Quantifying genotyping errors in noninvasive population genetics

The use of noninvasively collected samples greatly expands the range of ecological issues that may be investigated through population genetics. Furthermore, the difficulty of obtaining reliable genotypes with samples containing low quantities of amplifiable DNA may be overcome by designing optimal genotyping schemes. Such protocols are mainly determined by the rates of genotyping errors caused by false alleles and allelic dropouts. These errors may not be avoided through laboratory procedure and hence must be quantified. However, the definition of genotyping error rates remains elusive and various estimation methods have been reported in the literature. In this paper we proposed accurate codification for the frequencies of false alleles and allelic dropouts. We then reviewed other estimation methods employed in hair- or faeces-based population genetics studies and modelled the bias associated with erroneous methods. It is emphasized that error rates may be substantially underestimated when using an erroneous approach. Genotyping error rates may be important determinants of the outcome of noninvasive studies and hence should be carefully computed and reported.     

Recombination and the origin of sequence diversity in the DRB MHC class II locus in chamois (Rupicapra spp.)

We examined the evolutionary processes contributing to genetic diversity at the major histocompatibility complex (MHC) class II DRB locus in chamois (Rupicapra spp., subfamily Caprinae). We characterised the pattern of intragenic recombination (or homologous gene conversion) and quantified the amount of recombination in the genealogical history of the two chamois species, Pyrenean chamois (Rupicapra pyrenaica) and Alpine chamois (Rupicapra rupicapra). We found evidence for intragenic recombination, and the estimated amount of population recombination suggests that recombination has been a significant process in generating DRB allelic diversity in the genealogical history of the genus Rupicapra. Moreover, positive selection appears to act on the same peptide-binding residues in both analysed chamois species, but not in identical intensity. Recombination coupled with positive selection drives the rapid evolution at the peptide-binding sites in the MHC class II DRB gene. Many chamois MHC class II DRB alleles are thus much younger than previously assumed.     

Microsatellite genotyping errors: detection approaches, common sources and consequences for paternal exclusion

Microsatellite genotyping errors will be present in all but the smallest data sets and have the potential to undermine the conclusions of most downstream analyses. Despite this, little rigorous effort has been made to quantify the size of the problem and to identify the commonest sources of error. Here, we use a large data set comprising almost 2000 Antarctic fur seals Arctocephalus gazella genotyped at nine hypervariable microsatellite loci to explore error detection methods, common sources of error and the consequences of errors on paternal exclusion. We found good concordance among a range of contrasting approaches to error-rate estimation, our range being 0.0013 to 0.0074 per single locus PCR (polymerase chain reaction). The best approach probably involves blind repeat-genotyping, but this is also the most labour-intensive. We show that several other approaches are also effective at detecting errors, although the most convenient alternative, namely mother-offspring comparisons, yielded the lowest estimate of the error rate. In total, we found 75 errors, emphasizing their ubiquitous presence. The most common errors involved the misinterpretation of allele banding patterns (n = 60, 80%) and of these, over a third (n = 22, 36.7%) were due to confusion between homozygote and adjacent allele heterozygote genotypes. A specific test for whether a data set contains the expected number of adjacent allele heterozygotes could provide a useful tool with which workers can assess the likely size of the problem. Error rates are also positively correlated with both locus polymorphism and product size, again indicating aspects where extra effort at error reduction should be directed. Finally, we conducted simulations to explore the potential impact of genotyping errors on paternity exclusion. Error rates as low as 0.01 per allele resulted in a rate of false paternity exclusion exceeding 20%. Errors also led to reduced estimates of male reproductive skew and increases in the numbers of pups that matched more than one candidate male. Because even modest error rates can be strongly influential, we recommend that error rates should be routinely published and that researchers make an attempt to calculate how robust their analyses are to errors.     

用超显性模型估计遗传多态平衡的种群参数

基于超显性模型,采用数学方法阐述了种群遗传多样性的延续机制及平衡条件,从双等位基因导出的定律有较大的局限性,在许多教科书中常用的估计平衡态基因频数的定律只适用于双等位基因,本文用另一种方法导出一些公式,将其扩展到多等位基因,讨论了种群中基因数ｎ,遗传负载荷（Ｌ）,杂合频ＹＸ（Ｈｅ）和纯合频率（Ｈｏｍ）之间的关系。     

A multilevel latent Markov model for the evaluation of nursing homes' performance

The periodic evaluation of health care services is a primary concern for many institutions. We consider services provided by nursing homes with the aim of ranking a set of these structures with respect to their effect on resident health status. Since the overall health status is not directly observable, and given the longitudinal and multilevel structure of the available data, we rely on latent variable models and, in particular, on a multilevel latent Markov model where residents and nursing homes are the first and the second level units, respectively. The model includes individual covariates to account for resident characteristics. The impact of nursing home membership is modelled through a pair of random effects affecting the initial distribution and the transition probabilities between different levels of health status. Through the prediction of these random effects we obtain a ranking of the nursing homes. Furthermore, the proposed model accounts for nonignorable dropout due to resident death, which typically occurs in these contexts. The motivating dataset is gathered from the Long Term Care Facilities programme, a health care protocol implemented in Umbria (Italy). Our results show that differences in performance between nursing homes are statistically significant.     

Population genetic structure of the tropical moss Acanthorrhynchium papillatum as measured with microsatellite markers

Mosses and other bryophytes are vital components of forests, because they sustain a tremendous diversity of invertebrates and influence significant ecological functions. There have been few studies on moss population diversity in Southeast Asia, despite the escalating deforestation in this region of rich biodiversity. The genetic diversity of the tropical moss Acanthorrhynchium papillatum (Harv.) Fleisch., collected from forested areas in Singapore and Peninsular Malaysia, was elucidated using eight microsatellite markers developed for this species. Significant levels of allelic and haplotypic diversity were observed among clumps of the moss. Differences in allelic richness and genotypic diversity among the populations were higher in less disturbed forests compared to the more disturbed areas, suggesting that genetic diversity is affected by habitat quality. Genetic diversity levels within the clumps studied were low, indicating that vegetative reproduction was more important within clumps than sexual reproduction. However, multilocus genotypes of samples within the clumps studied were not all alike, providing evidence of microsatellite mutation or of occasional sexuality. Despite the isolation of populations, A. papillatum can introduce genetic variability by mutation among vegetatively propagated individuals. This study provides baseline information on the genetic diversity of A. papillatum tropical rain forests.     

Polymorphism in the acetylcholinesterase gene of Musca domestica L. field populations in Turkey

Acetylcholinesterase (AChE), encoded by the Ace gene, is the primary target of organophosphates (OPs) and carbamates (CBs) in insects. Ace mutations have been identified in OP and CB resistant strains of Musca domestica. In this study, the Ace gene was partially amplified and sequenced at amino acid positions 260, 342, and 407 to determine the frequencies of these mutations in housefly samples collected from farms and garbage disposal sites of 16 provinces in the Aegean and Mediterranean regions of Turkey. In addition, the percent remaining AChE activities in these samples were assayed by using three OPs (malaoxon, paraoxon, and dichlorvos) and one CB (carbaryl) compound as inhibitors. In all the analyzed samples, 13 different combinations at the three amino acid positions were identified and the L/V260-A/G342-F/Y407 combination was found in the highest frequency. No susceptible individual was detected. The highest mean percent remaining AChE activities were detected in the individuals having the L260-A/G342-F/Y407 genotype when malaoxon and paraoxon were used as inhibitors and in the individuals with the L260-A342-F/Y407 combination when dichlorvos and carbaryl were used as inhibitors. The obtained data were heterogeneous and there was no exact correlation between the molecular genetic background and the resistance phenotypes of the flies. The findings of this study at the molecular and biochemical levels indicate the presence of significant control problems in the field.     

Parasitism and maintenance of diversity in a fungal vegetative incompatibility system: the role of selection by deleterious cytoplasmic elements

In fungi, horizontal transmission of deleterious cytoplasmic elements is reduced by the vegetative incompatibility system. This self/non-self recognition system may select for greater diversity of fungal incompatibility phenotypes in a frequency-dependent manner but the link between the diversity of fungal phenotypes and the virulence of cytoplasmic parasites has been poorly studied. We used an epidemiological model to show that even when transmission between incompatibility types is permitted, parasite pressure can lead to high levels of polymorphism for vegetative incompatibility systems. Moreover, high levels of polymorphism in host populations can select for less virulent cytoplasmic parasites. This feedback mechanism between parasite virulence and vegetative incompatibility system polymorphism of host populations may account for the general avirulence of most known mycoviruses. Furthermore, this mechanism provides a new perspective on the particular ecology and evolution of the host/parasite interactions acting between fungi and their cytoplasmic parasites.     

Description of microsatellite markers and genotyping performances using feathers and buccal swabs for the Ivory gull (Pagophila eburnea)

We report 22 new polymorphic microsatellites for the Ivory gull (Pagophila eburnea), and we describe how they can be efficiently co-amplified using multiplexed polymerase chain reactions. In addition, we report DNA concentration, amplification success, rates of genotyping errors and the number of genotyping repetitions required to obtain reliable data with three types of noninvasive or nondestructive samples: shed feathers collected in colonies, feathers plucked from living individuals and buccal swabs. In two populations from Greenland (n=21) and Russia (Severnaya Zemlya Archipelago, n=21), the number of alleles per locus varied between 2 and 17, and expected heterozygosity per population ranged from 0.18 to 0.92. Twenty of the markers conformed to Hardy-Weinberg and linkage equilibrium expectations. Most markers were easily amplified and highly reliable when analysed from buccal swabs and plucked feathers, showing that buccal swabbing is a very efficient approach allowing good quality DNA retrieval. Although DNA amplification success using single shed feathers was generally high, the genotypes obtained from this type of samples were prone to error and thus need to be amplified several times. The set of microsatellite markers described here together with multiplex amplification conditions and genotyping error rates will be useful for population genetic studies of the Ivory gull.