Genetic evidence for a complex of species within Rugopharynx australis (Mönnig, 1926) (Nematoda: Stronglyloidea) from macropodid marsupials

An electrophoretic analysis using 17 enzyme loci was carried out on specimens of the gastric nematode of macropodid marsupials, Rugopharynx australis (Mönnig, 1926), collected from Macropus eugenii (Desmarest), M. fuliginosus (Desmarest), M. giganteus Shaw, M. robustus Gould, M. rufogriseus (Desmarest), M. rufus (Desmarest), Thylogale billardierii (Desmarest) and Wallabia bicolor (Desmarest) from south-eastern Australia. The extent of fixed genetic differences between nematodes from different host species ranged from 0–53%. The two distinct morphological forms of the parasite found in M. rufogriseus differed at 50% of loci. Specimens present in M. fuliginosus and M. giganteus were indistinguishable genetically, as were nematodes from M. rufus and M. robustus. Of the two morphologically distinct congeners included in the analysis as controls, Rugopharynx epsilon (Johnston & Mawson, 1939) was genetically distinct (46–69% fixed genetic differences) from all specimens of the R. australis complex while R. rufogrisea Magzoub, 1964 was closely related to one of the two species occuring in M. rufogriseus. It was concluded that R. australis is a species complex, with a genetically distinct species present in M. eugenii, M. fuliginosus/M. giganteus, M. robustus/M. rufus, W. bicolor and T. billardierii, and two species in M. rufogriseus.     

An electrophoretic and morphological analysis of Labiostrongylus (Labiomultiplex) uncinatus (Nematoda: Cloacinidae), with the description of a new species, L. contiguus, from Macropus parryi (Marsupialia: Macropodidae)

An electrophoretic study was conducted on Labiostrongylus (Labiomultiplex) uncinatus, nematodes that occur in the stomachs of two species of Australian macropodid, Macropus dorsalis and M. parryi. The allelic profiles of these nematodes were compared to those of a morphologically distinct species, L. (Lm.) billardierii, which infests Thylogale billardierii. Nematodes were genetically characterized at 17 enzymes encoding a presumptive 18 loci. The results revealed the existence of two species, one in M. dorsalis and the other in M. parryi, that had fixed genetic differences at 72% of loci. This level of fixed difference between these species is equivalent to that when each is compared to L. billardierii (87–89%). The new species in M. parryi, Labiostrongylus contiguus n. sp., is described herein. A morphological comparison with L. uncinatus revealed slight but consistent differences in the morphology of their anterior end; namely, rectangular rather than conical shaped lateral lips, small inconspicuous, not larger hook-shaped cephalic papillae, and convex rather than flat floor of the buccal capsule for L. (Lm.) contiguus as compared with L. (Lm.) uncinatus. Differentiation of L. contiguus from L. uncinatus is more clearly demonstrated by biochemical characters than morphological ones.     

A morphological and electrophoretic study of Rugopharynx zeta (Johnston & Mawson, 1939) (Nematoda: Strongyloidea), with the description of a new species,R. mawsonae,from the black-striped wallaby Macropus dorsalis (Marsupialia: Macropodidae)

Rugopharynx zeta (Johnston & Mawson) (Nematoda: Strongyloidea) is redescribed from the rock wallabies Petrogale penicillata penicillata, P. p. herberti, P. inornata and P. assimilis from Queensland and New South Wales, Australia. Specimens formerly assigned to this nematode taxon from the wallabies Macropus dorsalis and M. parma are treated as a new species, R. mawsonae. R. mawsonae n. sp. differs from R. zeta in the shape of the dorsal ray, length of spicules, morphology of spicule tip, length of female tail and position of deirid. The morphological differences are supported by electrophoretic data. R. zeta and R. mawsonae n. sp. had fixed genetic differences at 45.0% of the 21 enzyme loci examined, while each differed at 38.1% and 45.0% of loci respectively from the morphologically distinct R. delta (Johnston & Mawson). The known host and geographical distributions of R. zeta and R. mawsonae n. sp. are reviewed.     

Genetic structure of populations of Norway spruce (<Emphasis Type="Italic">Picea abies</Emphasis> (L.) Karst. from Ukrainian Carpathians

The genetic diversity, subdivision, and differentiation of nine populations of Norway spruce (Picea abies (L.) Karst.) in Ukrainian Carpathians were studied using electrophoretic analysis of variability of enzyme systems in 346 trees aged from 80 to 150 years. Based on electrophoretic fractionation of enzymes extracted from seed endosperms in vertical slabs of 7.5% polyacrylamide gel, 20 loci of nine enzyme systems (ADH, ACP, DIA, GDH, GOT, MDH, LAP, FDH, SOD) were identified, and 71 allele variant were revealed. Each tree was heterozygous on average in 15.8% of its genes. The populations were characterized by low subdivision (F _ST = 0.017) and differentiation (D _N = 0.005). The main contribution to heterogeneity of population genetic structure was made by loci Dia-3, Lap-1, and Sod-3. Clustering and multivariate analysis revealed no observed trends in geographical or altitudinal position of the populations.     

Differentiation of taeniid cestodes by enzyme electrophoresis

Le Riche P.D. and Sewell M.M.H. 1978. Differentiation of taeniid cestodes by enzyme electrophoresis. International Journal for Parasitology8: 479–483. Interspecific differences were shown by enzyme electrophoresis in thin starch gel between Echinococcus granulosus (from horses), Taenia hydatigena, T. multiceps, T. ovis, T. pisiformis, T. saginata, T. solium and T. taeniaeformis, Differences were seen with adenylate kinase, glucose phosphate isomerase, glutamate dehydrogenase, hexokinase and malate dehydrogenase. Multiple molecular forms of glucose phosphate isomerase were clear and most species could be differentiated by this enzyme alone.No differences attributable to anthelmintic effect were seen and zymogram patterns were identical for adult and cystic forms of each species. No differences were seen between specimens collected from various geographical locations.     

Allozyme variation in bank vole,Myodes glareolus (Mammalia: Rodentia) in Northern Anatolia

A total of 94 specimens from 16 populations of Myodes glareolus, collected between 2004 and 2007, from different altitudinal distributions were analyzed, using 16 enzyme systems. We found that 10 out of 22 loci (Idh-2, α-Gpdh, Me, Pgm, Pgd, Mdh-s, Ada, Est-1, Ldh-1, and Ldh-2) were polymorphic. Group 1 included population from altitudes ranging from 27 to 605 m above sea level (ASL), and Group 2 were from altitudes ranging from 1003 to 1288 m ASL. The summaries of the genetic parameters also displayed differences between the 2 groups. The possible reasons of such fragmentation between M. glareolus populations were discussed.     

Microsatellite variation and microevolution in the critically endangered San Clemente Island loggerhead shrike (Lanius ludovicianus mearnsi)

Polymorphic nuclear microsatellite loci were used to characterize genetic variation in contemporary and historic populations of the San Clemente Island loggerhead shrike (Lanius ludovicianus mearnsi), an endangered bird with a current population of 30 individuals that is endemic to to one of the California Channel Islands. We also compared the population of the shrike with two contemporary populations of the still abundant subspecies, L. l. gambeli, which live 120 km away on the adjacent mainland. The current population of L. l. mearnsi has 60 per cent of the genetic variation of the mainland shrike populations and is strongly differentiated from them. Comparison of living birds with 19 birds collected in 1915 shows that most of the variation within the island population was lost before the recent 90 per cent decline in population size, and the 20 per cent decrease in variation this century is probably attributable to genetic drift. Mitochondrial DNA control region sequence data from 80 year old specimens show that there may have been limited introgression to L. l. mearnsi, this century, from another island subspecies, L. l. anthonyi, found in the northern Channel Islands. Today, gene flow between L. l. mearnsi and mainland L. l. gambel is very low, even though a few mainland birds visit the island annually. The island subspecies population has evolved sufficient genetic independence to justify ongoing conservation efforts to counter demographic collapse and genetic erosion; the course of genetic erosion can now be monitored non-invasively, as demonstrated by this study, based on DNA amplified from feathers.     

Dispersal of the Japanese Pine Sawyer,Monochamus alternatus (Coleoptera: Cerambycidae), in Mainland China as Inferred from Molecular Data and Associations to Indices of Human Activity

The Japanese pine sawyer, Monochamus alternatus Hope (Coleoptera: Cerambycidae), is an important forest pest as well as the principal vector of the pinewood nematode (PWN), Bursaphelenchus xylophilus (Steiner et Buhrer), in mainland China. Despite the economic importance of this insect-disease complex, only a few studies are available on the population genetic structure of M. alternatus and the relationship between its historic dispersal pattern and various human activities. The aim of the present study was to further explore aspects of human activity on the population genetic structure of M. alternatus in mainland China. The molecular data based on the combined mitochondrial cox1 and cox2 gene fragments from 140 individuals representing 14 Chinese populations yielded 54 haplotypes. Overall, a historical (natural) expansion that originated from China’s eastern coast to the western interior was revealed by the haplotype network, as well as several recent, long-distant population exchanges. Correlation analysis suggested that regional economic status and proximity to marine ports significantly influenced the population genetic structure of M. alternatus as indicated by both the ratio of shared haplotypes and the haplotype diversity, however, the PWN distribution in China was significantly correlated with only the ratio of shared haplotypes. Our results suggested that the modern logistical network (i.e., the transportation system) in China is a key medium by which humans have brought about population exchange of M. alternatus in mainland China, likely through inadvertent movement of infested wood packaging material associated with trade, and that this genetic exchange was primarily from the economically well-developed east coast of China, westward, to the less-developed interior. In addition, this study demonstrated the existence of non-local M. alternatus in new PWN-infested localities in China, but not all sites with non-local M. alternatus were infested with PWN.     

Comparison of habitat selection by two sympatric macropods,Thylogale billardierii and Macropus rufogriseus rufogriseus,in a patchy eucalypt‐forestry environment

Abstract Population density estimates and patterns of habitat selection by sympatric red‐bellied pademelons (Thylogale billardierii (Marsupialia: Macropodidae)) and red‐necked wallabies (Macropus rufogriseus rufogriseus (Marsupialia: Macropodidae)) were examined within a patchy forestry environment in north‐west Tasmania. Population density of both species was relatively high. Selection indices from both population surveys and animal movement data showed that T. billardierii and M. rufogriseus had similar patterns of habitat selection at two spatio‐temporal scales; home range within the study area and habitats selected while foraging at night. Both species selected for young Eucalyptus nitens plantation with high weed‐cover within their home range. At night, T. billardierii and M. rufogriseus selected for open habitats (young plantation and grassland) and avoided closed habitats (native forest and 5–7 years old E. nitens plantation). There was no evidence for resource partitioning between species at these scales. In contrast, the two species differed in their selection for daytime sheltering habitat; T. billardierii selected native forest while M. rufogriseus selected older plantation. This may reflect differences in their predator avoidance strategies; that is, crypsis versus flight, rather than resource partitioning as a result of interspecific competition. The environment appears to be of high quality for both species, with patches of feeding and shelter habitats within close proximity of one another.     

GENETICS OF INCIPIENT SPECIATION IN DROSOPHILA MOJAVENSIS. III. LIFE‐HISTORY DIVERGENCE IN ALLOPATRY AND REPRODUCTIVE ISOLATION

We carried out a three‐tiered genetic analysis of egg‐to‐adult development time and viability in ancestral and derived populations of cactophilic Drosophila mojavensis to test the hypothesis that evolution of these life‐history characters has shaped premating reproductive isolation in this species. First, a common garden experiment with 11 populations from Baja California and mainland Mexico and Arizona reared on two host species revealed significant host plant X region and population interactions for viability and development time, evidence for host plant adaptation. Second, replicated line crosses with flies reared on both hosts revealed autosomal, X chromosome, cytoplasmic, and autosome X cactus influences on development time. Viability differences were influenced by host plants, autosomal dominance, and X chromosomal effects. Many of the F₁, F₂, and backcross generations showed evidence of heterosis for viability. Third, a QTL analysis of male courtship song and epicuticular hydrocarbon variation based on 1688 Baja × mainland F₂ males also revealed eight QTL influencing development time differences. Mainland alleles at six of these loci were associated with longer development times, consistent with population‐level differences. Eight G × E interactions were also detected caused by longer development times of mainland alleles expressed on a mainland host with smaller differences among Baja genotypes reared on the Baja host plant. Four QTL influenced both development time and epicuticular hydrocarbon differences associated with courtship success, and there was a significant QTL‐based correlation between development time and cuticular hydrocarbon variation. Thus, the regional shifts in life histories that evolved once D. mojavensis invaded mainland Mexico from Baja California by shifting host plants were genetically correlated with variation in cuticular hydrocarbon‐based mate preferences.     

Post-Translational Modification as a Potential Explanation of High Levels of Enzyme Polymorphism: Xanthine Dehydrogenase and Aldehyde Oxidase in DROSOPHILA MELANOGASTER

Xanthine dehydrogenase (XDH) and aldehyde oxidase (AO) in Drosophila melanogaster require for their activity the action of another unlinked locus, maroon-like (mal). While the XDH and AO loci are on chromosome 3, mal maps to the X chromosome. Although functional mal gene product is required for XDH and AO activity, it is possible to examine the effects of mutant mal alleles in those cases when pairs of mutants complement to produce a partial restoration of activity. To test whether mal mediates a post-translational modification of the XDH and AO proteins, we constructed several mal heteroallelic complementing stocks of Drosophila in which the third chromosomes were co-isogenic. Since all lines were co-isogenic for the XDH and AO structural genes, any variation in these enzymes seen when comparing these stocks must have been produced by post-translational modification by mal. We examined the XDH and AO proteins in these stocks by gel-sieving electrophoresis, a procedure that permits independent characterization of a protein's charge and shape, and is capable of discriminating many variants not detected in routine electrophoresis. In every mal heteroallelic combination, there is a significant alteration in protein shape, when compared to wild type. The magnitude of differences in shape of XDH and AO is correlated both with differences in their enzyme activities and with differences in their thermal stabilities. As the body of this variation appears heritable, any functional differences resulting from these variants are of real genetic and evolutionary interest. A similar post-translational modification of XDH and AO by yet another locus, lxd, was subsequently documented in an analogous manner. The pattern of electrophoretic differences produced by mal and lxd modification is similar to that reported for electrophoretic "alleles" of XDH in natural populations. The implication is that heritable variation in electrophoretic mobility at these two enzyme loci, and potentially at other loci, is not necessarily allelic to the structural gene loci.     

Wing morphometrics as a possible tool for the diagnosis of the Ceratitis fasciventris,C. anonae,C. rosa complex (Diptera,Tephritidae)

Previous attempts to resolve the Ceratitis FAR complex (Ceratitis fasciventris, Ceratitis anonae, Ceratitis rosa, Diptera, Tephritidae) showed contrasting results and revealed the occurrence of five microsatellite genotypic clusters (A, F1, F2, R1, R2). In this paper we explore the potential of wing morphometrics for the diagnosis of FAR morphospecies and genotypic clusters. We considered a set of 227 specimens previously morphologically identified and genotyped at 16 microsatellite loci. Seventeen wing landmarks and 6 wing band areas were used for morphometric analyses. Permutational multivariate analysis of variance detected significant differences both across morphospecies and genotypic clusters (for both males and females). Unconstrained and constrained ordinations did not properly resolve groups corresponding to morphospecies or genotypic clusters. However, posterior group membership probabilities (PGMPs) of the Discriminant Analysis of Principal Components (DAPC) allowed the consistent identification of a relevant proportion of specimens (but with performances differing across morphospecies and genotypic clusters). This study suggests that wing morphometrics and PGMPs might represent a possible tool for the diagnosis of species within the FAR complex. Here, we propose a tentative diagnostic method and provide a first reference library of morphometric measures that might be used for the identification of additional and unidentified FAR specimens.     

Genetic variation within the ticks Ixodes holocyclus and Ixodes cornuatus from south-eastern Australia

Ticks from mainland Australia (Victoria, New South Wales and Queensland) and Tasmania, identified morphologically as either Ixodes holocyclus or Ixodes cornuatus, were compared genetically using 24 enzyme loci. The results showed that ticks from three localities in Victoria were genetically similar to I. cornuatus in Tasmania, but both groups had fixed genetic differences at >45% of loci compared with other ticks on the mainland. In addition, there were fixed genetic differences at 0-60% of loci among I. holocyclus from different localities on the mainland. Ixodes holocyclus samples could be divided into four distinct clusters (with fixed genetic differences >15%), three of which were represented by one or two specimens. Nonetheless, these electrophoretic data suggest that I. holocyclus represents a species complex. The results also showed that the morphological criteria used to identify specimens were not always accurate because several specimens had been mis-identified morphologically. Despite limitations with the morphological identification, this study has demonstrated that I. cornuatus can be distinguished from the I. holocyclus species complex using six enzyme loci, providing the foundation for a re-examination of morphological characteristics. The present study has shown that I. cornuatus and the I. holocyclus complexes have a greater distribution than previously reported, with both occurring in sympatry at Cape Patterson, on the southern coastline of Victoria.     

Isozyme variability in species of the genus Drosophila. I. A multiple allelic isozyme system in Drosophila busckii: Inheritance and general considerations

Starch gel electrophoretic analysis of a triallelic leucine aminopeptidase polymorphism in a laboratory population of Drosophila busckii is described. The three alleles involved are expressed without dominance. A series of single-pair matings revealed an excess of heterozygous types in most segregating families, suggesting selection against the homozygous genotypes. A few cases of heterogeneity among progeny extracted from a single family were the result of matings that produced Mendelian ratios. These few cases had no clear genetic explanation, but there is a suggestion of two isoalleles for the electrophoretically intermediate enzyme.The research reported here was begun at the University of Hawaii and completed at the University of Texas and was supported (in part) by Public Health Service Research Grant No. GM 11609 to W. S. Stone and M. R. Wheeler and by Training Grant No. 2 T1-GM-337-06 and GM 00337-07 to R. P. Wagner et al., from the National Institutes of Health.     

Genetic Diversity among Rhizobium leguminosarum bv. Trifolii Strains Revealed by Allozyme and Restriction Fragment Length Polymorphism Analyses

Allozyme electrophoresis and restriction fragment length polymorphism (RFLP) analyses were used to examine the genetic diversity of a collection of 18 Rhizobium leguminosarum bv. trifolii, 1 R. leguminosarum bv. viciae, and 2 R. meliloti strains. Allozyme analysis at 28 loci revealed 16 electrophoretic types. The mean genetic distance between electrophoretic types of R. leguminosarum and R. meliloti was 0.83. Within R. leguminosarum, the single strain of bv. viciae differed at an average of 0.65 from strains of bv. trifolii, while electrophoretic types of bv. trifolii differed at a range of 0.23 to 0.62. Analysis of RFLPs around two chromosomal DNA probes also delineated 16 unique RFLP patterns and yielded genetic diversity similar to that revealed by the allozyme data. Analysis of RFLPs around three Sym (symbiotic) plasmid-derived probes demonstrated that the Sym plasmids reflect genetic divergence similar to that of their bacterial hosts. The large genetic distances between many strains precluded reliable estimates of their genetic relationships.     

Morphological Differences Between Radopholus citrophilus and R. similis

SEM observations of the external morphology of populations of Radopholus citrophilus and R. similis revealed several diagnostic differences. The cloaco-spicular orifice on males of R. citrophilus had three to seven genital papillae (anterior hypoptygmata), whereas males of R. similis were either smooth or had one or two shorter genital papillae (anterior hypoptygmata). Females of R. citrophilus had four annules in the region of the vulval opening, but R. similis had five annules in the same region. The labial disc and lateral lips appeared to be of diagnostic significance, but these areas were more susceptible to artifacts due to fixation. An unknown population of Radopholus from Puerto Rico with a chromosome number of n = 4 was morphologically similar to R. similis. These morphological differences provide additional support that R. citrophilus and R. similis are distinct species.     

Studies on Anoplotaenia dasyuri beddard, 1911 (Cestoda: Taeniidae), a parasite of the Tasmanian devil: Life-cycle and epidemiology

Anoplotaenia dasyuri was found commonly in Tasmanian devils (Sarcophilus harrisi) and was also found in a tiger cat (Dasyurus maculatus), in feral cats (Felis catus) and in rural dogs in Tasmania. The Bennett's wallaby (Macropus rufogriseus) was a suitable experimental intermediate host, metacestodes localising in the heart, lungs and skeletal muscles. Natural infections were detected in pademelons (Thylogale billardieri) and potoroos (Potorous apicalis) as well as in M. rufogriseus. Metacestodes from wallabies developed into gravid worms when fed to Tasmanians devils and to a tiger cat but failed to develop in dogs. The development in Tasmanian devils is described.     

Enzyme gene heterozygosity in small island populations of Philaenus spumarius (L.) (Homoptera)

Genetic polymorphism at about twenty enzyme loci in one mainland and in six differentially isolated island populations of Philaenus spumarius (L.) was studied by starch gel electrophoresis. The populations have different average degrees of heterozygosity, so that the most isolated population is the least polymorphic. The differences in heterozygosity seem to be correlated to the size of the population and the degree of isolation from other populations. With a single exception, the most common allele in each locus is the most common one everywhere. The results are compared with the differences observed in the color polymorphism of Philaenus island populations. The allele frequencies of enzyme loci are maintained by selection; the fact that the prevalent allele is the same in all populations may be due to selection and founder principle.Report no. 485 from the Tvärminne Zoological Station, University of Helsinki.     

Population genetic characterisation of dominant Cryptosporidium parvum subtype IIaA15G2R1

The subtype IIaA15G2R1 at the 60 kDa glycoprotein (gp60) gene locus is the most dominant Cryptosporidium parvum infecting dairy cattle and humans in industrialised nations. The reasons for its high transmissibility are not clear, and it remains to be determined whether this subtype represents a homogeneous parasite population. In this study, we sequence-characterised 26 IIaA15G2R subtype specimens and 26 non-IIaA15G2R subtype specimens from the United States, Canada, United Kingdom and Spain at seven other known polymorphic loci, including CP47, CP56, DZ-HRGP, MSC6-5, MSC6-7, RPGR and ZPT. Extensive heterogeneity within IIaA15G2R1 and discordance in typing results between gp60 and other genetic markers were observed. Results of inter-locus and intra-ZPT linkage disequilibrium and recombination analyses indicated that the heterogeneity within IIaA15G2R1 and discordance in typing results among genetic loci were largely due to the occurrence of genetic recombination, mostly within the gp60 subtype IIaA15G2R1. Although there was no clear population diversion between IIaA15G2R and non-IIaA15G2R subtypes, results of STRUCTURE and F_ST analyses suggested the presence of at least two subpopulations; subpopulation 1 had an epidemic population structure and was widely distributed, whereas subpopulation 2 had a clonal population structure and consisted of geographically segregated multilocus subtypes. Genetic recombination between epidemic and geographically segregated C. parvum populations appeared to be a driving force in the emergence of a hyper-transmissible IIaA15G2R1 subtype. Genetic recombination was observed even between the zoonotic IIa subtype family and anthroponotic subtype family IIc at CP56, MSC6-7 and ZPT. Thus, the IIaA15G2R1 subtype at gp60 is likely a fitness marker for C. parvum and the wide spread of IIaA15G2R1 subtype around the world is probably independent of the sequence characteristics at other genetic loci.     

Allozyme variation among Rana balcanica,R. levantina,and R. ridibunda (Amphibia: Anura)1

The allozyme variation among water frogs of the species R. balcanica, R. levantina and R. ridibunda, all formerly considered as one species (R. ridibunda Pallas, 1771), was studied using horizontal starch gel electrophoresis. Blood samples (N # 63) of frogs were collected from five populations in Greece and Israel. Samples (N = 9) of the hybrid frog R. esculenta collected from a locality in Germany were used as an outgroup for phylogenetic analyses. Fifteen enzymes controlled by twenty presumptive loci were identified. Thirteen loci were polymorphic within or among the studied populations. Genetic differentiation among the species was considerably greater than among populations of the same species. Even at Nestos River where R. ridibunda and R. balcanica occur in the same habitats, individuals could be assigned to either species due to characteristic differences of the genotypes (GPI1). This indicates reproductive isolation among these species. The reconstruction of phylogenetic relationships among the three species based on the allozyme data corroborated the model presented on the basis of bioacoustic data: R. ridibunda and R. balcanica (Nei's genetic distance D = 0.0820) are sibling species pertaining to an Eurasian lineage, whereas R. levantina (distance to the European species D = 0.1780 - 0.1955) together with R. perezi represent an independent afroasian lineage.