Central Haplotype Research Articles

First Histopathological and Molecular Characterization of Giant Thorny-headed Worm, Macracanthorhynchus hirudinaceus (Pallas, 1781) (Acanthocephala: Oligacanthorhynchidae) in Wild Boars, Sus scrofa Linnaeus, 1758 from Eastern Türkiye.

Macracanthorhynchus hirudinaceus (Pallas, 1781) is a zoonotic acanthocephalan that parasitizes the small intestine of wild boars. It is a pathogenic that causes economic losses, and poses a public health threat due to increased emergence. The aims of this study is describes histopathologically the damage caused by M. hirudinaceus in the small intestine of wild boar Sus scrofa Linnaeus, 1758, and molecularly characterize this parasite (sequences, haplotypes, phylogeny) for the first time in Elazig city, Türkiye. A high prevalence of infection was obtained. Upon separating the worms, it was discovered that there were ulcers resembling craters in the center, of the small intestine mucosa, surrounded by edema. The intestine wall where the parasite attached was damaged, with the villi epithelium and lamina propria in the mucosa being destroyed. The genomic DNA was isolated from all M. hirudinaceus samples, and PCR amplified the 489bp gene fragments were sequenced and confirmed that all 21 sequences were M. hirudinaceus. The haplotype analysis of the sequences revealed the presence of a central star-shaped haplotype, in addition to four other haplotypes. After conducting sequence analysis, the genetic differences between the M. hirudinaceus sequences obtained in this study and those reported from Europe and Japan suggest that this parasite is endemic to Türkiye's local wild boar population. Also, four haplotypes were identified, distinguishing it from other haplotypes by 1-5 mutation steps. It is essential to consider the worm's sequences and the formation of haplotypes, since these intrinsic characteristics may impact in the epidemiology and pathology of the worm in the future.

Symbiont infection and psyllid haplotype influence phenotypic plasticity during host switching events

Abstract Many herbivorous insect species exhibit phenotypic plasticity when using multiple hosts, which facilitates survival in heterogeneous host environments. Physiological host acclimation is an important part of it, yet the effects of host acclimation on insect feeding behaviour are not well studied, particularly for insect vectors of plant pathogens. We studied the combined effects of host acclimation and infection with a plant pathogenic symbiont on feeding behaviour of Bactericera cockerelli, an oligophagous psyllid widespread in both crop and natural habitats that feed primarily on Solanaceae and transmit an economically important plant pathogen, Candidatus Liberibacter solanacearum (CLso). We used a factorial design and the electrical penetration graphing technique to disentangle the effects of host acclimation, CLso infection and psyllid haplotype on the within‐plant feeding behaviour of B. cockerelli during conspecific and heterospecific host switches. This approach allows to connect phenotypic plasticity with the role of B. cockerelli as a vector by quantifying the frequency and duration of behaviours involved in CLso transmission. We found significant reductions in multiple metrics of B. cockerelli feeding efficiency, exacerbated by infection with CLso, which could lead to reduced transmission of this pathogen. Psyllid genotype was also important; the Central haplotype exhibited less dramatic changes in feeding efficiency than the Western haplotype during heterospecific host switches. Our study shows that host acclimation and heterospecific host switching directly alter feeding behaviours underlying pathogen transmission, and that the magnitude of feeding efficiency reductions depends on both host genotype and infection status.

Bacterial communities of the psyllid pest Bactericera cockerelli (Hemiptera: Triozidae) Central haplotype of tomato crops cultivated at different locations of Mexico.

The psyllid, Bactericera cockerelli, is an insect vector of 'Candidatus Liberibacter' causing "Zebra chip" disease that affects potato and other Solanaceae crops worldwide. In the present study, we analyzed the bacterial communities associated with the insect vector Bactericera cockerelli central haplotype of tomato crop fields in four regions from Mexico. PCR was used to amplify the mitochondrial cytochrome oxidase I gene (mtCOI) and then analyze the single nucleotide polymorphisms (SNP) and phylogenetic analysis for haplotype identification of the isolated B. cockerelli. Moreover, we carried out the microbial diversity analysis of several B. cockerelli collected from four regions of Mexico through the NGS sequencing of 16S rRNA V3 region. Finally, Wolbachia was detected by the wsp gene PCR amplification, which is the B. cockerelli facultative symbiont. Also we were able to confirm the relationship with several Wolbachia strains by phylogenetic analysis. Our results pointed that B. cockerelli collected in the four locations from Mexico (Central Mexico: Queretaro, and Northern Mexico: Sinaloa, Coahuila, and Nuevo Leon) were identified, such as the central haplotype. Analyses of the parameters of the composition, relative abundance, and diversity (Shannon index: 1.328 ± 0.472; Simpson index 0.582 ± 0.167), showing a notably relatively few microbial species in B. cockerelli. Analyses identified various facultative symbionts, particularly the Wolbachia (Rickettsiales: Anaplasmataceae) with a relative abundance higher. In contrast, the genera of Sodalis and 'Candidatus Carsonella' (Gammaproteobacteria: Oceanospirillales: Halomonadaceae) were identified with a relatively low abundance. On the other hand, the relative abundance for the genus 'Candidatus Liberibacter' was higher only for some of the locations analyzed. PCR amplification of a fragment of the gene encoding a surface protein (wsp) of Wolbachia and phylogenetic analysis corroborated the presence of this bacterium in the central haplotype. Beta-diversity analysis revealed that the presence of the genus 'Candidatus Liberibacter' influences the microbiota structure of this psyllid species. Our data support that the members with the highest representation in microbial community of B. cockerelli central haplotype, comprise their obligate symbiont, Carsonella, and facultative symbionts. We also found evidence that among the factors analyzed, the presence of the plant pathogen affects the structure and composition of the bacterial community associated with B. cockerelli.

A Preliminary mitochondrial cytochrome c oxidase-I-based phylogeographic and phylogenetic analysis of Eurasian Acanthocinus griseus (Coleoptera, Cerambycidae)

Acanthocinus griseus (Fabricius, 1792) (Coleoptera: Cerambycidae, Lamiinae, Acanthocinini) has long been known for its role in the decay process of the wood in the forest ecosystem, and two critical features of the species, inhabiting standing trees and being a vector of pine wood nematodes Bursaphelenchus spp., have been noted recently. Therefore, understanding the current relationships and possible migration scenarios has been further required to assess invasion risks. The present work provided a preliminary comprehension of the phylogenetic relationships of A. griseus based on the mitochondrial cytochrome c oxidase-I (COI) gene region (658 bp), with sequences produced in the present study, from the specimens collected from timberyards, ports and forests of Kocaeli Province, Turkey, and with available sequences in GenBank of inhabitants of Eurasia, and of intercepted specimens in ports. The intraspecific genetic distance of A. griseus was 1.37-0,3%, while the interspecific distance was 10,79-13,37%, except the closeness of an A. griseus haplotype (AGR1) to A. sachalinensis (0,3%) more than its conspecifics (4,71-5,47%). The ML and BI analyses suggested identical topologies. The statistical parsimony network drew a reticular branching diagram without grouping across countries or geographic regions, which addresses ongoing gene flow. Most haplotypes from Turkey were clustered around a central haplotype (AGR11), which may point to a bottleneck effect. A haplotype previously intercepted in USA ports was identical to a haplotype sampled in Kocaeli. The present study suggests that the relationship between A. griseus and A. sachalinensis should be reconsidered from both morphological and molecular points of view. In addition, the possible ongoing intraspecific gene flow within A. griseus might be due to facilitated migration by the international wood trade.

Genetic diversity of the polyphagous pest Rachiplusia nu across crops and locations in the Argentine Pampas

AbstractRachiplusia nu (Guenée) (Lepidoptera: Noctuidae, Plusiinae) is a defoliator of soybean (Glycine max L., Fabaceae), sunflower (Helianthus annuus L., Asteraceae), alfalfa (Medicago sativa L., Fabaceae), and horticultural crops in temperate southern South America; however, there is little genetic information available about this pest insect. Recently, concerns about this insect have increased because of its suspected resistance to Bt soybean and its spread to tropical regions. To better understand pest dynamics, and therefore implement suitable management strategies, a thorough knowledge of the genetic variability and spatial structuring of populations is required. In this paper, R. nu larvae were sampled from various crops and locations in central Argentina (where the pest is endemic) to assess diversity and population structure through mtDNA sequence analysis. In addition to the analysis of the standard cytochrome c oxidase subunit I (COI) ‘barcode’ fragment, a new set of primers was developed to explore genetic variation in the cytochrome b (Cytb) gene. COI and Cytb concatenated sequences revealed an overall high haplotype diversity (mean ± SD = 0.798 ± 0.037) and low nucleotide diversity (0.00137 ± 0.00013). The haplotype network showed a total of 19 haplotypes, separated by at most five mutational steps. A main central haplotype and four additional frequent haplotypes (accounting for 80% of the specimens examined) were distributed across populations with no obvious pattern related to host plant or geographic origin. Analysis of molecular variance further supported a weak population structure, where genetic variation within populations was the main source of total variation (ca. 96%). Pairwise comparisons showed only minor genetic differentiation among some of the populations studied. Implications of these findings in the context of R. nu control are discussed.

Acquisition and transmission of "Candidatus Liberibacter solanacearum" differs among Wolbachia-infected and -uninfected haplotypes of Bactericera cockerelli.

"Candidatus Liberibacter solanacearum" (Lso) causes disease symptoms and economic losses in potato, tomato, and other solanaceous crops in North America. Lso is transmitted to plants by potato psyllid, Bactericera cockerelli, which occurs as distinct haplotypes named western, central, and northwestern that differ in presence or absence of the bacterial endosymbiont, Wolbachia. Previous work showed that all three vector haplotypes can transmit Lso, but it was not clear whether acquisition and transmission rates of Lso were equal among the haplotypes. The goal of our study was to compare Lso infection rates among psyllids of the western, central, and northwestern haplotypes. Using data collected from several years of periodic testing of Lso infection of laboratory-reared potato psyllid colonies, we showed that psyllids of the western and central haplotypes are more likely to harbor Lso than are psyllids of the northwestern haplotype. We then used greenhouse assays to demonstrate that psyllids of the northwestern haplotype are less likely to acquire and transmit Lso compared with those of the western haplotype. Lso infection rates corresponded with Wolbachia infection among the three psyllid haplotypes. The Wolbachia-infected central and western haplotypes were more likely to harbor and transmit Lso compared with the Wolbachia-free northwestern haplotype. Results demonstrate that potato psyllids of the western and central haplotypes pose a greater risk for spread of Lso in crops and suggest a pattern between infection with Lso and Wolbachia in potato psyllid.

Mitochondrial Genetic Diversity of Eurasian Otter (<i>Lutra lutra</i>) from European Part of Russia and Transcaucasian Countries

In this study we examined mitochondrial DNA diversity of Eurasian otter from European part of Russia and Transcaucasian countries and compared it with other European populations. We used a fragment of mtDNA control region (255 bp) and also included previously detected haplotypes from NCBI. Six haplotypes were found in 75 samples from European part of Russia and Transcaucasian countries. Lut1 was the most common haplotype (62.1% of samples), Lut4 was detected in 17.6% of samples, other 4 haplotypes were newly detected. Haplotype diversity for European part of Russia and Transcaucasian countries were h = 0.56 ± 0.054, nucleotide diversity were π = 0.0016 ± 0.002. When we elongated the fragment of mtDNA (820 bp), haplotype (0.85 ± 0.03) and nucleotide diversity (π = 0.002 ± 0.001) were increased and 14 haplotypes were found. Geographic distribution of haplotypes depends neither on region, nor on river system. Central haplotype was detected throughout European part of Russia and Transcaucasian countries, as well as minor haplotypes, observed in lesser number of regions. Therefore, genetic diversity of Eurasian otters from European part of Russia and Transcaucasian countries is higher than in the rest of Europe. Besides, the population structure repeats European pattern with several regional features.

Genetic diversity of the stag beetle, Lucanus cervus (Insecta, Coleoptera) in Romania based on ND1 mitochondrial sequences

Lucanus cervus (Linnaeus, 1758) is one of the largest Coleoptera species in Europe and one of the most recognizable. It is a saproxylic species living in woodlands and wooded urban habitats and spending its larval stages inside tree trunks below or near soil level, feeding on decaying wood. The stag beetle has a wide distribution covering the entire Europe up to the Ural Mountains. In this paper, we attempted to describe the diversity and genetic structure of L. cervus populations in Romania using the NADH mitochondrial molecular marker. We identified 36 haplotypes arranged in a star-shaped structure with a central haplotype shared by many samples and linked by one or two mutational steps with other 35, represented by 1–3 samples. This pattern is frequently attributed to populations that have undergone recent range expansion, a phenomenon which is also sustained by the mismatch analysis. The analyzed populations revealed various degrees of genetic diversity. Many populations with relatively high levels of genetic diversity were found outside the Natura 2000 Network Special Areas of Conservation, while some populations included in protected areas proved to have low values of genetic variability. This underlines the need to review the designated protection sites for this species in Romania.

Health Equity in Sickle Cell Research and Access to Therapy

Health Equity in Sickle Cell Research and Access to Therapy

Phylogenetic Relationship and Molecular Divergence Dating Using SRY Gene Polymorphism about Four Ladoum Sheep Lineages in Senegal

Animal genetic resources are playing a vital role in livestock production and are essential to food security. The present study aims to contribute to a better understanding genetic local sheep breeds and to elucidate the phylogenetic relationships through the evolution of the SRY gene in four different lineages of Ladoum sheep raised in Senegal. After a brief analysis of genetic diversity, the phylogenetic relationships and molecular dating were inferred through haplotype networks and four phylogenetic reconstruction methods. The different haplotype networks are constructed with NETWORK ver. 5.0.0.0 using the Median-Joining method. Phylogenetic trees were reconstructed using neighbor-joining, maximum parsimony, maximum likelihood and Bayesian inference. The robustness of the nodes in phylogenetic trees of the three first methods was assessed by 1000 bootstraps. For Bayesian inference, the posterior probability distribution of the trees was estimated by 4 MCMC chains. 5,000,000 generations were performed for each of the chains by sampling the different parameters every 1000 generations. Results show a low polymorphism. Haplotypic diversity is much higher than the average nucleotide divergence between all pairs of haplotypes. The majority and central haplotype indicates a close relationship between “Batling” and “Tyson” individuals. “Birahim” lineage is very distinct from the rest. Phylogenetic trees confirm two genetically separate clades between “Birahim” and the other lineages. The period of divergence between “Birahim” lineage versus the common ancestor of the other three lineages was 2504 years ago. The polyphyly revealed in “Birahim” lindicates that this lineage does not contain the common ancestor of all individuals who compose it. It could therefore be derived from two or more sheep breeds with a common ancestor, Ovis aries. The monophyletic clade appears to be a group including a common ancestor and all of its genetic descendants. This group, bringing together the other three lineages, is in the process of being structured into sub-lineages. This study is the first to show that there are only two genetic lines within ladoum sheep in Senegal.

Molecular Diversity of Diplostomum spathaceum (Digenea: Diplostomidae) on the Capoeta umbla and Cyprinus carpio (Cypriniformes) Using Mitochondrial DNA Barcode

Diplostomid trematodes (Digenea) include a wide and diverse group of common digeneans. Diplostomid larval stages are significant pathogens that may exert grave effects on both natural fish and aquaculture populations. Diplostomum species, which use fish as a second intermediate host, is one of the most common trematode species, which affects the fish welfare negatively with the formation of cataracts by settling in the eye. This study is determined the molecular characterization of Diplostomum parasite in Cyprinus carpio and Capoeta umbla for the first time from Turkey, based on the mitochondrial COX1 sequence data. Diplostomum samples were determined as Diplostomum spathaceum according to the phylogenetic trees created in the light of the morphological and COX1 sequence data. Sequence results matched ~98-100% with D. spathaceum as a result of blast analysis. It was defined that this parasite was represented by three different haplotypes in Turkey. As a result of haplotype analysis performed on a total of 162 isolates in GenBank and obtained in this study, 40 polymorphic regions and 58 haplotypes were determined. This haplotype network had arranged within a star-like configuration with a main central haplotype. This shows that the variation within D. spathaceum species is quite high. The mean abundance, mean intensity, and prevalence of the parasite in C. carpio were 8.29 (5.80-11.24), 10.91±9.37 (1-39), and 76%, respectively. In the C umbla, mean abundance, mean intensity, and prevalence were calculated as 11.54 (10.16-12.92), 14.07±16.59 (1-67), and 82%, respectively.

Genetic diversity of the two-spotted stink bug Bathycoelia distincta (Pentatomidae) associated with macadamia orchards in South Africa.

The South African macadamia industry is severely affected by a complex of stink bugs, dominated by the two-spotted stink bug, Bathycoelia distincta Distant (Pentatomidae). This species was first discovered during the spring of 1984 in the Limpopo province. Although considerable effort has been spent trying to manage this pest, it continues to be a pest of concern for the macadamia industry. Information on the genetic diversity of this species is lacking, despite the potential relevance of such information for management strategies. The present study aimed to characterise the genetic diversity of B. distincta populations in South Africa. The Cytochrome c Oxidase Subunit 1 (COI) and cytochrome b (Cytb) gene regions were sequenced from individuals collected from the three main regions of macadamia production over three different seasons (2018-2020). An overall high haplotype diversity (COI = 0.744, Cytb = 0.549 and COI+Cytb = 0.875) was observed. Pairwise mean genetic distance between populations from each region varied from 0.2-0.4% in both datasets, which suggests the absence of cryptic species. The median joining network for both datasets consisted of one or two central haplotypes shared between the regions in addition to unique haplotypes observed in each region. Finally, low genetic differentiation (FST < 0.1), high gene flow (Nm > 1) and the absence of a correlation between genetic and geographic distance were estimated among populations. Overall, these results suggest that the B. distincta populations are not structured among the areas of macadamia production in South Africa. This might be due to its ability to feed and reproduce on various plants and its high dispersal (airborne) between the different growing regions of the country along with the rapid expansion of macadamia plantations in South Africa.

Bacterial Endosymbionts of Bactericera maculipennis and Three Mitochondrial Haplotypes of B. cockerelli (Hemiptera: Psylloidea: Triozidae)

Insects harbor bacterial endosymbionts that provide their hosts with nutritional benefit or with protection against natural enemies, plant defenses, insecticides, or abiotic stresses. We used directed sequencing of 16S rDNA to identify and compare endosymbionts of Bactericera maculipennis (Crawford) and the western, central, and northwestern haplotypes of B. cockerelli (Šulc) (Hemiptera: Psylloidea: Triozidae). Both species are native to North America, are known to harbor the plant pathogen 'Candidatus Liberibacter solanacearum' and develop on shared host plants within the Convolvulaceae. The Old-World species Heterotrioza chenopodii (Reuter) (Psylloidea: Triozidae), now found in North America, was included as an outgroup. 16S sequencing confirmed that both Bactericera species harbor 'Candidatus Liberibacter solanacearum' and revealed that both species harbor unique strains of Wolbachia and Sodalis. However, the presence of Wolbachia and Sodalis varied among haplotypes of B. cockerelli. The central and western haplotypes harbored the same strains of Wolbachia, which was confirmed by Sanger sequencing of the wsp and ftsZ genes. Wolbachia was also detected in very low abundance from the northwestern haplotype by high-throughput sequencing of 16S but was not detected from this haplotype by PCR screening. The northwestern and central haplotypes also harbored Sodalis, which was not detected in the western haplotype. Heterotrioza chenopodii harbored an entirely different community of potential endosymbionts compared with the Bactericera spp. that included Rickettsia and an unidentified bacterium in the Enterobacteriaceae. Results of this study provide a foundation for further research on the interactions between psyllids and their bacterial endosymbionts.

'Candidatus Liberibacter solanacearum' Infection of Physalis ixocarpa Brot. (Solanales: Solanaceae) in Saltillo, Mexico.

The potato psyllid Bactericera cockerelli (Šulc) (Hemiptera: Triozidae) is a pest of solanaceous crops (order Solanales), including potato (Solanum tuberosum L.) and tomato (S. lycopersicum L.). Feeding by high populations of nymphs causes psyllid yellows while adults and nymphs are vectors of the plant pathogen 'Candidatus Liberibacter solanacearum'. Foliar symptoms that were consistent with either 'Ca. L. solanacearum' infection or psyllid yellows were observed in 2019 on tomatillo (Physalis ixocarpa Brot.; family Solanaceae) grown within an experimental plot located near Saltillo, Mexico. This study had three primary objectives: 9i) determine whether the foliar symptoms observed on tomatillo were associated with 'Ca. L. solanacearum' infection, (ii) identify the haplotypes of 'Ca. L. solanacearum' and potato psyllids present in the symptomatic plot, and (iii) use gut content analysis to infer the plant sources of 'Ca. L. solanacearum'-infected psyllids. Results confirmed that 71% of symptomatic plants and 71% of psyllids collected from the plants were infected with 'Ca. L. solanacearum'. The detection of 'Ca. L. solanacearum' in plants and psyllids and the lack of nymphal populations associated with psyllid yellows strongly suggests that the observed foliar symptoms were caused by 'Ca. L. solanacearum' infection. All infected plants and insects harbored the more virulent 'Ca. L. solanacearum' haplotype B but one psyllid was also coinfected with haplotype A. The potato psyllids were predominantly of the central haplotype but one psyllid was identified as the western haplotype. Molecular gut content analysis of psyllids confirmed the movement of psyllids between noncrop habitats and tomatillo and indicated that 'Ca. L. solanacearum' infection of psyllids was associated with increased plant diversity in their diet.

Haplotyping the Potato Psyllid (Hemiptera: Triozidae) and the Associated Pathogenic Bacterium 'Candidatus Liberibacter solanacearum' in Non-crop Alternative Hosts in Southern Idaho.

Zebra chip, is a potato disease associated with the bacterium 'Candidatus Liberibacter solanacearum' (Lso) and vectored by the potato psyllid, Bactericera cockerelli Šulc. Potato psyllids are native to North America, where four haplotypes have been described. They are able to colonize a wide range of solanaceous species, crops, and weeds. The epidemiology of zebra chip disease is still poorly understood and might involve the different haplotypes of psyllids as well as two haplotypes of Lso. As several perennial weeds have been recognized as potential host for potato psyllids and Lso, a yearly monitoring of several patches of bittersweet nightshade (Solanum dulcamara) and field bindweed (Convolvulus arvensis) located in the potato-growing region of southern Idaho was conducted from 2013 to 2017, to gain insight into psyllid dynamics in non-potato hosts and Lso presence in the fields. Potato psyllids caught on each host were individually tested for Lso, and a subset were haplotyped based on the CO1 gene, along with the haplotyping of Lso in positive samples. On bittersweet nightshade, the Northwestern haplotype was numerically dominant, with around 2.7% of psyllids found to be carrying either Lso haplotype A or B, suggesting a limited role in zebra chip persistence, which has infected Idaho fields at a low occurrence since the 2012 outbreak. Field bindweed was found to be a transient, non-overwintering host for potato psyllid of Northwestern, Western and Central haplotypes late in the season, suggesting minor, if any, role in persistence of Lso and field infestation by potato psyllids.

Susceptibility of Physalis longifolia (Solanales: Solanaceae) to Bactericera cockerelli (Hemiptera: Triozidae) and 'Candidatus Liberibacter solanacearum'.

The potato psyllid, Bactericera cockerelli (Šulc), is a major pest of potato (Solanum tuberosum L.; Solanales: Solanaceae) as a vector of 'Candidatus Liberibacter solanacearum', the pathogen that causes zebra chip. Management of zebra chip is challenging in part because the noncrop sources of Liberibacter-infected psyllids arriving in potato remain unknown. Adding to this challenge is the occurrence of distinct genetic haplotypes of both potato psyllid and Liberibacter that differ in host range. Longleaf groundcherry (Physalis longifolia Nutt.) has been substantially overlooked in prior research as a potential noncrop source of Liberibacter-infected B. cockerelli colonizing fields of potato. The objective of this study was to assess the suitability of P. longifolia to the three common haplotypes of B. cockerelli (central, western, and northwestern haplotypes), and to two haplotypes of 'Ca. L. solanacearum' (Liberibacter A and B haplotypes). Greenhouse bioassays indicated that B. cockerelli of all three haplotypes produced more offspring on P. longifolia than on potato and preferred P. longifolia over potato during settling and egg-laying activities. Greenhouse and field trials showed that P. longifolia was also highly susceptible to Liberibacter. Additionally, we discovered that infected rhizomes survived winter and produced infected plants in late spring that could then be available for psyllid colonization and pathogen acquisition. Results show that P. longifolia is susceptible to both B. cockerelli and 'Ca. L. solanacearum' and must be considered as a potentially important source of infective B. cockerelli colonizing potato fields in the western United States.

High levels of population genetic differentiation in the American crocodile (Crocodylus acutus).

The American crocodile (Crocodylus acutus) is a widely distributed species across coastal and brackish areas of the Neotropical region of the Americas and the Greater Antilles. Available information on patterns of genetic differentiation in C. acutus shows a complex structuring influenced by interspecific interactions (mainly hybridization) and anthropogenic actions (mostly historical hunting, recent poaching, habitat loss and fragmentation, and unintentional translocation of individuals). In this study, we used data on mitochondrial DNA control region and 11 nuclear polymorphic microsatellite loci to assess the degree of population structure of C. acutus in South America, North America, Central America and the Greater Antilles. We used traditional genetic differentiation indices, Bayesian clustering and multivariate methods to create a more comprehensive picture of the genetic relationships within the species across its range. Analyses of mtDNA and microsatellite loci show evidence of a strong population genetic structure in the American crocodile, with unique populations in each sampling locality. Our results support previous findings showing large degrees of genetic differentiation between the continental and the Greater Antillean C. acutus. We report three new haplotypes unique to Venezuela, which are considerably less distant from the Central and North American haplotypes than to the Greater Antillean ones. Our findings reveal genetic population differentiation between Cuban and Jamaican C. acutus and offer the first evidence of strong genetic differentiation among the populations of Greater Antillean C. acutus.

First report of Echinococcus canadensis (G6/G7) by sequence analysis from the Khyber Pakhtunkhwa province of Pakistan

Cystic echinococcosis (CE) is a zoonotic parasitic disease that can result in human and animal health problems globally. Although the disease is known to be endemic in Asia and the Middle East, there are few epidemiological studies on CE in Pakistan. The purpose of the present study was to identify the Echinococcus granulosus sensu lato species and genotypes contributing to human CE cases in the Khyber Pakhtunkhwa (KPK) province of Pakistan. A total of fifty-six formalin fixed paraffin embedded (FFPE) CE cyst samples of human origin were collected from the Pathology Department, Rehman Medical Institute (RMI), KPK for the years 2012-2017. Cyst samples came from the liver (26/56; 46.4%), lungs (3/56; 5.3%), spleen (3/56; 5.3%), pelvis (1/56; 1.8%), breast (1/56; 1.8%), and thigh (1/56; 1.8%). The organ location for 21 of the cysts was not recorded. World Health Organization-Informal Working Group on Echinococcosis (WHO-IWGE) ultrasound-based cyst staging was available for 17 of the 26 (65.4%) hepatic cysts. Five of these cysts (29.4%) were CE3 (transitional), nine (52.9%) were CE4 (inactive), and three (17.6%) were CE5 (inactive). Most of the cysts were obtained from CE patients that were ethnically Afghan Pashtuns (44/56; 78.6%), while 12.5% (7/56) were from patients that were Pakistani Pashtuns. The majority (41/56; 73.2%) of patients reported having close interactions with dogs. Using 12SrRNA primers, 33 cyst samples were identified as being caused by E. granulosus sensu stricto (s.s.). Mitochondrially encoded cytochrome C oxidase 1 (mt-CO1) was evaluated for the remaining 23 samples. PCR product was obtained from six of these 23 samples. Of these six samples, one was identified as Echinococcus canadensis (G6/7). Haplotype analysis showed high haplotype and low nucleotide diversity for the mt-CO1 gene. There were 26 polymorphic sites for the mt-CO1 sequence, of which 65.3% (17/26) were parsimony informative. The E. canadensis mt-CO1 haplotype network consisted of 11 haplotypes, with a main central haplotype. In conclusion, it appears that E. granulosus s.s. and E. canadensis (G6/7) are circulating in the northwestern region of Pakistan. Further molecular epidemiological studies are needed to explore the local genetic diversity of the parasite.

Recovery from interglacial-related bottleneck likely triggered diversification of Lake Titicaca gastropod species flock

Ancient Lake Titicaca (Bolivia, Peru) is home to several endemic species flocks, including representatives of the caenogastropod genus Heleobia. Preliminary analyses indicated that past geological and limnological events had an important effect on intralacustrine speciation events in Heleobia spp. However, the role of allopatric speciation due to vicariance events has not been assessed in detail before, even though several biogeographic barriers are conceivable. This study therefore assesses evolutionary patterns in the Heleobia flock in space and time based on mitochondrial DNA sequencing data of 103 specimens from Lake Titicaca and the surrounding Altiplano. Specifically, the degree of genetic distinctness of populations at various geographical scales and the timing of population expansion events were inferred.A TCS network analysis indicated that many terminal haplotypes belong to Altiplano species, whereas central haplotypes are typically associated with Lake Titicaca taxa. AMOVAs based on geographical groupings resulted in a significant partitioning of variance in Altiplano vs. Titicaca taxa but not in taxa from the northern vs. southern basins of the lake. Mismatch analyses suggested that the Heleobia flock underwent sudden spatial and demographic expansions during the Middle/Late Pleistocene. Based on these findings, the working hypothesis of vicariant speciation has to be rejected. Instead, a Middle Pleistocene lake level lowstand (possibly interglacial MIS9), which resulted in the desiccation of the southern and the salinization of the northern basin, may have caused a severe bottleneck in Heleobia spp. Recovery from these limnological conditions might then have triggered sudden spatial and demographic expansion events.

Genetic variation in field voles (Microtus agrestis) from the British Isles: selective sweeps or population bottlenecks?

The Eurasian field vole (Microtus agrestis) comprises three evolutionarily significant units (ESUs). The northern ESU is found at higher latitudes across the western Palaearctic region and includes six, largely allopatric, mitochondrial DNA lineages that were derived from population bottlenecks. One of these lineages is found in southern Britain and nearby areas of continental Europe. A prominent sub-lineage is nested within, and therefore derived from, the part of this lineage occupying southern Britain. The sub-lineage consists of an abundant central haplotype together with a series of closely related haplotypes, a distribution that would result from either a recent population bottleneck or a selective sweep. To distinguish between these, we sequenced a Y-chromosome marker in 167 field voles from Britain and Europe, and analysed a panel of 13 autosomal microsatellite loci in 144 field voles from eight populations in Britain. The Y-chromosome marker showed a continental-scale pattern of variation that was not aligned with that of the mitochondrial marker, while microsatellite variation did not show any evidence for a bottleneck, tentatively favouring selection instead. This implies a role for both stochastic and selective processes in generating phylogeographical patterns at different scales in the field vole.