Sperm morphology in the Malagasy rodents (Muroidea: Nesomyinae)

The morphology of the spermatozoon of representative species of the subfamily Nesomyinae (Muroidea: Nesomyidae), a monophyletic group of rodents endemic to Madagascar, was examined by light and electron microscopy to determine the sperm head shape and tail length across the species. Marked interspecific differences were found to occur in both the form of the sperm head and length of the tail. The species that possess a sperm head with an apical hook, which largely contains acrosomal material, generally displayed longer sperm tails, and a species with a spatulate sperm head had the shortest tail. The association between sperm head shape and tail length mirrors that previously found in Eurasian and Australasian murine rodents. Thus, the repeated association between sperm head shape and tail length across these groups of muroid rodents clearly indicates a functional relationship between these two features. A comparison of sperm morphology of the nesomyines to that of related muroid rodents on the mainland of Africa suggests that the possession of an apical hook is the ancestral condition. J. Morphol., 2010. © 2010 Wiley‐Liss, Inc.     

Morphological diversity and evolution of the spermatozoon in the mouse‐related clade of rodents

Most species in the three highly speciose families of the mouse‐related clade of rodents, the Muridae, Cricetidae, and Nesomyidae (superfamily Muroidea), have a highly complex sperm head in which there is an apical hook but there are few data available for the other related families of these rodents. In the current study, using light and electron microscopies, we investigated the structure of the spermatozoon in representative species of four other families within the mouse‐related clade, the Dipodidae, Spalacidae, Pedetidae, and Heteromyidae, that diverged at or near the base of the muroid lineage. Our results indicate that a diverse array of sperm head shapes and tail lengths occurs but none of the species in the families Spalacidae, Dipodidae, or Pedetidae has a sperm head with an apical hook. By contrast, a rostrally extending apical hook is present in spermatozoa of members of the Family Heteromyidae which also invariably have comparatively long sperm tails. These findings suggest that the hook‐shaped sperm head in the murid, cricetid, and nesomyid rodents evolved after divergence of this lineage from its common ancestor with the other families of the mouse‐related clade, and that separate, and independent, convergent evolution of a similar sperm head form, and long sperm tail, occurred in the Heteromyidae. J. Morphol. 275:540–547, 2014. © 2013 Wiley Periodicals, Inc.     

The spermatozoon of Eurasian murine rodents: Its morphological diversity and evolution

The murine rodents are the most speciose subfamily of mammals. Here the morphology of the spermatozoon, as determined by scanning and transmission electron microscopy of representative species from four Eurasian clades, is described. Much interspecific variability in all components of the spermatozoon was found to occur, although most species have a bilaterally flattened sperm head with a single apical hook of variable length and orientation. Ultrastructural observations indicate that this apical hook invariably contains a nuclear projection as well as a large extension of the subacrosomal cytoskeleton, as a perforatorium rostrally, and a complex asymmetrical acrosomal extension. These spermatozoa also have relatively long tails that are attached to the lower concave surface of the sperm head. Uniquely, in species in the Apodemus clade, the apical hook is orientated caudally. In a few species a highly derived sperm head morphotype that does not contain an apical hook is present. These sperm heads vary in morphology from being globular in two species of Bandicota, to bilaterally flattened and paddle-shaped in Tokudaia and Micromys. In spermatozoa of the latter two genera the subacrosomal cytoskeleton, which is less extensive than in species with a hooked sperm head, forms an apical extension, but that is not the case in Bandicota. In all species where the sperm head lacks an apical hook the acrosome is more symmetrical. The sperm tail is much shorter in these species, with attachment to the head occurring on the ventral surface in Tokudaia and basal in Micromys and the two species of Bandicota. As the sperm head morphotype with a complex apical hook is present in all the major clades of murine rodents, it is likely to be a plesiomorphic character within each of these clades, with the nonhooked sperm heads, which vary greatly in structure between species of the different lineages, probably being independently derived. The ultrastructural organization of the sperm head of Bandicota, but not those of Micromys or Tokudaia, suggest divergence in some of the morphological events associated with sperm-egg interaction at the time of fertilization.     

The Gerbil Spermatozoon – interspecific variation in morphology does not invariably follow murine rodent evolutionary trends in response to predicted intermale sperm competition

In various groups of mammals, the intensity of intermale sperm competition relates to relative testes mass (RTM) with some evidence suggesting that this may also be the case for some aspects of sperm form. In murid rodents, a large RTM generally correlates with a streamlined sperm head, long apical hook and long tail with most data coming from species in the subfamily Murinae. In this study, RTM and sperm form are compared across 15 species of gerbils, seven from the Tribe Taterillini and eight from the Tribe Gerbillini. Marked interspecific differences in RTM and sperm morphology were observed. However, the Gerbilliscus species with the largest RTM do not have a sperm head with an apical hook nor a longer sperm tail than other species with smaller RTM whereas, by contrast, in the Tribe Gerbillini, species where the sperm head lacks a hook have a relatively small testes mass. We thus suggest that in gerbils, unlike in murine rodents, high levels of postcopulatory sexual selection have not invariably resulted in the evolution of a spermatozoon with a long apical hook and long sperm tail. The possible reasons for this are briefly discussed.     

Is the hook of muroid rodent's sperm related to sperm train formation?

Competition between spermatozoa of rival males to gain fertilizations has led to a wide array of modifications in sperm structure and function. Sperm cells of most muroid rodents have hook‐shaped extensions in the apical–ventral tip of the head, but the function of this structure is largely unknown. These ‘hooks’ may facilitate aggregation of spermatozoa in so‐called ‘trains’, as an adaptation to sperm competition, because sperm in trains may swim faster than free‐swimming cells. However, there is controversy regarding the role of the hook in train formation, and in relation to whether it is selected by sperm competition. We examined spermatozoa from muroid rodents with varying levels of sperm competition to assess whether (i) sperm aggregates are common in these taxa, (ii) presence of a hook relates to the formation of sperm aggregations, and (iii) formation of sperm aggregations is explained by sperm competition. Our analyses in 25 muroid species revealed that > 92% of spermatozoa swim individually in all species, with the exception of the wood mouse, Apodemus sylvaticus, which has ~50% spermatozoa swimming freely. Species with hooked spermatozoa had higher sperm competition levels and longer sperm than species whose sperm lack a hook. Neither the presence of hook nor sperm competition levels were related to the percentage of sperm in aggregations. Thus, (i) sperm aggregates in muroid rodents are an exceptional trait found only in a few species, (ii) evolution of the sperm hook is associated to sperm competition levels, but (iii) the hook is unlikely to be related to the formation of sperm aggregates. The evolutionary significance of the sperm head hook thus remains elusive, and future studies should examine potential roles of this pervasive structure in sperm's hydrodynamic efficiency and sperm–female tract interactions.     

Sperm competition and the evolution of the sperm hook in house mice

Sperm morphology varies considerably both between and within species. The sperm of many muroid rodents bear an apical hook at the proximal end of the head. The curvature of the sperm hook varies greatly across species, however the adaptive significance of the sperm hook is currently not known. In wood mice the apical hooks intertwine to form sperm ‘trains’, which exhibit faster swimming velocities than single cells. Thus, it has been suggested that if sperm ‘trains’ were advantageous in a competitive situation, then the apical sperm hook might be an evolutionary product of selection via sperm competition. A comparative study of rodent species provided support for the hypothesis, and showed that species with higher levels of sperm competition had more reflected sperm hooks. Here, we tested this hypothesis at the intraspecific level. We quantified sperm hook morphology from seven house mouse populations, and found that interpopulation variation in hook curvature was not explained by variation in sperm competition risk. Furthermore, observations of ejaculated sperm revealed that sperm groups are not a common characteristic of mouse ejaculates. We suggest that selection for sperm attachment to the oviduct epithelium, and thus better retainment of sperm fertilizing potential, may provide a more general explanation of the evolutionary relationship between sperm competition risk and the curvature of the sperm hook among rodents, and provide a phylogenetic comparison among rodent species that supports our hypothesis.     

The spermatozoon of the Old Endemic Australo‐Papuan and Philippine rodents – its morphological diversity and evolution

Breed, W.G. and Leigh, C.M. 2010. The spermatozoon of the Old Endemic Australo‐Papuan and Philippine rodents – its morphological diversity and evolution.—Acta Zoologica (Stockholm) 91 : 279–294 The spermatozoon of most murine rodents contains a head in which there is a characteristic apical hook, whereas most old endemic Australian murines, which are part of a broader group of species that also occur in New Guinea and the Philippines, have a far more complex sperm form with two additional ventral processes. Here we ask the question: what is the sperm morphology of the New Guinea and Philippines species and what are the trends in evolutionary changes of sperm form within this group? The results show that, within New Guinea, most species have a highly complex sperm morphology like the Australian rodents, but within the Pogonomys Division some species have a simpler sperm morphology with no ventral processes. Amongst the Philippines species, many have a sperm head with a single apical hook, but in three Apomys species the sperm head contains two additional small ventral processes, with two others having cockle‐shaped sperm heads. When these findings are plotted on a molecular phylogeny, the results suggest that independent and convergent evolution of highly complex sperm heads containing two ventral processes has evolved in several separate lineages. These accessory structures may support the sperm head apical hook during egg coat penetration.     

No evidence of sperm conjugate formation in an Australian mouse bearing sperm with three hooks

Sperm conjugation occurs when two or more sperm physically unite for motility or transport through the female reproductive tract. In many muroid rodent species, sperm conjugates have been shown to form by a single, conspicuous apical hook located on the sperm head. These sperm “trains” have been reported to be highly variable in size and, despite all the heads pointing in roughly the same direction, exhibit a relatively disordered arrangement. In some species, sperm “trains” have been shown to enhance sperm swimming speed, and thus have been suggested to be advantageous in sperm competition. Here, we assessed the behavior of sperm in the sandy inland mouse (Pseudomys hermannsburgensis), a muroid rodent that bears sperm with three apical hooks. First, we accrued genetic evidence of multiple paternity within “wild” litters to unequivocally show that sperm competition does occur in this species. Following this we utilized both in vitro and in vivo methodologies to determine whether sandy inland mouse sperm conjugate to form motile trains. Our observations of in vitro preparations of active sperm revealed that sandy inland mouse sperm exhibit rapid, progressive motility as individual cells only. Similarly, histological sections of the reproductive tracts of mated females revealed no in vivo evidence of sperm conjugate formation. We conclude that the unique, three‐hooked morphology of the sandy inland mouse sperm does not facilitate the formation of motile conjugates, and discuss our findings in relation to the different hypotheses for the evolution of the muroid rodent hook/s.     

Phylogeny of muroid rodents: relationships within and among major lineages as determined by IRBP gene sequences

The rodent family Muridae is the single most diverse family of mammals with over 1300 recognized species. We used DNA sequences from the first exon ( approximately 1200bp) of the IRBP gene to infer phylogenetic relationships within and among the major lineages of muroid rodents. We included sequences from every recognized muroid subfamily except Platacanthomyinae and from all genera within the endemic Malagasy subfamily Nesomyinae, all recognized tribes of Sigmodontinae, and a broad sample of genera in Murinae. Phylogenetic analysis of the IRBP data suggest that muroid rodents can be sorted into five major lineages: (1) a basal clade containing the fossorial rodents in the subfamilies Spalacinae, Myospalacinae, and Rhizomyinae, (2) a clade of African and Malagasy genera comprising the subfamilies Petromyscinae, Mystromyinae, Cricetomyinae, Nesomyinae, and core dendromurines, (3) a clade of Old World taxa belonging to Murinae, Otomyinae, Gerbillinae, Acomyinae, and Lophiomyinae, (4) a clade uniting the subfamilies Sigmodontinae, Arvicolinae, and Cricetinae, and (5) a unique lineage containing the monotypic Calomyscinae. Although relationships among the latter four clades cannot be resolved, several well-supported supergeneric groupings within each are identified. A preliminary examination of molar tooth morphology on the resulting phylogeny suggests the triserial murid molar pattern as conceived by evolved at least three times during the course of muroid evolution.     

Spermatozoa of murid rodents from Africa: morphological diversity and evolutionary trends

The diversity of the structural organization of the spermatozoa of African murid rodents is described at the light and transmission electron microscopical level of resolution. In most species the sperm head is falciform in shape but it varies somewhat in overall breadth, width, and length. A typical perforatorium is present and the acrosome splits into a large head cap over the convex surface and a smaller ventral segment similar to the sperm head of most Asian and Australasian murids. In a few species, however, the morphology is very different. In Acomys and Uranomys spermatozoa, the apical hook is more bilaterally flattened, has a large apical acrosomal region, and no separate ventral segment. Two species of Aethomys have, in addition to an apical hook, a 4μ long extension of the cytoskeletal material that projects from the concave surface of the sperm head, whereas in Dasymys two large ventral processes extend from the upper concave region which contain nuclear material basally and a huge extension of cytoskeleton apically. In Aethomys chrysophilus type B, the sperm nucleus is unique in form and often has a central region in which threads of chromatin can be seen; it is capped by a massive acrosome whose apical segment is complex and convoluted in structure. Stochomys longicaudatus appears to have a conical sperm head, and in all three Lophuromys species the sperm head is spatulate in shape with the flat, plate-like nucleus capped by a thin acrosome. The evolutionary trends in changes of sperm head shape and design of these rodents are discussed. It is suggested that some of the differences in morphology may relate to the variation in structural organization of the coats around the egg through which the spermatozoon has to pass in order for fertilization to occur.     

Divergent patterns of breakpoint reuse in Muroid rodents

Multiple Genome Rearrangement (MGR) analysis was used to define the trajectory and pattern of chromosome rearrangement within muroid rodents. MGR was applied using 107 chromosome homologies between Mus, Rattus, Peromyscus, the muroid sister taxon Cricetulus griseus, and Sciurus carolinensis as a non-Muroidea outgroup, with specific attention paid to breakpoint reuse and centromere evolution. This analysis revealed a high level of chromosome breakpoint conservation between Rattus and Peromyscus and indicated that the chromosomes of Mus are highly derived. This analysis identified several conserved evolutionary breakpoints that have been reused multiple times during karyotypic evolution in rodents. Our data demonstrate a high level of reuse of breakpoints among muroid rodents, further supporting the “Fragile Breakage Model” of chromosome evolution. We provide the first analysis of rodent centromeres with respect to evolutionary breakpoints. By analyzing closely related rodent species we were able to clarify muroid rodent karyotypic evolution. We were also able to derive several high-resolution ancestral karyotypes and identify rearrangements specific to various stages of Muroidea evolution. These data were useful in further characterizing lineage-specific modes of chromosome evolution.     

Interspecific variation of sperm morphology in the Australian rodent genus Zyzomys

The sperm head morphology and tail length of two species of Australian rock rats, Zyzomys argurus and Zyzomys pedunculatus, are presented. In Z. argurus the sperm head has an apical hook together with two ventral processes extending from the upper concave surface that are largely composed of cytoskeletal material, and the sperm tail is about 135 µm in length. By contrast, in Z. pedunculatus the sperm head is paddle‐shaped with the nucleus capped by an acrosome that has a large apical segment and is surrounded by a thin layer of cytoskeletal material, and the sperm tail is only around 85 µm in length. Since the structure of the spermatozoon of Z. argurus is similar to that of most of the old endemic Australian rodents it is presumed to be the ancestral condition within the Zyzomys genus with that of Z. pedunculatus being highly derived and showing convergence with the sperm structure in some other orders of mammals.     

Sperm competition does not influence sperm hook morphology in selection lines of house mice

Sperm show a remarkable degree of variation in size, shape and complexity. Murine rodents exhibit a sperm head morphology that differs greatly from the ovoid shape that is characteristic of most mammals. These rodents have sperm that bear one or more apical hooks, the function of which is currently surrounded by much controversy. It has been suggested that the hook serves to facilitate the formation of sperm groups, which in some species exhibit relatively faster velocities than single cells and thus, may provide an advantage when ejaculates are competing for fertilisations. In support of this hypothesis, a comparative study reported a positive association between the strength of sperm competition (estimated from testes size) and the curvature of the sperm hook amongst 37 murine species. Here, we assessed whether sperm competition influences sperm hookedness at the intra-specific level. Following 16 generations of selection, we used geometric morphometry (GM) to describe sperm head morphology in selection lines of house mice evolving with (polygamous) and without (monogamous) sperm competition. Although the GM analysis returned two relative warps that described variation in the curvature of the sperm hook, we found no evidence of divergence between the selection lines. Thus, we can conclude that sperm competition does not influence the degree of sperm hookedness in house mice.     

Ecological and Ecomorphological Specialization Are Not Associated with Diversification Rates in Muroid Rodents (Rodentia: Muroidea)

Multiple diversification rate shifts explain uneven clade richness in muroid rodents. Previous muroid studies have shown that extrinsic factors, notwithstanding ecological opportunity, are poor predictors of clade diversity. Here, we use a 297-muroid species chronogram that is sampled proportional to total clade diversity, along with various trait-dependent diversification approaches to investigate the association between diversification rates with intrinsic attributes—diet, habitat, body mass, and relative tail length. We found some association between both dietary specialization and body mass, as well as between habitat specialization with relative tail lengths using phylogenetic analyses of variance. However, there was no significant association between diversification rates with the evolution of these traits in muroid rodents. We also show that several of the state-dependent diversification approaches are highly susceptible to Type I error—a result that is in accordance with recent criticisms of these methods. Finally, we discuss several potential causes for the lack of association between the examined trait data with diversification rates, ranging from methodological biases (e.g. method conservativism) to biology (e.g. behavioral plasticity and ecological opportunism of muroid rodents).     

Morphology of the spermatozoon of a murid rodent from Africa, Dasymys incomtus

The structural organization of the head of the spermatozoon from the African murid rodent Dasymys incomtus was investigated by light and transmission electron microscopy and after incubation in various chemical agents. The results show that it is falciform in shape with a bifurcated rostral extension lying just caudal to the apical hook. The nucleus protrudes into the base of this extension but it is largely composed of a massive elaboration of the sperm head cytoskeleton which is very resistant to dispersion by chemical agents. This elaborate accessory structure of the sperm head shows similar morphology to the ventral processes on the sperm head of Australasian hydromyine rodents. The possible functional and evolutionary significance of these findings are discussed.     

Changes in the curvature of sperm apical hooks in murine rodents

Sperm apical hooks in murine rodents play an important role in sperm competition. Apical hooks are more curved and longer in species with relatively larger testes, that is in species with a higher risk of sperm competition. The sperm can form aggregations, ‘trains’, that can move faster than individual sperm, thus reaching the egg earlier as was observed in Apodemus sylvaticus. The apical hook plays an important role for train formation. This study focuses on the changes in the curvature of sperm apical hooks during the final stages of spermiogenesis and stages before fertilization (sperm-life span). Apical hook curvatures of field mice (A. agrarius and A. sylvaticus) vary significantly between dormant and active sperm. In contrast, there are no significant differences among the stages in the eastern house mouse. Since there are high ranges of angle values in all stages, the mean angles of apical hook curvature are not appropriate for evaluating risk of sperm competiton. The ranges of angle values point to a level of flexibility of the apical hooks. The lengths of sperm hooks in individual species do not change during particular stages. The length and flexibility of the sperm apical hooks are important for the formation of sperm aggregations, thus these sperm characters indicate the risk of sperm competition and the sperm strategies in murine rodents.     

Intra-individual variation in sperm tail length in murine rodents

In eutherian mammals, there are marked interspecific differences in sperm head shape and tail length. In a few species, sperm head variability occurs but intra-individual variation in sperm tail length has rarely been investigated or commented upon. Here, we ask the question: Do murine rodent species that have variable sperm head shapes exhibit greater intra-individual variation in sperm midpiece and total tail lengths than closely related species where little, or no, sperm head variability occurs? From three separate lineages, we selected three pairs of murine rodents, one of which has monomorphic, and the other variable, sperm head shape. These were from southern Asia the bandicoot rats Bandicota bengalensis and Bandicota indica , from southern Africa the veld rats, Aethomys chrysophilus and Aethomys ineptus and from Australia the fawn hopping mouse Notomys cervinus and the spinifex hopping mouse Notomys alexis . Cauda epididymal sperm smears were prepared and sperm midpiece and total tail lengths were determined. A linear mixed-effects model was used to estimate intra-individual variance. The results showed that in all three species where there are variable sperm head shapes ( B. indica , A. ineptus and N. alexis ), statistically significantly greater intra-individual variability of sperm midpiece and total tail lengths occurs ( P <0.0001 in all cases). These species all have relatively smaller testes mass compared with the closely related species with monomorphic sperm populations. This suggests that depressed levels of intermale sperm competition may result in the occurrence of variability in not only the divergent sperm head shape but also in the length of the midpiece as well as that of the total length of the sperm tail.     

Phylogeny and divergence-date estimates of rapid radiations in muroid rodents based on multiple nuclear genes

The muroid rodents are the largest superfamily of mammals, containing nearly one third of all mammal species. We report on a phylogenetic study comprising 53 genera sequenced for four nuclear genes, GHR, BRCA1, RAG1, and c-myc, totaling up to 6400 nucleotides. Most relationships among the subfamilies are resolved. All four genes yield nearly identical phylogenies, differing only in five key regions, four of which may represent particularly rapid radiations. Support is very strong for a fundamental division of the mole rats of the subfamilies Spalacinae and Rhizomyinae from all other muroids. Among the other "core" muroids, a rapid radiation led to at least four distinct lineages: Asian Calomyscus, an African clade of at least four endemic subfamilies, including the diverse Nesomyinae of Madagascar, a hamster clade with maximum diversity in the New World, and an Old World clade including gerbils and the diverse Old World mice and rats (Murinae). The Deomyinae, recently removed from the Murinae, is well supported as the sister group to the gerbils (Gerbillinae). Four key regions appear to represent rapid radiations and, despite a large amount of sequence data, remain poorly resolved: the base of the "core" muroids, among the five cricetid (hamster) subfamilies, within a large clade of Sigmodontinae endemic to South America, and among major geographic lineages of Old World Murinae. Because of the detailed taxon sampling within the Murinae, we are able to refine the fossil calibration of a rate-smoothed molecular clock and apply this clock to date key events in muroid evolution. We calculate rate differences among the gene regions and relate those differences to relative contribution of each gene to the support for various nodes. The among-gene variance in support is greatest for the shortest branches. We present a revised classification for this largest but most unsettled mammalian superfamily.     

Variation in Apical Hook Length Reflects the Intensity of Sperm Competition in Murine Rodents

Background

Post-copulatory sexual selection has been shown to shape morphology of male gametes. Both directional and stabilizing selection on sperm phenotype have been documented in vertebrates in response to sexual promiscuity.

Methodology

Here we investigated the degree of variance in apical hook length and tail length in six taxa of murine rodents.

Conclusions

Tail sperm length and apical hook length were positively associated with relative testis mass, our proxy for levels of sperm competition, thus indicating directional post-copulatory selection on sperm phenotypes. Moreover, our study shows that increased levels of sperm competition lead to the reduction of variance in the hook length, indicating stabilizing selection. Hence, the higher risk of sperm competition affects increasing hook length together with decreasing variance in the hook length. Species-specific post-copulatory sexual selection likely optimizes sperm morphology.     

Co‐evolution of gametes of the Greater Bandicoot Rat,Bandicota indica – a murine rodent from South‐East Asia

Most Old World mice and rats, subfamily Murinae, have a spermatozoon with an apical hook, a long tail and, as seen typically in eutherian mammals, a bilaterally flattened head. Dramatically different from this are the sperm of the Greater Bandicoot Rat, Bandicota indica. Here, we ask the question has the structure of the sperm head co‐evolved with that of the egg coat, the zona pellucida? For this, we first summarise the morphological features of the spermatozoon of B. indica that may relate to zona pellucida penetration at the time of fertilisation, and we confirm that the sperm head is generally round, not bilaterally flattened, in profile and has a huge acrosome. We then show that the zona pellucida around oocytes in tertiary follicles also differs from that of the other murine rodents in being only about 4 μm thick and, as demonstrated by lectin staining, has an unusual abundance of alpha‐L‐fucose. These findings indicate that both the male and female gametes of this South‐East Asian murine rodent are highly divergent in their structural organisation. One of the functional implications of this probably relates to sperm–zona interactions and the release of acrosomal enzymes that probably facilitate penetration by digestion of the zona matrix at the time of fertilisation.