Genomic characterization of mutant laboratory mouse strains by exome sequencing and annotation lift-over

Background

Exome sequencing has become a popular method to evaluate undirected mutagenesis experiments in mice. However, the most suitable mouse strain for the biological model may be relatively distant from the standard mouse reference genome. For pinpointing causative variants, a matching reference with gene annotations is essential, but not always readily available.

Results

We present an approach that allows to use murine Ensembl annotations on alternative mouse strain assemblies. We resolved ENU-induced mutation screening for 8 phenotypic mutant lines generated on C3HeB/FeJ background aligning the sequences against the closely related, but not annotated reference of C3H/HeJ. Variants occurring in all strains were filtered out as specific for the C3HeB/FeJ strain but unrelated to mutagenesis. Variants occurring exclusively in all individuals of one mutant line and matching the inheritance model were selected as mutagenesis-related. These variants were annotated with gene and exon names lifted over from the standard murine reference mm9 to C3H/HeJ using megablast. For each mutant line, we could restrict the results to exonic variants in between 1 and 23 genes.

Conclusions

The presented method of exonic annotation lift-over proved to be a valuable tool in the search for mutagenesis-derived coding genomic variants and the assessment of genotype-phenotype relationships.     

Survival of Mycobacterium avium ssp. paratuberculosis in yoghurt and in commercial fermented milk products containing probiotic cultures

Aims: To assess the survival of Mycobacterium avium ssp. paratuberculosis (MAP) in yoghurt and commercial fermented milk products containing probiotic strains. Methods and Results: Whole and skimmed UHT milk artificially inoculated with MAP were used to manufacture yoghurt, using two different yoghurt starter cultures. Five commercial fermented milk products were inoculated with MAP. Two different MAP strains were studied. The survival of MAP in all products was monitored by culture over a 6‐week storage period at 6°C. In yoghurt, MAP counts did not change appreciably during the storage period. Fat content and type of yoghurt starter culture had no consistent effect on the survival of MAP. In the fermented milk products, survival patterns varied but resulted in a 1·5 to ≥3·8 log reduction for the Niebüll strain and a 1·2–2·2 log reduction for the NIZO strain after 6 weeks, depending on the probiotic starters present in the product. Conclusions: MAP easily survived in yoghurt but MAP numbers decreased in fermented milk products containing probiotic cultures. Significance and Impact of the Study: The results contribute to the lack of knowledge on the behaviour of MAP in yoghurt and fermented milk products containing probiotic cultures. This knowledge is valuable in the context of the risk of MAP transmission to humans via yoghurt and the possible contribution of probiotic fermented milk products to the elimination of MAP.     

Evolution-based strategy to generate non-genetically modified organisms Saccharomyces cerevisiae strains impaired in sulfate assimilation pathway

Aims: An evolution‐based strategy was designed to screen novel yeast strains impaired in sulfate assimilation. Specifically, molybdate and chromate resistance was used as selectable phenotype to select sulfate permease–deficient variants that unable to produce sulfites and hydrogen sulfide (H₂S). Methods and Results: Four Saccharomyces cerevisiae parent strains were induced to sporulate. After tetrad digestion, spore suspensions were observed under the microscope to monitor the conjugation of gametes. Then, the cell suspension was inoculated in tubes containing YPD medium supplemented with ammonium molybdate or potassium chromate. Forty‐four resistant strains were obtained and then tested in microvinifications. Three strains with a low sulfite production (SO₂ <10 mg l^?1) and with an impaired H₂S production in grape must without added sulfites were selected. Conclusions: Our strategy enabled the selection of improved yeasts with desired oenological characteristics. Particularly, resistance to toxic analogues of sulfate allowed us to detect strains that unable to assimilate sulfates. Significance and Impact of the Study: This strategy that combines the sexual recombination of spores and application of a specific selective pressure provides a rapid screening method to generate genetic variants and select improved wine yeast strains with an impaired metabolism regarding the production of sulfites and H₂S.     

Mathematical modeling of fungal infection in immune compromised individuals: implications for drug treatment

We present a mathematical model that describes treatment of a fungal infection in an immune compromised patient in which both susceptible and resistant strains are present. The resulting nonlinear differential equations model the biological outcome, in terms of strain growth and cell number, when an individual, who has both a susceptible and a resistant population of fungus, is treated with a fungicidal or fungistatic drug. The model demonstrates that when the drug is only successful at treating the susceptible strain, low levels of the drug cause both strains to be in stable co-existence and high levels eradicate the susceptible strain while allowing the resistant strain to persist or to multiply unchecked. A modified model is then described in which the drug is changed to one in which both strains are susceptible, and subsequently, at the appropriate level of treatment, complete eradication of both fungal strains ensues. We discuss the model and implications for treatment options within the context of an immune compromised patient.     

Antidepressant-like responses to lithium in genetically diverse mouse strains

A mood stabilizing and antidepressant response to lithium is only found in a subgroup of patients with bipolar disorder and depression. Identifying strains of mice that manifest differential behavioral responses to lithium may assist in the identification of genomic and other biologic factors that play a role in lithium responsiveness. Mouse strains were tested in the forced swim test (FST), tail suspension test (TST) and open-field test after acute and chronic systemic and intracerebroventricular (ICV) lithium treatments. Serum and brain lithium levels were measured. Three (129S6/SvEvTac, C3H/HeNHsd and C57BL/6J) of the eight inbred strains tested, and one (CD-1) of the three outbred strains, showed an antidepressant-like response in the FST following acute systemic administration of lithium. The three responsive inbred strains, as well as the DBA/2J strain, displayed antidepressant-like responses to lithium in the FST after chronic administration of lithium. However, in the TST, acute lithium resulted in an antidepressant-like effect only in C3H/HeNHsd mice. Only C57BL/6J and DBA/2J showed an antidepressant-like response to lithium in the TST after chronic administration. ICV lithium administration resulted in a similar response profile in BALB/cJ (non-responsive) and C57BL/6J (responsive) strains. Serum and brain lithium concentrations showed that behavioral results were not because of differential pharmacokinetics of lithium in individual strains, suggesting that genetic factors likely regulate these behavioral responses to lithium. Our results indicate that antidepressant-like responses to lithium in tests of antidepressant efficacy varies among genetically diverse mouse strains. These results will assist in identifying genomic factors associated with lithium responsiveness and the mechanisms of lithium action.     

Genetic influences on exercise-induced adult hippocampal neurogenesis across 12 divergent mouse strains

New neurons are continuously born in the hippocampus of several mammalian species throughout adulthood. Adult neurogenesis represents a natural model for understanding how to grow and incorporate new nerve cells into preexisting circuits in the brain. Finding molecules or biological pathways that increase neurogenesis has broad potential for regenerative medicine. One strategy is to identify mouse strains that display large vs. small increases in neurogenesis in response to wheel running so that the strains can be contrasted to find common genes or biological pathways associated with enhanced neuron formation. Therefore, mice from 12 different isogenic strains were housed with or without running wheels for 43 days to measure the genetic regulation of exercise-induced neurogenesis. During the first 10 days mice received daily injections of 5-bromo-2'-deoxyuridine (BrdU) to label dividing cells. Neurogenesis was measured as the total number of BrdU cells co-expressing NeuN mature neuronal marker in the hippocampal granule cell layer by immunohistochemistry. Exercise increased neurogenesis in all strains, but the magnitude significantly depended on genotype. Strain means for distance run on wheels, but not distance traveled in cages without wheels, were significantly correlated with strain mean level of neurogenesis. Furthermore, certain strains displayed greater neurogenesis than others for a fixed level of running. Strain means for neurogenesis under sedentary conditions were not correlated with neurogenesis under runner conditions suggesting that different genes influence baseline vs. exercise-induced neurogenesis. Genetic contributions to exercise-induced hippocampal neurogenesis suggest that it may be possible to identify genes and pathways associated with enhanced neuroplastic responses to exercise.     

21株马特组镰刀菌遗传多样性的ISSR分析

为明确马特组镰刀菌种间和种内的遗传差异与亲缘关系,本文利用ISSR分子标记技术对21个马特组菌株进行了遗传多样性分析.结果表明:利用筛选出的15条引物对3种供试菌株进行扩增,共扩增出239条条带,其中多态性条带230条,多态性位点比例为96.2％,平均每条引物产生条带数为15.3条.21个菌株间的遗传相似系数范围为0.494～0.933,平均为0.640.在遗传相似系数为0.593时,供试的21株马特组镰刀菌可明显分成2个ISSR类群(IG),IG-Ⅰ包括1～17号菌株,为Fusarium solani和F solani var.coeruleum;IG-Ⅱ包括18 ～21号菌株,全部为F.ventricosum.在遗传相似系数为0.933时,供试的21个菌株可被全部区分开.供试的镰刀菌基因组在SSR区域具有丰富的多态性;ISSR类群划分与菌种分类之间存在一定相关性,但与菌株的地理来源没有相关性;而同一类群中,不同菌株之间的遗传相似性与菌株的地理来源存在一定的相关性.同一地区同种寄主的相同菌种,其菌株间也存在一定的遗传差异.     

专利微生物菌种的保藏与发放统计分析

对WIPO公开的"国际保藏机构2001～2010年专利微生物保藏与发放"数据进行了统计分析。结果表明:全世界的专利菌种保藏量与发放量处于快速增长中;美国是专利菌种保藏与发放量最多的国家;近年中国的专利菌种年保藏量实现三连冠,但专利菌种的发放率处于极低水平。     

Mucoid Helicobacter pylori isolates with fast growth under microaerobic and aerobic conditions

Background: Helicobacter pylori is microaerobic and turns into coccoid under aerobic conditions. In this study, two mucoid strains, A and D, were isolated from gastric biopsies which grew well on blood agar after 24‐hour incubation under aerobic as well as microaerobic conditions. The aim of this study was to identify these strains and compare their growth under aerobic and microaerobic conditions with that of control H. pylori. Materials and Methods: The two isolates A and D were identified as H. pylori according to microscopic morphology, urease, catalase and oxidase tests. Their growth under humidified aerobic and microaerobic conditions was compared with that of control H. pylori which grew only under microaerobic conditions. They were further identified by amplification of 16S rRNA, vacA alleles, cagA and ureAB genes by PCR. Their susceptibility to current antimicrobials was also examined. Results: The strains A and D produced mucoid colonies under aerobic and microaerobic conditions after 24‐hour, exhibiting the typical spiral morphology of H. pylori. The results of urease, catalase and oxidase tests were positive. Sequencing of amplified products showed 99–100% homology with those of the reference H. pylori strains in GenBank. Both strains exhibited resistance to the high concentrations of antimicrobials. Conclusions: This study reports the isolation of two mucoid strains of H. pylori with confluent growth under aerobic and microaerobic conditions. It appears that production of exopolysaccharide (EXP) could serve as a physical barrier to reduce oxygen diffusion into the bacterial cell and uptake of antibiotics. EXP protected the mucoid H. pylori isolates against stressful conditions, the result of which could be persistence of bacterial infection in the stomach.     

Host preference of Callosobruchus maculatus: a comparison of life history characteristics for three strains of beetles on two varieties of cowpea

Abstract: The reproductive success of Callosobruchus maculatus Fabricius, the main insect pest of stored cowpea, may vary between strains of this beetle and between varieties of the host seeds. Life history parameters of beetle strains from three different origins in West Africa were compared on two susceptible varieties of cowpea, Vigna unguiculata (L.) Walp. All beetle strains were assayed in a no‐choice and a two‐choice test. No major differences were found between the beetle strains. In a no‐choice situation, the developmental period from egg to adult was prolonged on the bean variety Kpodjiguegue. In a two‐choice situation, the beetles showed a strong preference for the Californian blackeyed bean variety to oviposit on. Here again the development took longer on Kpodjiguegue beans and the intrinsic rate of increase of the beetle population was lower. Using either equal numbers of beans of the same size or equal weights of beans of undetermined size of the two bean varieties did not affect the outcome of the test.