabbits, rats, mice and guinea-pigs.