Dysfunction of dimorphic sperm impairs male fertility in the silkworm

Sperm, which have a vital role in sexual reproduction in the animal kingdom, can display heteromorphism in some species. The regulation of sperm dichotomy remains a longstanding puzzle even though the phenomenon has been widely documented for over a century. Here we use Bombyx mori as a model to study a form of sperm dimorphism (eupyrene and apyrene sperm), which is nearly universal among Lepidoptera. We demonstrate that B. mori Sex-lethal (BmSxl) is crucial for apyrene sperm development, and that B. mori poly(A)-specific ribonuclease-like domain-containing 1 (BmPnldc1) is required for eupyrene sperm development. BmSXL is distributed in the nuclei and cytoplasm of somatic cyst cells in a mesh-like pattern and in the cytoplasm of germ cells enclosed in spermatocysts and sperm bundles. Cytological analyses of dimorphic sperm in BmSxl mutants (∆BmSxl) showed deficient apyrene sperm with abnormal nuclei, as well as loss of motility associated with malformed mitochondrial derivatives. We define the crucial function of apyrene sperm in the process of fertilization as assisting the migration of eupyrene spermatozoa from bursa copulatrix to spermatheca. By contrast, BmPnldc1 deficiency (∆BmPnldc1) caused eupyrene sperm abnormalities and impaired the release of eupyrene sperm bundles during spermiation. Although apyrene or eupyrene sperm defects impaired fertility of the mutated males, double copulation of a wild-type female with ∆BmSxl and ∆BmPnldc1 males could rescue the sterility phenotypes induced by single copulation with either gene-deficient male. Our findings demonstrate the crucial functions of BmSxl and BmPnldc1 in the development of sperm dimorphism and the indispensable roles of nonfertile apyrene sperm in fertilization.

Sperm dimorphism in the free-living marine nematode Terschellingia glabricutis (Nematoda: Monhysterida: Linhomoeidae)

The structure and development of the dimorphic spermatozoa of the free-living marine nematode Terschellingia glabricutis (Monhysterida, Linhomoeina, Linhomoeidae) were studied with transmission electron microscope (TEM). The linhomoeid pattern of spermatozoa includes absence of fibrous bodies (FB) in spermatocytes and their appearance in spermatids as spindle-shaped bodies never associating with membranous organelles (MO); presence of centrioles near the sperm nucleus; occurrence of filopodia in the spermatids and immature spermatozoa; unpolarised concentric structure of the mature spermatozoa; microtubule-like elements in spermatids and spermatozoa. The linhomoeins differ from their closest relatives, monhysterins (Xyalidae, Sphaerolaimidae), by total absence of MO at all the stages of spermatogenesis. This peculiarity is also found in the sperm development of some chromadorids, desmodorids and tylenchomorphs. In T. glabricutis the anterior testis produces large (20 μm) 'macrospermatozoa' of the linhomoeid pattern; the posterior testis produces much smaller (10 μm) 'microspermatozoa' of simplified structure. The female genital tubes also contain two types of amoeboid spermatozoa of different size and internal structure. Thus, T. glabricutis demonstrates a pattern of sperm dimorphism in which each testis of the diorchic male produces its own type of spermatozoa. Both types are found in the genital tube of fertilised females. Such sperm dimorphism is known from light microscope observations for several distantly related marine species although the biological significance of this dimorphism is obscure.

Partial formation of sperm dimorphism from spermatocytes of the cottoid fish, Hemilepidotus gilberti in cell culture

SummaryPolymorphism of sperm is considered to be significant for the reproductive strategy in some animal species. The phenomenon is thought to occur in the species-specific stage of spermatogenesis, but how the identical germ cells are differentiated towards polymorphic sperm remains unknown. We here performed a germ cell culture in the cottoid fish, Hemilepidotus gilberti, whose sperm exhibit dimorphism with fertilizable eusperm and unfertilizable parasperm. In the culture, germ cells, which were obtained with an identical morphology, a spherical shape of 5–7 µm in diameter, differentiated into smaller spherical cells with a single nucleus, a moving flagellum and localized mitochondria. In addition, large retroflex-shaped cells with two elongated nuclei were also observed in the cell culture. Germ cells that had each morphological feature were histologically also observed in some cysts of the spermatogenetic testis, suggesting that the former type of cell corresponded to developing eusperm and the latter corresponded to developing parasperm. When BrdU was incorporated into germ cells in the culture, it was detected in both cells with eusperm-like and those with parasperm-like morphologies. These findings suggest that DNA-duplicating spermatocytes are potent to autonomously progress a part of spermatogenesis to form dimorphic sperm.

Ultrastructural evidence of sperm dimorphism in Deladenus sp. (Tylenchomorpha: Sphaerularioidea: Allantonematidae)

The dimorphic spermatozoa of the insect-parasitic nematode Deladenus sp. (Tylenchomorpha: Sphaerularioidea: Allantonematidae) were studied for the first time with a transmission electron microscope (TEM). The immature spermatozoa from the testis of mycetophagous males are 10-12 μm diam. and 4-5 μm long unpolarised cells with a centrally located nucleus without a nuclear envelope. The cytoplasm contains mitochondria and specific components, membranous organelles (MO) and fibrous bodies (FB). The MO are spherical vesicles with an internal system of finger-like invaginations of the membrane; the spindle-shaped FB consist of tightly packed parallel fibres. Spermatozoa from the uteri of infective females of Deladenus sp. are vastly different in size being tiny cells ca 2 μm diam. with a spherical or oval nucleus. Each cell contains several mitochondria and MO. Although each individual of Deladenus sp. contains only monomorphic spermatozoa, sperm dimorphism was revealed after analysis of the whole life cycle. Despite a difference in size the cytological characters of both types of spermatozoa conform to the typical rhabditid pattern. The presence of both MO and FB in sphaerularioidid spermatozoa differentiates the superfamily Sphaerularioidea from Tylenchoidea whose representatives lack MO in the spermatogenic cells.

Riders on the sperm: sperm dimorphism and spermatozeugmata in nematodes from the genus Steinernema (Rhabditida: Steinernematidae)

A comparative ultrastructural study of the male gametes in entomopathogenic nematodes of the genus Steinernema (Rhabditida: Steinernematidae) is based on two species producing monomorphic (S. feltiae) and dimorphic (S. tami) spermatozoa. The spermatozoa of both species are basically the same as in many other Rhabditida. Immature spermatozoa from the testis are unpolarised cells containing a nucleus without a nuclear envelope, mitochondria and membranous organelles (MO), a unique component of the nematode spermatozoa. Mature spermatozoa have a nucleus, mitochondria and emptied MO, each attached to a sperm plasmalemma and opening to the exterior via a pore. Monomorphic mature spermatozoa of S. feltiae are 5 μm in diam., bear pseudopods, and form chains of several cells joined by gap junctions. The dimorphic immature spermatozoa of S. tami form spermatozeugmata where the single giant megaspermatozoon (30-35 μm diam.) bears numerous tiny, 3 μm microspermatozoa, intimately attached to the megaspermatozoon surface by gap junctions. Mature megaspermatozoa from the uterus are motile cells 35-50 μm diam.; they have prominent pseudopods and convey immotile, 2 μm microspermatozoa towards the oviduct. Unlike many other invertebrates, sperm dimorphism in S. tami does not change the basic set and structure of organelles, only their number. The genus Steinernema has two strategies for sperm agglomeration: i) chains of monomorphic spermatozoa of normal size, as in S. feltiae; and ii) spermatozeugmata based on a dramatic size difference between spermatozoa formed in the testis, as exemplified by S. tami. According to the types of sperm agglomeration, the genus Steinernema is divided into two main clades.