Prevalence and multilocus genotyping of Giardia duodenalis in zoo animals in three cities in China.

Giardia duodenalis is a flagellated parasitic microorganism that parasitizes in the intestines of humans and animals. Although asymptomatic infections commonly exist in both humans and animals, some enteric symptoms have been reported in immunocompromised individuals, posing a threat to public health. Children could be infected with G. duodenalis through an environment contaminated by infective animals. Thus, the investigation of the prevalence and genotypes of G. duodenalis in zoo animals is important. In this study, 672 fecal samples of 113 species of animals, including non-human primates, artiodactyla, perissodactyla, proboscidian, marsupial, birds, carnivora, and rodents, were collected from three zoos in Hangzhou city, Dalian city, and Suzhou city in China. The samples were screened for the positivity of G. duodenalis by nested PCR based on the ß-giardin (bg), triosephosphate isomerase (tpi), and glutamate dehydrogenase (gdh) gene loci. The overall G. duodenalis prevalence was 10.6% (71/672). The prevalence in non-human primates, artiodactyla, perissodactyla, proboscidian, marsupial, birds, carnivora, and rodent was 6.9% (10/144), 9.0% (12/133), 17.1% (6/35), 0% (0/6), 8.7% (2/23), 13.3% (28/211), 6.7% (7/105), and 40.0% (6/15), respectively. The region and category were considered risk factors for G. duodenalis infection in zoo animals (p < 0.001). Additionally, four genotypes of G. duodenalis were identified in zoo animals, including assemblage E (n = 46), assemblage A (n = 18), assemblage B (n = 6), and assemblage D (n = 1). The assemblages A, B, D, and E are also genotypes observed in humans and other animals. Among the sequences obtained in our study, one multilocus genotype (MLG) of the sub-assemblage AI was observed within assemblage A. Furthermore, three MLGs were detected within assemblage B. These findings reveal G. duodenalis genetic variability in zoo animals in three cities in China and suggest that zoo animals could be a potential source of human infection with G. duodenalis.

High Genetic Diversity in Predominantly Clonal Populations of the Powdery Mildew Fungus Podosphaera leucotricha from U.S. Apple Orchards.

Apple powdery mildew (APM), caused by Podosphaera leucotricha, is a constant threat to apple production worldwide. Very little is known about the biology and population structure of this pathogen in the United States and other growing regions, which affects APM management. A total of 253 P. leucotricha isolates, sampled from 10 apple orchards in Washington, New York, and Virginia, were genetically characterized with novel single sequence repeat and mating type markers. Eighty-three multilocus genotypes (MLGs) were identified, most of which were unique to a given orchard. Each isolate carried either a MAT1-1 or a MAT1-2 idiomorph at the mating type locus, indicating that P. leucotricha is heterothallic. Virulence tests on detached apple leaves showed that the 10 most frequent P. leucotricha MLGs were avirulent on a line containing a major resistance gene. Analysis of molecular variance showed significant differentiation (P < 0.001) among populations, a result supported by principal coordinate analysis revealing three genetic groups, each represented by nonoverlapping MLGs from Washington, New York, and Virginia. A Bayesian cluster analysis showed genetic heterogeneity between Washington populations, and a relative migration analysis indicated substantial gene flow among neighboring orchards. Random mating tests indicated that APM epidemics during the active cycle were dominated by clonal reproduction. However, the presence of sexual structures in orchards, the likelihood that five repeated MLGs resulted from sexual reproduction, and high genotypic diversity observed in some populations suggest that sexual spores play some role in APM epidemics. IMPORTANCE Understanding the population biology and epidemiology of plant pathogens is essential to develop effective strategies for controlling plant diseases. Herein, we gathered insights into the population biology of P. leucotricha populations from conventional and organic apple orchards in the United States. We showed genetic heterogeneity between P. leucotricha populations in Washington and structure between populations from different U.S. regions, suggesting that short-distance spore dispersal plays an important role in the disease's epidemiology. We presented evidence that P. leucotricha is heterothallic and that populations likely result from a mixed (i.e., sexual and asexual) reproductive system, revealing that the sexual stage contributes to apple powdery mildew epidemics. We showed that the major resistance gene Pl-1 is valuable for apple breeding because virulent isolates have most likely not emerged yet in U.S. commercial orchards. These results will be important to achieve sustainability of disease management strategies and maintenance of plant health in apple orchards.

Detection, Diversity, and Population Dynamics of Waterborne Phytophthora ramorum Populations.

Sudden oak death, the tree disease caused by Phytophthora ramorum, has significant environmental and economic impacts on natural forests on the U.S. west coast, plantations in the United Kingdom, and in the worldwide nursery trade. Stream baiting is vital for monitoring and early detection of the pathogen in high-risk areas and is performed routinely; however, little is known about the nature of water-borne P. ramorum populations. Two drainages in an infested California forest were monitored intensively using stream-baiting for 2 years between 2009 and 2011. Pathogen presence was determined both by isolation and polymerase chain reaction (PCR) from symptomatic bait leaves. Isolates were analyzed using simple sequence repeats to study population dynamics and genetic structure through time. Isolation was successful primarily only during spring conditions, while PCR extended the period of pathogen detection to most of the year. Water populations were extremely diverse, and changed between seasons and years. A few abundant genotypes dominated the water during conditions considered optimal for aerial populations, and matched those dominant in aerial populations. Temporal patterns of genotypic diversification and evenness were identical among aerial, soil, and water populations, indicating that all three substrates are part of the same epidemiological cycle, strongly influenced by rainfall and sporulation on leaves. However, there was structuring between substrates, likely arising due to reduced selection pressure in the water. Additionally, water populations showed wholesale mixing of genotypes without the evident spatial autocorrelation present in leaf and soil populations.

Multilocus genetic analysis of Trypanosoma cruzi supports non-domestic intrusion into domestic transmission in an endemic region of Colombia.

Trypanosoma cruzi, the causative agent of Chagas disease, is primarily transmitted to humans by hematophagous bugs of the Triatominae subfamily. In the Colombian Caribbean region, particularly on Margarita Island, T. cruzi transmission is highly endemic and associated with vectors such as Triatoma maculata and Rhodnius pallescens. Additionally, T. cruzi-infected Didelphis marsupialis are commonly found in close proximity to human dwellings. Given the complex transmission dynamics involving various domestic and non-domestic hosts, this study aimed to analyze 145 T. cruzi clones from twelve strains isolated from T. maculata, R. pallescens, and D. marsupialis using spliced leader intergenic region (SL-IR) sequences and nine polymorphic microsatellite loci. The results indicate the presence of a single polymorphic T. cruzi population, suggesting sustained local transmission dynamics between triatomines adapted to A. butyracea forests and peridomestic areas inhabited by synanthropic mammal reservoir such as D. marsupialis. Notably, this population appears to lack substructure, highlighting the importance of adopting an alternative eco-health approach to complement traditional chemical vector control methods for more effective and sustainable interruption of transmission.

Estimation of Genetic Diversity and Number of Unique Genotypes of Cassava Germplasm from Burkina Faso Using Microsatellite Markers.

Genetic diversity is very important in crop improvement. This study was carried out to assess the genetic diversity and the number of unique multilocus genotypes (MLGs) in a cassava collection in Burkina Faso. To achieve this objective, 130 cassava accessions were genotyped using 32 simple sequence repeat (SSR) markers. The results revealed that among these markers, twelve (12) were highly informative, with polymorphic information content (PIC) values greater than 0.50; twelve (12) were moderately informative, with PIC values ranging between 0.25 and 0.50; and eight (8) were not very informative, with PIC values lower than 0.25. A moderate level of genetic diversity was found for the population, indicated by the average expected heterozygosity (0.45) and the observed heterozygosity (0.48). About 83.8% of unique multilocus genotypes were found in the cassava collection, indicating that SSR markers seem to be most appropriate for MLG identification. Population structure analysis based on hierarchical clustering identified two subpopulations and the Bayesian approach suggested five clusters. Additionally, discriminant analysis of principal components (DAPC) separated the cassava accessions into 13 subpopulations. A comparison of these results and those of a previous study using single nucleotide polymorphisms (SNP) suggests that each type of marker can be used to assess the genetic structure of cassava grown in Burkina Faso.

Genetic diversity of the submerged macrophyte Ceratophyllum demersum depends on habitat hydrology and habitat fragmentation.

The adaptability of plant populations to a changing environment depends on their genetic diversity, which in turn is influenced by the degree of sexual reproduction and gene flow from distant areas. Aquatic macrophytes can reproduce both sexually and asexually, and their reproductive fragments are spread in various ways (e.g. by water). Although these plants are obviously exposed to hydrological changes, the degree of vulnerability may depend on the types of their reproduction and distribution, as well as the hydrological differences of habitats. The aim of this study was to investigate the genetic diversity of the cosmopolitan macrophyte Ceratophyllum demersum in hydrologically different aquatic habitats, i.e. rivers and backwaters separated from the main river bed to a different extent. For this purpose, the first microsatellite primer set was developed for this species. Using 10 developed primer pairs, a high level of genetic variation was explored in C. demersum populations. Overall, more than 80% of the loci were found to be polymorphic, a total of 46 different multilocus genotypes and 18 private alleles were detected in the 63 individuals examined. The results demonstrated that microsatellite polymorphism in this species depends on habitat hydrology. The greatest genetic variability was revealed in populations of rivers, where flowing water provides constant longitudinal connections with distant habitats. The populations of the hydrologically isolated backwaters showed the lowest microsatellite polymorphism, while plants from an oxbow occasionally flooded by the main river had medium genetic diversity. The results highlight that in contrast to species that spread independently of water flow or among hydrologically isolated water bodies, macrophytes with exclusive or dominant hydrochory may be most severely affected by habitat fragmentation, for example due to climate change.

Giardia duodenalis in Hu sheep: occurrence and environmental contamination on large-scale housing farms.

Giardia duodenalis is a common zoonotic intestinal parasitic protozoan and sheep are among its hosts; however, limited information is available on sheep kept in large-scale housing. The Hu sheep is a first-class protected local livestock breed in China. In this study, we investigated the seasonal dynamics of G. duodenalis infection in Hu sheep and the environmental contamination of large-scale sheep farms. We collected 474 fecal samples and 312 environmental samples from Hu sheep on a large-scale sheep farm in Henan, China. The prevalence of G. duodenalis was determined by nested PCR targeting the ß­giardin (bg) gene. The assemblages and multilocus genotypes (MLGs) were investigated based on analyses of three genetic loci, i.e. bg, glutamate dehydrogenase (gdh), and triosephosphate isomerase (tpi). To detect mixed infections of different assemblages, assemblage A/E-specific PCRs were performed to amplify the tpi gene. The prevalence of G. duodenalis infection in sheep was 17.9% (81/474) and the positivity rate in environmental samples was 0.96% (3/312). Genetic analysis revealed the presence of two assemblages (assemblages A and E), with assemblage E being detected in both fecal and environmental samples, and assemblage A detected only in fecal samples. A total of 23 MLGs were obtained in fecal and environmental samples, all of which belonged to assemblage E. These results indicate the seasonal dynamics of G. duodenalis infection in sheep and environmental contamination on large-scale housing sheep farms and provide an important reference for the prevention and control of G. duodenalis on large-scale housing sheep farms.

Title: Giardia duodenalis chez les moutons Hu : occurrence et contamination environnementale dans les fermes d'élevage à grande échelle. Abstract: Giardia duodenalis est un protozoaire parasitaire intestinal zoonotique commun et les moutons font partie de ses hôtes, mais peu d'informations sont disponibles sur les moutons élevés dans des stabulations à grande échelle. Le mouton Hu est une race de bétail locale protégée de première classe en Chine. Dans cette étude, nous avons étudié la dynamique saisonnière de l'infection à G. duodenalis chez les moutons Hu et la contamination environnementale des élevages ovins à grande échelle. Nous avons collecté 474 échantillons fécaux et 312 échantillons environnementaux de moutons Hu dans une ferme ovine à grande échelle dans le Henan, en Chine. La prévalence de G. duodenalis a été déterminée par PCR nichée ciblant le gène de la ß­giardine (bg). Les assemblages et les génotypes multilocus (MLG) ont été étudiés sur la base de l'analyse de trois locus génétiques, à savoir bg, glutamate déshydrogénase (gdh) et triosephosphate isomérase (tpi). Pour détecter des infections mixtes de différents assemblages, des PCR spécifiques aux assemblages A et E ont été réalisées pour amplifier le gène tpi. La prévalence de l'infection à G. duodenalis chez les ovins était de 17,9 % (81/474) et le taux de positivité dans les échantillons environnementaux était de 0,96 % (3/312). L'analyse génétique a révélé la présence de deux assemblages (assemblages A et E), l'assemblage E étant détecté à la fois dans les échantillons fécaux et environnementaux, et l'assemblage A détecté uniquement dans les échantillons fécaux. Au total, 23 MLG ont été détectés dans des échantillons fécaux et environnementaux, tous appartenant à l'assemblage E. Ces résultats montrent la dynamique saisonnière de l'infection à G. duodenalis chez les ovins et la contamination environnementale dans les élevages ovins à grande échelle et fournissent une référence importante pour la prévention et le contrôle de G. duodenalis dans les élevages ovins à grande échelle.

Epistatic interactions in idiopathic pulmonary arterial hypertension.

BACKGROUND: Idiopathic pulmonary arterial hypertension (IPAH) is a poorly understood complex disorder, which results in progressive remodeling of the pulmonary artery that ultimately leads to right ventricular failure. A two-hit hypothesis has been implicated in pathogenesis of IPAH, according to which the vascular abnormalities characteristic of PAH are triggered by the accumulation of genetic and/or environmental insults in an already existing genetic background. The multifactor dimensionality reduction (MDR) analysis is a statistical method used to identify gene-gene interaction or epistasis and gene-environment interactions that are associated with a particular disease. The MDR method collapses high-dimensional genetic data into a single dimension, thus permitting interactions to be detected in relatively small sample sizes. AIM: To identify and characterize polymorphisms/genes that increases the susceptibility to IPAH using MDR analysis. MATERIALS AND METHODS: A total of 77 IPAH patients and 100 controls were genotyped for eight polymorphisms of five genes (5HTT, EDN1, NOS3, ALK-1, and PPAR-γ2). MDR method was adopted to determine gene-gene interactions that increase the risk of IPAH. RESULTS: With MDR method, the single-locus model of 5HTT (L/S) polymorphism and the combination of 5HTT(L/S), EDN1(K198N), and NOS3(G894T) polymorphisms in the three-locus model were attributed to be the best models for predicting susceptibility to IPAH, with a P value of 0.05. CONCLUSION: MDR method can be useful in understanding the role of epistatic and gene-environmental interactions in pathogenesis of IPAH.

Prevalence and multilocus genotyping of Giardia duodenalis in Holstein cattle in Yunnan, China.

Giardia duodenalis is an important zoonotic protozoon, which can infect a variety of animals, causing diarrhea and even death of animals or humans. Dairy cattle have been implicated as important sources of human G. duodenalis. However, the information about the prevalence and genetic diversity of G. duodenalis in dairy cattle in China's Yunnan Province remains limited. This study investigated the occurrence and multilocus genotyping of G. duodenalis of Holstein cattle in Yunnan Province, China. A total of 524 fresh fecal samples of Holstein cattle were randomly collected from 8 farms in Yunnan. In this study, 27.5% (144/524) of tested samples were positive for G. duodenalis infection. The highest infection ratio was found in preweaned calves (33.7%), and the infection rates of postweaned calves, growing cattle, and adult cattle were 24.5%, 23.0%, and 17.3%, respectively. The sequence analysis of SSU rRNA gene showed that the predominant assemblage of G. duodenalis in this study was assemblage E (97.9%, 141/144), whereas assemblage A was identified only in three samples (2.1%, 3/144). All G. duodenalis-positive samples were further assayed with nested polymerase chain reaction (PCR) targeting ß-giardin (bg), triosephosphate isomerase (tpi), and glutamate dehydrogenase (gdh) genes, and 87, 41, and 81 sequences were obtained, respectively. Mixed infection of assemblages A and E of G. duodenalis was detected in three samples. Multilocus genotyping yielded 23 multilocus genotypes (MLGs). This is the first study that reveals the prevalence data of G. duodenalis in Holstein cattle in Yunnan Province, and the results of this study provided baseline data for the prevention and control of G. duodenalis infection in Holstein cattle in Yunnan Province, China.

A high genetic diversity of Enterocytozoon bieneusi in diarrheic pigs in southern China.

Enterocytozoon bieneusi is an important pathogen that is responsible for over 90% of documented cases of human microsporidiosis worldwide, causing a threat to public health and husbandry development. In immunocompromised patients, it can cause persistent diarrhoea, wasting diathesis and malabsorption and developing life-threatening chronic diarrhoea. However, there was little information on the prevalence and multilocus genotypes of E. bieneusi in diarrheic pigs in three provinces of southern China. In this study, 1254 faecal samples of diarrheic pigs were collected from 37 pig farms in Hunan, Jiangxi, and Fujian provinces in southern China, and were investigated the prevalence and genotypes of E. bieneusi by nested polymerase chain reaction (PCR) based on the internal transcribed spacer (ITS) region of the nuclear ribosomal DNA gene. The overall prevalence of E. bieneusi was 5.7% (72/1254) in three provinces. Furthermore, the difference was statistically significant (p < 0.001) in the prevalence of E. bieneusi in age groups. ITS sequence analysis revealed that 13 E. bieneusi genotypes were identified, including 8 known genotypes (EbpC, n = 30; Henan-IV, n = 21; CH5, n = 6; EbpA, n = 3; KIN-1, n = 2; O, n = 1; GX3, n = 1; CHS5, n = 1) and 5 novel genotypes (JX1, n = 2; JX2, n = 1; JX3, n = 2; FJ1, n = 1; FJ2, n = 1), and the genotype EbpC was the preponderant genotype. Phylogenetic analysis indicated that all genotypes of E. bieneusi were clustered as the zoonotic group 1. Moreover, a high genetic diversity of E. bieneusi were identified in this study, which the 64, 57, 52 and 64 samples were identified by multilocus sequence typing (MLST) at MS1, MS3, MS4 and MS7 loci, respectively. Then, 45 samples were successfully amplified and sequenced at four loci, forming 41 distinct multilocus genotypes (MLGs). These findings suggest that diarrheic pigs may potentially threaten to transmit E. bieneusi to humans, revealing E. bieneusi genetic variability in diarrheic pigs in three provinces of southern China.

"Guess Who's Coming to Dinner": Molecular Tools to Reconstruct multilocus Genetic Profiles from Wild Canid Consumption Remains.

Non-invasive genetic sampling is a practical tool to monitor pivotal ecological parameters and population dynamic patterns of endangered species. It can be particularly suitable when applied to elusive carnivores such as the Apennine wolf (Canis lupus italicus) and the European wildcat (Felis silvestris silvestris), which can live in overlapping ecological contexts and sometimes share their habitats with their domestic free-ranging relatives, increasing the risk of anthropogenic hybridisation. In this case study, we exploited all the ecological and genetic information contained in a single biological canid faecal sample, collected in a forested area of central Italy, to detect any sign of trophic interactions between wolves and European wildcats or their domestic counterparts. Firstly, the faecal finding was morphologically examined, showing the presence of felid hair and claw fragment remains. Subsequently, total genomic DNA contained in the hair and claw samples was extracted and genotyped, through a multiple-tube approach, at canid and felid diagnostic panels of microsatellite loci. Finally, the obtained individual multilocus genotypes were analysed with reference wild and domestic canid and felid populations to assess their correct taxonomic status using Bayesian clustering procedures. Assignment analyses classified the genotype obtained from the endothelial cells present on the hair sample as a wolf with slight signals of dog ancestry, showing a qi = 0.954 (C.I. 0.780-1.000) to the wolf cluster, and the genotype obtained from the claw as a domestic cat, showing a qi = 0.996 (95% C.I. = 0.982-1.000) to the domestic cat cluster. Our results clearly show how a non-invasive multidisciplinary approach allows the cost-effective identification of both prey and predator genetic profiles and their taxonomic status, contributing to the improvement of our knowledge about feeding habits, predatory dynamics, and anthropogenic hybridisation risk in threatened species.

Geographical pattern of genetic diversity in Capsella bursa-pastoris (Brassicaceae)-A global perspective.

We analyzed the global genetic variation pattern of Capsella bursa-pastoris (Brassicaceae) as expressed in allozymic (within-locus) diversity and isozymic (between-locus) diversity. Results are based on a global sampling of more than 20,000 C. bursa-pastoris individuals randomly taken from 1,469 natural provenances in the native and introduced range, covering a broad spectrum of the species' geographic distribution. We evaluated data for population genetic parameters and F-statistics, and Mantel tests and AMOVA were performed. Geographical distribution patterns of alleles and multilocus genotypes are shown in maps and tables. Genetic diversity of introduced populations is only moderately reduced in comparison with native populations. Global population structure was analyzed with structure, and the obtained cluster affiliation was tested independently with classification approaches and macroclimatic data using species distribution modeling. Analyses revealed two main clusters: one distributed predominantly in warm arid to semiarid climate regions and the other predominantly in more temperate humid to semihumid climate regions. We observed admixture between the two lineages predominantly in regions with intermediate humidity in both the native and non-native ranges. The genetically derived clusters are strongly supported in macroclimatic data space. The worldwide distribution patterns of genetic variation in the range of C. bursa-pastoris can be explained by intensive intra- and intercontinental migration, but environmental filtering due to climate preadaption seems also involved. Multiple independent introductions of genotypes from different source regions are obvious. "Endemic" genotypes might be the outcome of admixture or of de novo mutation. We conclude that today's successfully established Capsella genotypes were preadapted and found matching niche conditions in the colonized range parts.

Multilocus Genotyping and Intergenic Spacer Single Nucleotide Polymorphisms of Amylostereum areolatum (Russulales: Amylostereacea) Symbionts of Native and Non-Native Sirex Species.

Sirex noctilio along with its mutualistic fungal symbiont, Amylostereum areolatum (a white rot fungus), is an invasive pest that causes excessive damage to Pinus plantations in Northeast China. In 2015, S. noctilio were found to attack Pinus sylvestris var. mongolica, and often share larval habitat with the native woodwasp, S. nitobei. The objective of this study was to determine the possible origin(s) of the introduced pest complex in China and analyse the genetic diversity between A. areolatum isolated from invasive S. noctilio, native S. nitobei and other woodwasps collected from Europe (native range) and other countries. Phylogenetic analyses were performed using the intergenic spacer (IGS) dataset and the combined 4-locus dataset (the internal transcribed spacer region (ITS), translation elongation factor alpha 1 (tef1), DNA-directed ribosomal polymerase II (RPB2), and mitochondrial small subunit (mtSSU)) of three Amylostereum taxa. The multilocus genotyping of nuclear ribosomal regions and protein coding genes revealed at least three distinct multilocus genotypes (MLGs) of the fungus associated with invasive S. noctilio populations in Northeast China, which may have come from North America or Europe. The IGS region of A. areolatum carried by S. noctilio from China was designated type B1D2. Our results showed a lack of fidelity (the paradigm of obligate fidelity to a single fungus per wasp species) between woodwasp hosts and A. areolatum. We found that the native S. nitobei predominantly carried A. areolatum IGS-D2, but a low percentage of females instead carried A. areolatum IGS-B1D2 (MLG A13), which was presumably due to horizontal transmission from S. noctilio, during the sequential use of the same wood for larval development. The precise identification of the A. areolatum genotypes provides valuable insight into co-evolution between Siricidae and their symbionts, as well as understanding of the geographical origin and history of both Sirex species and their associated fungi.

Genetic Diversity and Conservation Status of Helianthus verticillatus, an Endangered Sunflower of the Southern United States.

Evaluating species diversity and patterns of population genetic variation is an essential aspect of conservation biology to determine appropriate management strategies and preserve the biodiversity of native plants. Habitat fragmentation and potential habitat loss are often an outcome of a reduction in naturally occurring wildfires and controlled prescribed burning, as seen in Helianthus verticillatus (whorled sunflower). This endangered, wild relative of the common sunflower, Helianthus annuus, is endemic to four locations in Alabama, Georgia, and Tennessee, United States. Despite its endangered status, there is no recovery plan for H. verticillatus, and knowledge related to its basic plant biology and importance in ecosystem services is mostly unknown. In this study, we utilized 14 microsatellite loci to investigate fine-scale population structure and genetic diversity of H. verticillatus individuals found on two sampling sites within the Georgia population. Our results indicated moderate genetic diversity and the presence of two distinct genetic clusters. Analyses of molecular variance indicated that the majority of variance was individually based, thus confirming high genetic differentiation and limited gene flow between H. verticillatus collection sites. The evidence of a population bottleneck in these sites suggests a recent reduction in population size that could be explained by habitat loss and population fragmentation. Also, high levels of linkage disequilibrium were detected, putatively suggesting clonal reproduction among these individuals. Our study provides a better understanding of fine-scale genetic diversity and spatial distribution of H. verticillatus populations in Georgia. Our results can underpin an original recovery plan for H. verticillatus that could be utilized for the conservation of this endangered species and to promote its persistence in the wild.

Asexual reproduction of a few genotypes favored the invasion of the cereal aphid Rhopalosiphum padi in Chile.

BACKGROUND: Aphids (Hemiptera: Aphididae) are insects with one of the highest potentials for invasion. Several aphid species are present globally due to introduction events; they represent important pests of agroecosystems. The bird cherry-oat aphid Rhopalosiphum padi (Linnaeus) is a major pest of cereals and pasture grasses worldwide. Here, we report the genetic features of populations of R. padi that colonize different cereal crops in central Chile. METHODS: Rhopalosiphum padi individuals were collected in central Chile and genotyped at six microsatellite loci. The most frequent multilocus genotype (MLG) was then studied further to assess its reproductive performance across cereal hosts under laboratory conditions. RESULTS: Populations of R. padi in Chile are characterized by a low clonal diversity (G/N = 62/377 = 0.16) and the overrepresentation of a few widely distributed MLGs. One of the MLGs constituted roughly half of the sample and was observed in all sampled populations at high frequencies. Furthermore, this putative aphid "superclone" exhibited variations in its reproductive performance on cereals most commonly cultivated in Chile. The sampled populations also exhibited weak signs of genetic differentiation among hosts and localities. Our findings suggest that (1) obligate parthenogenesis is the primary reproductive mode of R. padi in Chile in the sampled range and (2) its introduction involved the arrival of a few genotypes that multiplied asexually.

Occurrence and multilocus genotyping of Giardia duodenalis in black-boned sheep and goats in southwestern China.

BACKGROUND: Giardia duodenalis is an important intestinal protozoan infecting both humans and animals, causing significant public health concern and immeasurable economic losses to animal husbandry. Sheep and goats have been reported as common reservoirs of G. duodenalis, but only a limited amount of information is available for native breeds of these small ruminants in China. The present study investigated the prevalence and multilocus genotypes of G. duodenalis in black-boned sheep and goats, two important native breeds in Yunnan Province, southwestern China. METHODS: Fecal samples were collected from 336 black-boned goats and 325 black-boned sheep from five counties (Meishui, Shanshu, Shilin, Yongsheng and Nanping) of Yunnan Province and the genomic DNA was extracted from these feces. The prevalence of G. duodenalis was determined by the nested PCR targeting the ß-giardin (bg) gene. The assemblages and multilocus genotypes (MLGs) were investigated based on analyses of three genetic loci, i.e. bg, glutamate dehydrogenase (gdh) and triosephosphate isomerase (tpi). RESULTS: Giardia duodenalis infection was detected in both black-boned sheep and goats, and the prevalence of G. duodenalis in black-boned sheep (21.8%, 71/325) was significantly higher (χ2 = 36.63, df = 1, P < 0.001) than that in black-boned goats (4.8%, 16/336). Significant differences in prevalence were also observed in goats and sheep from different counties (χ2 = 39.83, df = 4, P < 0.001) and age groups (χ2 = 97.33, df = 3, P < 0.001). Zoonotic assemblage A and animal-specific assemblage E were identified in both black-boned sheep and goats with the latter as the predominant assemblage. Based on sequences obtained from the three genetic loci (bg, gdh and tpi), 16 MLGs were obtained in black-boned sheep and goats, including 15 MLGs in assemblage E and one MLG in assemblage A. CONCLUSIONS: Our results not only extended the host range of G. duodenalis, but also revealed high genetic variations in G. duodenalis assemblages. The findings of the present study also provide baseline data for preventing and controlling G. duodenalis infection in black-boned sheep and goats in Yunnan Province.

Microgeographic variation in recruitment under adult trees: arrival of new genotypes or perpetuation of the existing ones?

Investigating spatial variation in the relative importance of sexual reproduction and clonal propagation is critical to obtain more accurate estimates of future effective population sizes and genetic diversity, as well as to identify ecological correlates of clonality. We combined a stratified sampling scheme with microsatellite genetic analyses to estimate variation in the proportion of sexual versus clonal recruits among saplings in five populations of the tree Pyrus bourgaeana. Using a likelihood framework, we identified clones among the genotypes analysed and examined variation among populations regarding the proportion of saplings coming from clonal propagation. We also examined the relationship between the relative abundance of clonal shoots across the studied populations and their herbivory levels. Our results revealed that one third of the saplings examined (N = 225 saplings) had a probability above 0.9 of being clones of nearby (<10 m) trees, with the ratio between clonal propagation and sexual recruitment varying up to eight-fold among populations. A small portion of these putative clonal shoots reached sexual maturity. Relative abundance of clonal shoots did not significantly relate to the herbivory by ungulates. Our results call into question optimistic expectations of previous studies reporting sufficient levels of recruitment under parental trees without animal seed dispersal services. Nevertheless, given that some of these clonal shoots reach sexual maturity, clonal propagation can ultimately facilitate the long-term persistence of populations during adverse periods (e.g. environmental stress, impoverished pollinator communities, seed dispersal limitation).

cloncase: Estimation of sex frequency and effective population size by clonemate resampling in partially clonal organisms.

Inferring reproductive and demographic parameters of populations is crucial to our understanding of species ecology and evolutionary potential but can be challenging, especially in partially clonal organisms. Here, we describe a new and accurate method, cloncase, for estimating both the rate of sexual vs. asexual reproduction and the effective population size, based on the frequency of clonemate resampling across generations. Simulations showed that our method provides reliable estimates of sex frequency and effective population size for a wide range of parameters. The cloncase method was applied to Puccinia striiformis f.sp. tritici, a fungal pathogen causing stripe/yellow rust, an important wheat disease. This fungus is highly clonal in Europe but has been suggested to recombine in Asia. Using two temporally spaced samples of P. striiformis f.sp. tritici in China, the estimated sex frequency was 75% (i.e. three-quarter of individuals being sexually derived during the yearly sexual cycle), indicating strong contribution of sexual reproduction to the life cycle of the pathogen in this area. The inferred effective population size of this partially clonal organism (Nc  = 998) was in good agreement with estimates obtained using methods based on temporal variations in allelic frequencies. The cloncase estimator presented herein is the first method allowing accurate inference of both sex frequency and effective population size from population data without knowledge of recombination or mutation rates. cloncase can be applied to population genetic data from any organism with cyclical parthenogenesis and should in particular be very useful for improving our understanding of pest and microbial population biology.

Geographical segregation of Cryptosporidium parvum multilocus genotypes in Europe.

Cryptosporidium parvum is a common enteric protozoan pathogen of humans and livestock. Multilocus genotyping based on simple sequence repeat polymorphisms has been used extensively to identify transmission cycles and to investigate the structure of C. parvum populations and of the related pathogen Cryptosporidiumhominis. Using such methods, the zoonotic transmission of C. parvum has been shown to be epidemiologically important. Because different genetic markers have been used in different surveys, the comparison of Cryptosporidium genotypes across different laboratories is often not feasible. Therefore, few comparisons of Cryptosporidium populations across wide geographical areas have been published and our understanding of the epidemiology of cryptosporidiosis is fragmented. Here we report on the genotypic analysis of a large collection of 692 C. parvum isolates originating primarily from cattle and other ruminants from Italy, Ireland and Scotland. Because the same genotypic markers were used in these surveys, it was possible to merge the data. We found significant geographical segregation and a correlation between genetic and geographic distance, consistent with a model of isolation by distance. The presence of strong LD and positive IA(S) values in the combined MLG dataset suggest departure from panmixia, with different population structures of the parasite prevailing in each country.

DifferInt: compositional differentiation among populations at three levels of genetic integration.

In many fields of study, it is important to know how different populations are genetically. Commonly used measures such as FST and its derivatives based on gene identity probabilities do not reliably reflect difference, as they can be maximal when almost all populations are identical and very small when populations are completely distinct. Compositional differentiation, in contrast, is maximal only for completely distinct populations. Moreover, underlying forces of fragmentation that act on single- and multilocus genotypes may be missed if only the allelic level is viewed. For these reasons, descriptive measures of compositional differentiation based on untransformed distributions of genetic types at different levels of genetic integration (alleles, single- and multilocus genotypes) were derived. Here, two measures of complementary differentiation and two new measures of dispersive differentiation are described. One of each considers genic differences between individuals, ensuring their consistency across integration levels. An increase from one level to the next indicates differences among the populations in their forms of gene association. The computer program DifferInt calculates these measures for one or more codominantly expressed gene loci at the gene-pool, single-locus and multilocus levels. Effects of gene pools and gene association on differentiation are compared by permutation analysis. Snail diagrams depict the contribution of each population to differentiation. Results are demonstrated using a numerical example. The applicability of the measures calculated by this program in conservation, phylogeography, mating system analysis and adaptation studies is discussed.