Molecular evolution and phylogeny of theDrosophila virilis species group as inferred by two-dimensional electrophoresis

Summary Systematic relationship among the 12 species of theDrosophila virilis species group, andDrosophila robusta, were investigated by the use of two-dimensional electrophoresis (2-DE). A total of 389 protein characters (about 200 loci) were scored and analyzed both phylogenetically and phenetically. The resulting phylogeny was found to be largely concordant with the current views of evolution among these species based on other independent morphological, chromosomal, electrophoretic, and immunological data sets, although some notable differences were observed. The 2-DE data also appeared to be useful for constructing a molecular clock to date the absolute times of divergence among the species. It appears from this analysis that the evolution of the major clades within the species group occurred about 20 million years ago. Previous suggestions that the rate of molecular evolution was different between the virilis and montana phylads was not confirmed. The technique of 2-DE seems to be an excellent tool for reconstructing phylogenies and should be particularly valuable for examining relatively closely related species.     

Sequence evolution of mitochondrial tRNA genes and deep-branch animal phylogenetics

Mitochondrial DNA sequences are often used to construct molecular phylogenetic trees among closely related animals. In order to examine the usefulness of mtDNA sequences for deep-branch phylogenetics, genes in previously reported mtDNA sequences were analyzed among several animals that diverged 20–600 million years ago. Unambiguous alignment was achieved for stem-forming regions of mitochondrial tRNA genes by virtue of their conservative secondary structures. Sequences derived from stem parts of the mitochondrial tRNA genes appeared to accumulate much variation linearly for a long period of time: nearly 100 Myr for transition differences and more than 350 Myr for transversion differences. This characteristic could be attributed, in part, to the structural variability of mitochondrial tRNAs, which have fewer restrictions on their tertiary structure than do nonmitochondrial tRNAs. The tRNA sequence data served to reconstruct a well-established phylogeny of the animals with 100% bootstrap probabilities by both maximum parsimony and neighbor joining methods. By contrast, mitochondrial protein genes coding for cytochrome b and cytochrome oxidase subunit I did not reconstruct the established phylogeny or did so only weakly, although a variety of fractions of the protein gene sequences were subjected to tree-building. This discouraging phylogenetic performance of mitochondrial protein genes, especially with respect to branches originating over 300 Myr ago, was not simply due to high randomness in the data. It may have been due to the relative susceptibility of the protein genes to natural selection as compared with the stem parts of mitochondrial tRNA genes. On the basis of these results, it is proposed that mitochondrial tRNA genes may be useful in resolving deep branches in animal phylogenies with divergences that occurred some hundreds of Myr ago. For this purpose, we designed a set of primers with which mtDNA fragments encompassing clustered tRNA genes were successfully amplified from various vertebrates by the polymerase chain reaction.Abbreviations AA stem amino acid-acceptor stem - AC stem anticodon stem - COI cytochrome oxidase subunit I - cytb cytochrome b - D stem dihydrouridine stem - MP maximum parsimony - mtDNA mitochondrial DNA - Myr million years - NJ neighbor joining - PCR polymerase chain reaction - Ti transition - T stem tC stem - Tv transversion Correspondence to: Y. Kumazawa     

Evolution of nested genes with special reference to cuticle proteins inDrosophila melanogaster

Summary One of the pupal cuticle protein (PCP) genes has been found within an intron of aDrosophila housekeeping gene (theGart locus) that encodes three enzymes involved in the purine pathway. This intronic gene has been described as a gene within a gene, and the gene is now called a “nested” gene. Because the intronic PCP gene has sequence similarity with the larval cuticle protein (LCP) gene, it may have been derived from one of the LCP genes or their ancestral gene. We have studied possible phylogenetic relationships among these five genes by comparing nucleotide sequences of four LCP genes with that of the PCP gene. The results obtained suggest that the PCP gene may have originated from an ancestral gene before duplication of the LCP genes occurred. Using the number of synonymous (silent) substitutions, we then estimated the divergence time between the PCP gene and the LCP genes to be about 70 million years (Myr). The divergence time estimated is much larger than that for the sibling species ofD. melanogaster (about 2.5 Myr), indicating that the “nested” gene structure can be seen not only inDrosophila melanogaster, but also in other distantly relatedDrosophila species.     

Phylogenetic reconstruction of South American felids defined by protein electrophoresis

Phylogenetic associations among six closely related South American felid species were defined by changes in protein-encoding gene loci. We analyzed proteins isolated from skin fibroblasts using two-dimensional electrophoresis and allozymes extracted from blood cells. Genotypes were determined for multiple individuals of ocelot, margay, tigrina, Geoffroy's cat, kodkod, and pampas cat at 548 loci resolved by two-dimensional electrophoresis and 44 allozyme loci. Phenograms were constructed using the methods of Fitch-Margoliash and neighbor-joining on a matrix of Nei's unbiased genetic distances for all pairs of species. Results of a relative-rate test indicate changes in two-dimensional electrophoresis data are constant among all South American felids with respect to a hyena outgroup. Allelic frequencies were transformed to discrete character states for maximum parsimony analysis. Phylogenetic reconstruction indicates a major split occurred approximately 5–6 million years ago, leading to three groups within the ocelot lineage. The earliest divergence led to Leopardus tigrina, followed by a split between an ancestor of an unresolved trichotomy of three species (Oncifelis guigna, O. geoffroyi, and Lynchaduris colocolo) and a recent common ancestor of Leopardus pardalis and L. wiedii. The results suggest that modern South American felids are monophyletic and evolved rapidly after the formation of the Panama land bridge between North and South America.Correspondence to: J. Pecon Slattery     

Timing of butterfly parasitization of a plant–ant–scale symbiosis

In the Southeast Asian tropics, Arhopala lycaenid butterflies feed on Macaranga ant-plants inhabited by Crematogaster (subgenus Decacrema) ants tending Coccus-scale insects. A recent phylogenetic study showed that (1) the plants and ants have been codiversifying for the past 20–16 million years (Myr), and that (2) the tripartite symbiosis was formed 9–7 Myr ago, when the scale insects became involved in the plant–ant mutualism. To determine when the lycaenids first parasitized the Macaranga tripartite symbiosis, we constructed a molecular phylogeny of the lycaenids that feed on Macaranga by using mitochondrial and nuclear DNA sequence data and estimated their divergence times based on the cytochrome oxidase I molecular clock. The minimum age of the lycaenids was estimated by the time-calibrated phylogeny to be 2.05 Myr, about one-tenth the age of the plant–ant association, suggesting that the lycaenids are latecomers that associated themselves with the pre-existing symbiosis of plant, ant, and scale insects.     

Estimation of molecular clocks for ITS and 28S rDNA in Erysiphales

Based on a comparative phylogenetic analysis of Goloviomyces and their host tribes of the Asteraceae, we speculate that Golovinomyces first acquired parasitism to the Asteraceae after migration of the family into the Northern Hemisphere and before the divergence of the tribe Carduaeae. The divergence time of the Carduaeae is estimated to be 25.2Myr ago based on the molecular clock of rbcL sequences of the Asteraceae. When 25.2Myr is given at the node of the first split of the phylogenetic tree of Golovinomyces, nucleotide substitution rates of the Erysiphales are calculated to be 2.52 × 10^–9 per site per year (0.01D = 3.97Myr) in the ITS region and 6.5 × 10^–10 per site per year (0.01D = 15.4Myr) in the D1 and D2 regions of the 28S rDNA.     

Phylogeny and Evolutionary History of the Ground Squirrels (Rodentia: Marmotinae)

Although ground squirrels (Spermophilus) and prairie dogs (Cynomys) are among the most intensively studied groups of mammals with respect to their ecology and behavior, a well-resolved phylogeny has not been available to provide a framework for comparative and historical analyses. We used complete mitochondrial cytochrome b sequences to construct a phylogeny that includes all 43 currently recognized species in the two genera, as well as representatives of two closely related genera (Marmota and Ammospermophilus). In addition, divergence times for ground squirrel lineages were estimated using Bayesian techniques that do not assume a molecular clock. All methods of phylogenetic analysis recovered the same major clades, and showed the genus Spermophilus to be paraphyletic with respect to both Marmota and Cynomys. Not only is the phylogeny at odds with previous hypotheses of ground squirrel relationships, but it suggests that convergence in morphology has been a common theme in ground squirrel evolution. A well-supported basal clade, including Ammospermophilus and two species in the subgenus Otospermophilus, diverged from all other ground squirrels an estimated 17.5 million years ago. Between 10 and 14 million years ago, a relatively rapid diversification gave rise to lineages leading to marmots and to several distinct groups of ground squirrels. The Eurasian ground squirrels diverged from their North American relatives during this period, far earlier than previously hypothesized. This period of diversification corresponded to warming climate and spread of grasslands in western North America and Eurasia. Close geographic proximity of related forms suggests that most species evolved in or near their current ranges.     

Phylogeny of Taxaceae and Cephalotaxaceae Genera Inferred from Chloroplast matK Gene and Nuclear rDNA ITS Region

Phylogeny of the Taxaceae genera and the monotypic family Cephalotaxaceae has been extraordinarily controversial. In this paper chloroplast matK genes and nuclear ITS sequences were determined for all six genera of the two families and representatives of other conifer families. Analysis using either the nonsynonymous sites or the deduced amino acid sequences of matK genes strongly indicates that taxad genera and Cephalotaxaceae are monophyletic, with the Taxodiaceae/Cupressaceae clade as their sister group. Cephalotaxus is basal to the taxad genera, among which two clades, Torreya/Amentotaxus and Taxus/Pseudotaxus/Austrotaxus, are resolved. They correspond to Janchen's two tribes, Torreyeae and Taxeae. In Taxeae, Austrotaxus is the first to branch off. Analyses of the nuclear ITS sequence data corroborated the topology of the matK gene tree. These results refute the views that Cephalotaxaceae has no alliance with Taxaceae and that Austrotaxus and Amentotaxus should be excluded from the Taxaceae. We estimated the divergence time between the Taxodiaceae/Cupressaceae and the Cephalotaxaceae/Taxaceae clades to be 192–230 Myr ago and the divergence time between taxads and Cephalotaxus to be 149–179 Myr ago. Soon after the latter divergence event, within 6–8 Myr, the two taxad tribes originated. In conclusion, our data do not support Florin's claim that taxads could be traced to Devonian psilophytes (359–395 Myr ago).     

Molecular phylogeny and evolution of subsection Magnicellulatae (Erysiphaceae: Podosphaera) with special reference to host plants

The subsection Magnicellulatae of the genus Podosphaera section Sphaerotheca belongs to the tribe Cystotheceae of the Erysiphaceae, which has the characteristic of producing catenate conidia with distinct fibrosin bodies. In this study, we newly determined the nucleotide sequences of the D1/D2 domains of the 28S rDNA region and the sequences of the rDNA internal transcribed spacer (ITS) region to investigate the relationships between the phylogeny of this fungal group and their host plants. The results indicated that the 28S rDNA region is too conservative for phylogenetic analysis of this fungal group. The phylogenetic analysis using 95 ITS sequences demonstrated that two or more Magnicellulatae taxa often infect the same plant genus or species. Although there is a close relationship between Magnicellulatae and asteraceous hosts, this association seems to be not as strict as that between Golovinomyces and the Asteraceae. The difference between the two fungal groups may be explained by their different evolutionary timing.     

Teleomorph-anamorph relationships and reclassification of <Emphasis Type="Italic">Cordyceps cuboidea</Emphasis> and its allied species

Based on morphological characteristics and molecular phylogeny, we reclassified Cordyceps cuboidea and allied species C. alboperitheciata, C. prolifica, and Ophiocordyceps ryogamiensis. We investigated their teleomorph-anamorph relationships and revealed that these four species have Hirsutella-like anamorphs with morphological differences between them. By analyzing their molecular phylogeny, inferred from DNA sequences of internal transcribed spacer (ITS) and large subunit (LSU) D1/D2 region of rDNA, they were separated into four close-knit clades. Although C. prolifica and O. ryogamiensis formed their own clades, isolates of C. cuboidea separated into two clades, i.e., a true C. cuboidea clade and one resembling a new species, the O. paracuboidea clade. The latter two species are distinguished by the fruiting region of the stroma. In addition, C. alboperitheciata is regarded as a synonym of C. cuboidea. From the morphology, teleomorph-anamorph relationships, and molecular phylogeny, we concluded these species should be assigned to the genus Ophiocordyceps.     

Nucleotide divergence of the rp49 gene region between Drosophila melanogaster and two species of the Obscura group of Drosophila

Summary A 2.1-kb SStI fragment including the rp49 gene and the 3 end of the -serendipity gene has been cloned and sequenced in Drosophila pseudoobscura. rp49 maps at region 62 on the tip of chromosome II of this species. Both the coding and flanking regions have been aligned and compared with those of D. subobscura. There is no evidence for heterogeneity in the rate of silent substitution between the rp49 coding region and the rate of substitutions in flanking regions, the overall silent divergence per site being 0.19. Noncoding regions also differ between both species by different insertions/deletions, some of which are related to repeated sequences. The rp49 region of D. pseudoobscura shows a strong codon bias similar to those of D. subobscura and D. melanogaster. Comparison of the rates of silent (K _S ) and nonsilent (K _a ) substitutions of the rp49 gene and other genes completely sequenced in D. pseudoobscura and D. melanogaster confirms previous results indicating that rp49 is evolving slowly both at silent and nonsilent sites. According to the data for the rp49 region, D. pseudoobscura and D. subobscura lineages would have diverged some 9 Myr ago, if one assumes a divergence time of 30 Myr for the melanogaster and obscura groups.Offprint requests to: C. Segarra     

Phylogenetic relationships of 34 passerines based on mitochondrial Cytochrome b sequences

Phylogenetic relationships of 34 passerines were studied based on mitochondrial Cytochrome b (Cyt b) sequences. Phylogenetic trees were constructed using Neighbor-Joining, Maximum-Parsimony and Minimum evolution methods. The results show that the divergence between Fringillids and Emberizids reaches a family level and they should be grouped into family Fringillidae and Emberizidae, respectively; Accentors has a relatively close relationship with Fringillids and Emberizids; the divergence between robins and flycatchers does not reach a family level and they should be member to family Muscicapidae; long-tailed tits and sylviids should all be listed into families; barn swallow, crowtits and long-tailed tits have close relationships with Sylviidae; in the Fringillidae, brambling should be member to one subfamily Fringillinae, several other birds under the subfamily Carduelinae; in the Sylviidae, although lanceolated warblers and scaly-headed stubtails have a relatively far relationship, they should be member to one subfamily Acrocephalinae and warblers to Phylloscopinae. Muscicapidae, Fringillidae and Emberidae are all monophylies, but Sylviidae is not. The substitution rates of major clades are thought to be the same according to relative rate tests. Divergence time of major clades is estimated at the rate of 1.6% per million years, thus the estimated divergence time between Fringillidae and Emberizidae is 10.5 million years, robins and flycatchers 9.0 Myr, Acrocephalinae and Phylloscopinae 9.0 Myr, Carduelis flammea and Carpodacus erythrinus 7.5 Myr, Luscinia cyane and Tarsiger cyanurus 7.8 Myr, two outgroups 13.5 Myr.     

Performance of 18S rRNA in Littorinid Phylogeny (Gastropoda: Caenogastropoda)

In the past, 18S rRNA sequences have proved to be very useful for tracing ancient divergences but were rarely used for resolving more recent ones. Moreover, it was suggested that the molecule does not contain useful information to resolve divergences which took place during less than 40 Myr. The present paper takes littorinid phylogeny as a case study to reevaluate the utility of the molecule for resolving recent divergences. Two data sets for nine species of the snail family Littorinidae were analyzed, both separately and combined. One data set comprised 7 new complete 18S rRNA sequences aligned with 2 published littorinid sequences; the other comprised 12 morphological, 1 biochemical, and 2 18S rRNA secondary structure characters. On the basis of its ability to confirm generally accepted relationships and the congruence of results derived from the different data sets, it is concluded that 18S rRNA sequences do contain information to resolve ``rapid' cladogenetic events, provided that they occurred in the not too distant past. 18S rRNA sequences yielded support for (1) the branching order (L. littorea, (L. obtusata, (L. saxatilis, L. compressa))) and (2) the basal position of L. striata in the Littorina clade. Received: 6 February 1998 / Accepted: 20 March 1998     

A view of early vertebrate evolution inferred from the phylogeny of polystome parasites (Monogenea: Polystomatidae)

The Polystomatidae is the only family within the Monogenea to parasitize sarcopterygians such as the Australian lungfish Neoceratodus poisteri and freshwater tetrapods (lissamphibians and chelonians). We present a phylogeny based on partial 18S rDNA sequences of 26 species of Polystomatidae and three taxon from the infrasubclass Oligonchoinea (= Polyopisthocotylea) obtained from the gills of teleost fishes. The basal position of the polystome from lungfish within the Polystomatidae suggests that the family arose during the evolutionary transition between actinopterygians and sarcopterygians, ca. 425 million years (Myr) ago. The monophyly of the polystomatid lineages from chelonian and lissamphibian hosts, in addition to estimates of the divergence times, indicate that polystomatids from turtles radiated ca. 191 Myr ago, following a switch from an aquatic amniote presumed to be extinct to turtles, which diversified in the Upper Triassic. Within polystomatids from lissamphibians, we observe a polytomy of four lineages, namely caudatan, neobatrachian, pelobatid and pipid polystomatid lineages, which occurred ca. 246 Myr ago according to molecular divergence-time estimates. This suggests that the first polystomatids of amphibians originated during the evolution and diversification of lissamphibian orders and suborders ca. 250 Myr ago. Finally, we report a vicariance event between two major groups of neobatrachian polystomes, which is probably linked to the separation of South America from Africa ca. 100 Myr ago.     

Phylogeny of the genus Hesperodiaptomus (Copepoda) based on nucleotide sequence data of the nuclear ribosomal gene

The genus Hesperodiaptomus Light, 1938, one of the most diverse groups of freshwater copepods that occur in North and Central America, plays a major role in the food webs of the alpine freshwater communities. Phylogenetic relationships of these taxa remain poorly understood due to difficulties in obtaining reliable morphological characters for phylogenetic analyses. To understand the phylogenetic relationships within this group, we reconstructed a partial phylogeny of the genus Hesperodiaptomus based on nuclear ribosomal gene sequences. Phylogenetic analyses based upon the taxa examined supported the monophyly the genus and revealed two clades. The eiseni clade comprised species that are morphologically similar to Hesperodiaptomus eiseni (Lilljeborg, 1889), and the shoshone clade included species morphologically similar to Hesperodiaptomus shoshone (S.A. Forbes, 1882). The two groups can be distinguished by a modification of the right basis, the arrangement of spinules on the distal pad of the second exopod, and the degree of presence the inner lamellar expansion of the right coxa. Handling editor: P. Spaak     

Phylogeny and biogeography of Chinese and Australasian Polystichum ferns as inferred from chloroplast trnL-F and rps4-trnS sequence data

Polystichum, one of the largest genera of ferns, occurs worldwide with the greatest diversity in southwest China and adjacent regions. Although there have been studies of Chinese Polystichum on its traditional classification, geographic distributions, and even a few on its molecular systematics, its relationships to other species outside China remain little known. Here, we investigated the phylogeny and biogeography of the Polystichum species from China and Australasia. The evolutionary relationships among 42 Polystichum species found in China (29 taxa) and Australasia (13 taxa) were inferred from phylogenetic analyses of two chloroplast DNA sequence data sets: rps4-trnS and trnL-F intergenic spacers. The divergence time between Chinese and Australasian Polystichum was estimated. The results indicated that the Australasian species comprise a monophyletic group that is nested within the Chinese diversity, and that the New Zealand species are likewise a monophyletic group nested within the Australasian species. The divergence time estimates suggested that Chinese Polystichum migrated into Australasia from around 40 Ma ago, and from there to New Zealand from about 14 Ma. The diversification of the New Zealand Polystichum species began about 10 Ma. These results indicated that Polystichum probably originated in eastern Asia and migrated into Australasia: first into Australia and then into New Zealand.     

Evolution in bacteria: Evidence for a universal substitution rate in cellular genomes

Summary This paper constructs a temporal scale for bacterial evolution by tying ecological events that took place at known times in the geological past to specific branch points in the genealogical tree relating the 16S ribosomal RNAs of eubacteria, mitochondria, and chloroplasts. One thus obtains a relationship between time and bacterial RNA divergence which can be used to estimate times of divergence between other branches in the bacterial tree. According to this approach,Salmonella typhimurium andEscherichia coli diverged between 120 and 160 million years (Myr) ago, a date which fits with evidence that the chief habitats occupied now by these two enteric species became available that long ago.The median extent of divergence betweenS. typhimurium andE. coli at synonymous sites for 21 kilobases of protein-coding DNA is 100%. This implies a silent substitution rate of 0.7–0.8%/Myr—a rate remarkably similar to that observed in the nuclear genes of mammals, invertebrates, and flowering plants. Similarities in the substitution rates of eucaryotes and procaryotes are not limited to silent substitutions in protein-coding regions. The average substitution rate for 16S rRNA in eubacteria is about 1%/50 Myr, similar to the average rate for 18S rRNA in vertebrates and flowering plants. Likewise, we estimate a mean rate of roughly 1%/25 Myr for 5S rRNA in both eubacteria and eucaryotes.For a few protein-coding genes of these enteric bacteria, the extent of silent substitution since the divergence ofS. typhimurium andE. coli is much lower than 100%, owing to extreme bias in the usage of synonymous codons. Furthermore, in these bacteria, rates of amino acid replacement were about 20 times lower, on average, than the silent rate. By contranst, for the mammalian genes studied to date, the average replacement rate is only four to five times lower than the rate of silent substitution.     

Mitochondrial DNA evolution in theDrosophila nasuta subgroup of species

Summary TheDrosophila nasuta group consists of about 12 closely related species distributed throughout the Indo-Pacific region. They are of great interest because of their evolutionary idiosyncrasies including little morphological differentiation, the ability to intercross in the laboratory often producing fertile offspring, and substantial chromosomal evolution. Studies of metric traits, reproductive isolation, and chromosomal and enzyme polymorphisms have failed to resolve the phylogeny of the species. We report the results of a survey of the mitochondrial DNA (mtDNA) restriction patterns of the species. The phylogeny obtained is consistent with other available information and suggests thatD. albomicans may represent the ancestral lineage of the group. The amount of polymorphism in local populations (=1.0% per site) is within the typical range observed in other animals, includingDrosophila. The degree of differentiation between species is, however, low: the origin of the group is tentatively dated about 6–8 million years ago. This study confirms the usefulness of mtDNA restriction patterns for ascertaining the phylogeny of closely related species.     

DNA hybridization evidence of hominoid phylogeny: Results from an expanded data set

Summary The living hominoids are human, the two species of chimpanzees, gorilla, orangutan, and nine species of gibbons. The cercopithecoids (Old World monkeys) are the sister group of the hominoids. A consensus about the phylogeny of the hominoids has been reached for the branching order of the gibbons (earliest) and the orangutan (next earliest), but the branching order among gorilla, chimpanzees, and human remains in contention. In 1984 we presented DNA-DNA hybridization data, based on 183 DNA hybrids, that we interpreted as evidence that the branching order, from oldest to most recent, was gibbons, orangutan, gorilla, chimpanzees, and human. In the present paper we report on an expanded data set totaling 514 DNA hybrids, which supports the branching order given above. The ranges for the datings of divergence nodes are Old World monkeys, 25–34 million years (Myr) ago; gibbons, 16.4–23 Myr ago; orangutan, 12.2–17 Myr ago; gorilla, 7.7–11 Myr ago; chimpanzees-human, 5.5–7.7 Myr ago. The possible effects of differences in age at first breeding are discussed, and some speculations about average genomic rates of evolution are presented.     

Ceratomyxa (Myxozoa: Bivalvulida): Robust taxon or genus of convenience?

The genus Ceratomyxa (Myxozoa: Myxosporea: Bivalvulida) contains parasites that typically infect the gall bladders of marine teleosts. Species of this genus have also been recorded from elasmobranchs, while the best known species (Ceratomyxa shasta) is a systemic pathogen of fresh water salmonid fishes. Here we characterise 10 new species of Ceratomyxa from marine teleosts using morphometric and rDNA sequence data. A phylogeny of all Ceratomyxa species for which ssrDNA sequence is available was estimated by parsimony, maximum likelihood and Bayesian analyses. Mapping host fish taxonomy, geographic locality and morphology onto the phylogenetic tree provided some concordance of these characters to groups of Ceratomyxa species, but in no case was it consistent throughout the inferred phylogeny. The position of C. shasta as a sister species to the Ceratomyxa clade contradicts previous estimates of marine myxozoan phylogeny which suggested C. shasta was an unrelated lineage. Comparative DNA sequence data is available for more than 17% of some 200 described Ceratomyxa species and the genus now represents one of the most cohesive lineages within the Myxozoa. The independent branching of all atypical Ceratomyxa species and Palliatus indecorus, indicates a review of the diagnostic characters and possible division into more genera is warranted when further data are available.