Spermatophoric reaction reappraised: Novel insights into the functioning of the loliginid spermatophore based on Doryteuthis plei (Mollusca: Cephalopoda)

During copulation, spermatophores produced by male coleoid cephalopods undergo the spermatophoric reaction, a complex process of evagination that culminates in the attachment of the spermatangium (everted spermatophore containing the sperm mass) on the female's body. To better understand this complicated phenomenon, the present study investigated the functional morphology of the spermatophore of the squid Doryteuthis plei applying in vitro analysis of the reaction, as well as light and electron microscopy investigation of spermatangia obtained either in vitro, or naturally attached on females. Hitherto unnoticed functional features of the loliginid spermatophore require a reappraisal of some important processes involved in the spermatophoric reaction. The most striking findings concern the attachment mechanism, which is not carried out solely by cement adhesive material, as previously believed, but rather by an autonomous, complex process performed by multiple structures during the spermatophoric reaction. During evagination, the ejaculatory apparatus provides anchorage on the targeted tissue, presumably due to the minute stellate particles present in the exposed spiral filament. Consequently, the ejaculatory apparatus maintains the attachment of the tip of the evaginating spermatophore until the cement body is extruded. Subsequently, the cement body passes through a complex structural rearrangement, which leads to the injection of both its viscid contents and pointed oral region onto the targeted tissue. The inner membrane at the oral region of the cement body contains numerous stellate particles attached at its inner side; eversion of this membrane exposes these sharp structures, which presumably adhere to the tissue and augment attachment. Several naturally attached spermatangia were found with their bases implanted at the deposition sites, and the possible mechanisms of perforation are discussed based on present evidence. The function of the complex squid spermatophore and its spermatophoric reaction is revisited in light of these findings. J. Morphol. 2012. © 2011 Wiley Periodicals, Inc.     

Detailed developmental morphology of the spermatophore of the Mediterranean field cricket, Gryllus bimaculatus (De Geer) (Orthoptera: Gryllidae)

The detailed developmental morphology of the spermatophore of a cricket (Gryllus bimaculatus, De Geer) is described for the first time. A specific behavioural reference point is identified from which time developmental processes proceed in a more or less robotic manner. The structures previously identified as forming part of the fully formed spermatophore, that is, the ampulla, the sperm tube and the attachment plate are all identified in detail during their development. Interestingly the attachment plate material is extruded as an amorphous mass from the ejaculatory duct prior to its insertion into the dorsal pouch, the sperm do not enter the sperm sac until the end of spermatophore formation, and the sperm tube itself is formed from a flattened layer of material which rolls up into a tube and involves the use of a die mechanism.     

Spermatangium formation and sperm discharge in the Japanese pygmy squid Idiosepius paradoxus

In cephalopods, sperm discharge is an important event not only for sperm transfer but also influencing sperm storage capacity of attached spermatangia (everted spermatophores). To investigate sperm discharge from spermatangia and the condition of naturally attached spermatangia in Japanese pygmy squid (Idiosepius paradoxus) we (i) investigated the morphology of spermatophores and spermatangia, and the process of spermatophore evagination and sperm discharge from spermatangia obtained in vitro; (ii) observed spermatangia that were naturally attached to female squids at 6, 12, 18, 24 and 48 h after copulation to investigate alterations in naturally attached spermatangia with time. The spermatophore of I. paradoxus is slender and cylindrical and consists of a sperm mass, a cement body and an ejaculatory apparatus, which is similar to those of loliginid squids. The spermatangium is fishhook-shaped, its distal end being open and narrow. After the spermatangium is formed, the sperm mass gradually moves to the open end of the spermatangium, from where sperm are released. Sperm discharge is a rapid process immediately after the beginning of sperm release, but within 5 min changes to an intermittent release of sperm. Although the volume of residual spermatozoa differed among spermatangia that were naturally attached to a single individual, the probability that spermatangia would be empty increased with time. Most naturally attached spermatangia discharged almost all of their spermatozoa within 24 h after copulation, and no spermatangia were attached to females 48 h after copulation. These results suggest that sperm transfer from the spermatangium to the seminal receptacle must occur within 24 h, and that the spermatangium functions as a transient sperm storage organ in pygmy squids.     

Fine structure of the spermatophores and their ejaculated forms,sperm reservoirs,of the Japanese common squid,Todarodes pacificus

Spermatophores in a squid, Todarodes pacificus, were observed by light and electron microscopy and were further analyzed by X-ray microanalysis (XMA) of frozen thin sections. Each spermatophore consists of a sperm mass, a cement body, an ejaculatory apparatus, and some fluid materials, all of which are covered by an outer tunic. The outer tunic consists of about 20 membranous layers, each containing straight, parallel microgrooves. Each layer's microgroove pattern is roughly in an orthogonal arrangement with respect to the next layer's pattern. The sperm mass, which is the only cellular component, consists of a sperm rope which is coiled more than 500 times. Most of the spermatozoa in the rope are arranged regularly and are enveloped in materials which are well-stained by Alcian blue. The cement body is located between the sperm mass and ejaculatory apparatus and has a hard outer shell with an arrowhead-like structure, presumably for penetration into the tissue of the female. Calcium and phosphorus are present in the shell of the cement body, which also has an affinity for alizarin red. The ejaculatory apparatus consists of two tubes, designated as the inner tunic and the inner membrane. After the spermatophoric reaction, a sperm reservoir is formed at the anterior end of the extruded and inverted ejaculatory apparatus. The sperm reservoir, which encases the sperm mass, is composed of the cement body at the anterior end and the inner tunic of the ejaculatory apparatus at the posterior end.     

Fine structure and histochemistry of the freshly extruded and hardened spermatophore of the spiny lobster,Panulirus interruptus

Freshly extruded and hardened spermatophores of the spiny lobster, Panulirus interruptus, were compared using light and electron microscopy (EM). The spermatophore is composed of a sperm tube embedded in an acellular matrix. The sperm tube consists of tightly packed spherical cavities in an acellular material within which the sperm lie. The extruded spermatophore is white, soft, and sticky on all surfaces. The highly coiled sperm tube can be seen near the surface of the foot of the spermatophore, which is the side that will attach to the exoskeleton of the female. The opposite surface, the cap, will harden and darken after exposure to seawater. In the soft spermatophore, the matrix surrounding the sperm tube and extending from foot to cap is composed of small (2-μm) granules embedded in a loose weave of filaments. In the hardened spermatophore, the matrix is composed of small (4-μm) empty spheres. At the cap region the matrix darkens, and at the foot the granules dissolve to form a thick layer characterized by vertical striations. The structure of this spermatophore is compared to those spermatophores of other decapods that have been described at the EM level. The chemical composition and possible function(s) of the various components are discussed.     

Unraveling the structure of squids’ spermatophores: a combined approach based on Doryteuthis plei (Blainville, 1823) (Cephalopoda: Loliginidae)

Marian, J.E.A.R. and Domaneschi, O. 2012. Unraveling the structure of squids’ spermatophores: a combined approach based on Doryteuthis plei (Blainville, 1823) (Cephalopoda: Loliginidae). —Acta Zoologica (Stockholm) 93 : 281–307. Male coleoid cephalopods produce elaborate spermatophores, which function autonomously outside the male body during copulation, undergoing a complicated process of evagination. In order to contribute to the understanding of this unique structure, this study investigated the morphology of the spermatophore of Doryteuthis plei applying several microscopy techniques. A hitherto unreported, much more complex structural arrangement was revealed for the loliginid spermatophore, the most striking findings being: (1) the complex, layered structure of the middle membrane, which bears an additional, chemically distinct segment surrounding part of the cement body; (2) the presence of a space between the inner tunic and middle membrane filled with a fine reticulated material; (3) the presence of stellate particles not only embedded in the spiral filament, but also closely applied to the inner membrane at the level of the cement body; (4) the presence of a pre‐oral chamber in the cap region; and (5) the complex organization of the cement body, formed by two distinct layers encompassing contents of different chemical and textural properties. Careful literature reassessment suggests several of these features are common to loliginids, and to some extent to other squids. Their possible functional implications are discussed in light of our knowledge of the spermatophoric reaction mechanics.     

Two intromittent organs in Zorotypus caudelli (Insecta,Zoraptera): the paradoxical coexistence of an extremely long tube and a large spermatophore

Very unusual genitalia of the species Zorotypus caudelli are described. It contains the unique configuration of two different intromittent organs, one of them strongly elongated. Hyper elongated genitalia are known in different groups of insects. Males have to accommodate these unwieldy structures in the limited spaces of the abdomen and manipulate them acutely during copulation. A crucial question is how do species with elongated genitalia cope with these requirements? To investigate this, we studied key features enabling storage, insertion, and withdrawal of the elongated genitalia. The co‐existence of an elongated narrow tube and a bulky spermatophore is a highly unusual and apparently paradoxical condition. However, we demonstrate that the tube is not involved in sperm transmission, whereas the large spermatophore is transferred to females by a membranous fold of the genitalia. The movement of the spermatophore is caused by haemolymph pressure, which likely also promotes the insertion of both intromittent organs. A comparison with the genital anatomy and reproductive mode in related groups suggests that the elongated tube and its accommodating pouch is a de novo structure, and that the ancestral sperm transport via spermatophore is a preadaptive condition for the acquisition of this unusual structure. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112 , 40–54.     

The ultrastructure of the ejaculatory duct in the springtail Orchesella villosa (Geoffroy) (Hexapoda, Collembola) and the formation of the spermatophore

The initial part of the ejaculatory duct of Orchesella villosa contains a “valve” and a “sorter” avoiding respectively the reflow and allowing the separation of the secretion for the spermatophore stalk from the sperm fluid. For most of its length, the ejaculatory duct lumen is divided into two parts: in the dorsal part the sperm fluid flows while in the ventral district the secretion for the stalk occurs. Laterally, on both sides of the duct, longitudinal muscle fibers are present. The epithelium of the dorsal region consists of two types of long secretory cells; the most peculiar of them are those provided with extracellular cisterns flowing directly into the duct lumen as it occurs in 1st type of epidermal cells. These cells could be involved in the control of the viscosity of the sperm fluid. The second type of cells produce a secretion probably involved in the formation of the outer coat of the apical sperm droplet. The ventral epithelium consists of short cells contributing to the enrichment of the secretion for the spermatophore stalk and perhaps also to the viscosity of the secretion flowing in the lumen. In the distal part of the ejaculatory duct, the ventral district is provided with a thick layer of muscle fibers and with 3 + 3 cuticular laminae dividing the lumen into a series of slits through which the secretion of the stalk is squeezed out into filaments. This organization allows the twisting and hardening of these filaments. A drop of sperm fluid is laid on top of the long and rigid spermatophore stalk.     

The role of cercal sensory feedback during spermatophore transfer in the cricket, Acheta domesticus

The role of the cerci in the spermatophore transfer behavior of the cricket Acheta domesticus was examined. During transfer, the male cerci were held close to the female abdomen where they produced small flicking movements. Male cercal ablation significantly decreased mating success by reducing both the ability of the male to hook the epiphallus on to the female subgenital plate and to transfer the spermatophore. During spermatophore transfer, the male must thread the spermatophore tube into the female genital papilla and attach the spermatophore, via its attachment plate, to the base of the ovipositor. Extracellular recordings from the male genital nerve revealed that a centrally driven, rhythmic bursting activity of genital efferents produced the rhythmic contractions of the five pairs of genital muscles responsible for spermatophore threading. Tactile stimulation of campaniform sensilla on the medial aspect of each cercus inhibited the activity of those motor units responsible for advancing the spermatophore tube during threading, while simultaneously activating the motor units responsible for adjusting the position of the epiphallus. We conclude that mechanosensory neurons on the cerci of the male cricket supply important information on female position to the motor program responsible for spermatophore threading and transfer.     

Condition-dependent spermatophore size is correlated with male's age in a bushcricket (Orthoptera: Phaneropteridae)

During mating, male bushcrickets transfer a spermatophore that consists of a sperm-containing ampulla and a product of the accessory glands, the spermatophylax, which females directly ingest. In the present study, we demonstrate male spermatophore allocation in the bushcricket Poecilimon zimmeri . Males of this species show condition-dependent spermatophore investment. This investment depended upon the age at first mating of males, with older individuals transferring larger spermatophores than younger ones of the same body mass. Independently of age, heavier males transfer larger spermatophores, but the size of males (as measured by femur length) was not a good predictor. Heavier males allocate a lower proportion of their mass to spermatophores and reach their maximal investment point earlier than less heavy males. Spermatophylax production levelled off to a species specific maximum earlier than that of sperm investment (measured as ampulla mass), suggesting that males face high levels of sperm competition.  © 2009 The Linnean Society of London, Biological Journal of the Linnean Society , 2009, 96 , 354–360.     

Spernathecal morphology,sperm transfer and a novel mechanism of sperm displacement in the rove beetle,Aleochara curtula (Coleoptera,Staphylinidae)

Summary The mechanism by which sperm are transferred from the male's spermatophore to the female's storing cage is described for the rove beetle Aleochara curtula, emphasizing a novel mechanism of sperm displacement by competing males. The cuticular, U-shaped spermatheca is equipped with a valve structure and two sclerotized teeth. The tube of the spermatophore extends into the spermathecal duct through the guidance of the flagellum of the male endophallus. Further elongation of the spermatophore tube, however, occurs only after separation of the pair. A primary tube bursts at its tip after passing through the valve. Within the lumen of the primary tube, a second tube passes through the valve and continues to extend up to the apical bulb of the spermatheca, doubles back on itself and swells to form a balloon filling most of the spermatheca. The balloon of the spermatophore is pierced within the spermatheca by tooth-like structures pressed against the spermatophore through contraction of the spermathecal muscle. The same process of spermatophore growing and swelling is also observed in mated females. Sperm from previous copulations are backflushed through the valve and the spermathecal duct, indicative of last-male sperm predominance.Abbreviations ad adhesive secretion covering the sperm - sac am amorphous secretion of the spermatophore - as ascending portion of the spermatophore - ds descending portion of the spermatophore - end parts of the male endophallus - ext extended tube - f flagellum - gs genital segment - lt large tooth - m muscle of the spermatheca - nsc non sclerotized cuticle - op opening of the spermathecal gland - pt primary tube - sc sclerotized cuticle - sd spermathecal duct - se secretion of the spermathecal gland - sf secretion flowing out of the primary tube - sg spermathecal gland - sm sperm - smt small tooth - sp spermatheca - ss sperm sac - st secondary tube - vm vaginal muscle     

The structure and maturation of the spermatozoa of Sciara coprophila

The axial filament of Sciara coprophila does not conform to the usual 9 + 2 filament pattern but consists, rather, of as many as 76 pairs of filaments which decrease in number from the anterior to the posterior region of the sperm. It is first seen at the base of the head in the shape of an indented oval. The axial filament varies in configuration along the remaining length of the sperm as one whorl or two connected whorls of filament pairs. The other structures of the sperm revealed by the light and electron microscopes are a homogeneous, dense, spear-shaped nucleus, a row of spherical dense bodies in the middle piece enclosed by the axial filament and of unknown nature and function and a single mitochondrial derivative. The mitochondrial nebenkern derivative consists of a large electron transparent region bordered by cristae and a smaller paracrystalline region located adjacent to the axial filament. The derivative arises as paracrystalline material in a medial nuclear indentation. The electron transparent material is first seen at the anterior end of the middle piece. Unlike other known insect sperm, but reminiscent of sperm capacitation in mammals, sperm maturation is completed in the spermathecae of Sciara 7 to 9 hours after insemination. It consists of the acquisition of sperm motility and elimination of the electron transparent region of the mitochondrial nebenkern derivative. The electron microscope reveals in mature sperm that the axial filament doublets have changed configuration and consist of a single whorl which encloses the paracrystalline rod. The process by which the major portion of the nebenkern derivative is eliminated occurs in four identifiable stages. Since sperm maturation does not appear to be intrinsically controlled, factors in the spermathecal fluid may play a role in its completion.     

The male genital tract and aedeagal components of the Diptera with a discussion of their phylogenetic significance

The male genital tract of Diptera is surveyed, based on whole dissections, with emphasis on nematocerous infraorders and especially the Culicomorpha. The genital tracts of all available nematocerous families are described and illustrated, including, for the first time, those of the families Thaumaleidae, Dixidae, Hesperinidae, and Pachyneuridae, and, as an outgroup, the mecopteran family Nannochoristidae (Mecoptera). On the basis of outgroup comparison, the Diptera + Mecoptera + Siphonaptera share the synapomorphies of accessory gland continuous with the vas deferens and U-shaped vasa deferentia. Further synapomorphies of the male genital tract provide evidence of the monophyly of the Mecoptera + Siphonaptera, Boreidae + Siphonaptera, and the Blephariceridae + Psychodidae + Trichoceridae + Anisopodidae + Brachycera. The Bibionomorpha, Culicomorpha, and Ptychopteromorpha share the synapomorphy of paired, two- to three-chambered accessory gland complexes, with secondary losses in Corethrellidae + Culicidae. It is concluded that there is no convincing evidence for the monophyly of the Chironomoidea. A sperm pump with an ejaculatory apodeme and the absence of a spermatophore are considered to be part of the ground plan of Diptera, but, because of uncertainty as to the identity of the basal lineage within Diptera and the homology of the sperm pump outside of Diptera, the polarity of these features cannot be presently interpreted.  © 2007 The Linnean Society of London, Zoological Journal of the Linnean Society , 2007, 150 , 711–742.     

Unusual granules in the ejaculatory duct of a Microphthalmus species (Polychaeta,Annelida)

Summary The paired prominent ejaculatory ducts of the hermaphroditic polychaete Microphthalmus cf. listensis are surrounded by gland cells the processes of which penetrate the ducts themselves. These cells produce, in separate regions, two different types of spherical granules. Type I is composed of an electron dense and an electron lucent part. Type II granules contain a tubular filament that forms a single or double spiral in the periphery of a more or less unstructured electron dense material. Golgi vesicles give rise to this granule type. During the passage of sperm, these granules are obviously discharged into the lumen of the duct. Here they change form and probably dissolve. Their function is as yet unknown; capacitation of sperm is assumed.     

FEMALE CONTROL OF SPERM TRANSFER AND INTRASPECIFIC VARIATION IN SPERM PRECEDENCE: ANTECEDENTS TO THE EVOLUTION OF A COURTSHIP FOOD GIFT

Manipulation of ejaculates is believed to be an important avenue of female choice throughout the animal kingdom, but evidence of its importance to sexual selection remains scarce. In crickets, such manipulation is manifest in the premature removal of the externally attached spermatophore, which may afford females an important means of postcopulatory mate choice. We tested the hypothesis that premature spermatophore removal contributes significantly to intraspecific variation in sperm precedence by (1) experimentally manipulating spermatophore attachment durations of competing male Gryllodes sigillatus and (2) employing protein electrophoresis to determine the paternity of doubly mated females. The relative spermatophore attachment durations of competing males had a significant influence on male paternity, but the pattern of sperm precedence deviated significantly from the predictions of an ideal lottery. Instead, paternity data and morphological evidence accorded best with a model of partial sperm displacement derived here. Our model is similar to a displacement model of Parker et al. in that sperm of the second male mixes instantaneously with that of the first throughout the displacement process, but the novel feature of our model is that the number of sperm displaced is only a fraction of the number of sperm transferred by the second male. Regardless of the underlying mechanism, female G. sigillatus can clearly alter the paternity of their offspring through their spermatophore-removal behavior, and employ such cryptic choice in favoring larger males and those providing larger courtship food gifts. We discuss how female control of sperm transfer and intraspecific variation in sperm precedence may be important precursors to the evolution of gift giving in insects.     

Delayed spermatophore removal in the land snail Euhadra peliomphala

Although many studies have demonstrated that variation in male genital morphology can influence male fertilization success, it remains unclear in many cases which mechanism(s) of sexual selection and sexual conflict produce the morphological divergence of genitalia. Distinguishing between such mechanisms requires, at the very least, information on the process by which genital morphology influences fertilization success. The length of the spermatophore‐producing organ (SPO) of the stylommatophoran land snails is a case in point. The length of the spermatophore may have an important influence on the transfer of the spermatophore to the recipient's genital tract. To test this hypothesis, we investigated in Euhadra peliomphala (Pfeiffer, 1850) the relationship between the length of the SPO in sperm donors and the position of their spermatophore in sperm recipients at 72, 84, and 96 h after mating. The results showed that longer spermatophores required longer times to reach a gametolytic organ and to begin to be digested. These findings suggest that this delay in spermatophore digestion explains the mechanism for the high fertilization success shown by snails with a long SPO. © 2013 The Linnean Society of London     

Spermatophore feeding and mating behaviour in praying mantids (Mantodea: Liturgusidae)

In a number of insects, males transfer sperm to females via an externally attached spermatophore, which females subsequently remove and consume. Here I report the first study of spermatophore feeding in a praying mantid. While studies of praying mantid mating behaviour have largely focused on sexual cannibalism, this behaviour is not known in the genus Ciulfina . The general patterns of mating behaviour and spermatophore feeding are described for four species: Ciulfina rentzi , Ciulfina klassi , Ciulfina biseriata and Ciulfina baldersoni . Copulation duration and postcopulatory spermatophore attachment duration were found to vary both inter- and intraspecifically. Ciulfina rentzi exhibited a considerably longer mean copulation duration and a considerably shorter mean postcopulatory spermatophore attachment duration than the other Ciulfina species. Smaller males copulated for longer durations in C. rentzi and C. klassi . For C. baldersoni , both copulation duration and postcopulatory spermatophore attachment duration increased with female mass. The genus Ciulfina is highlighted as an intriguing new system in which to test hypotheses surrounding the function of spermatophore feeding and the significance of copulation duration.     

Female alpine newts (Triturus alpestris) mate initially with males signalling fertility benefits

Gametic asymmetry implies that females invest more per gamete than males do and thus sperm is considered to be a relatively cheap resource. However, contrary to this classic view, sperm has been shown to be frequently in short supply; hence, selection favouring females that mate for fertility benefits should occur. For this reason, we determined whether males signalling fertility are preferred by female newts of the species Triturus alpestris . We performed paired female–male trials using unmated and previously inseminated females to determine potential criteria for female interest in a courting male, to establish what factors lead to successful mating and to assess the importance of female choice for direct and indirect benefits. We found that female interest in any potential mate and mating success decreased once mating had occurred. Furthermore, we detected an increase in spermatophore deposition rate and rapid spermatophore transfer in encounters that resulted in a successful mating. The results obtained indicate that female alpine newts are attracted to males showing signs of relatively high fertility and that females exhibit a decreased propensity to mate once initial sperm reserves have been acquired. Our results support the theory of initial female choice for fertility benefits.  © 2007 The Linnean Society of London, Biological Journal of the Linnean Society , 2007, 91 , 483–491.     

Floral morphogenesis in Sinofranchetia (Lardizabalaceae) and its systematic significance

The floral organs of Sinofranchetia chinensis Hemsl. (Lardizabalaceae) are all spiral in initiation. Stamen and petal (nectar‐leaf) primordia initiate independently and are different in shape. The petals and three stamens in the first whorl are retarded in the early developmental stages. The carpel primordia are conduplicate; the stigma is formed around the upper part of the ventral slit and the style is not differentiated. The functionally unisexual flowers are bisexual in organization in the early developmental stages. The development of the flowers on the inflorescence is spiral and centripetal. Some floral characteristics of Sinofranchetia appear to be plesiomorphic in Lardizabalaceae. © 2009 The Linnean Society of London, Botanical Journal of the Linnean Society, 2009, 160 , 82–92.     

Molecular dissection of nuptial gifts in divergent strains of Ostrinia moths

Seminal fluid proteins (SFPs) produced in the male accessory glands and ejaculatory duct are subject to strong sexual selection, often evolve rapidly and therefore may play a key role in reproductive isolation and species formation. However, little is known about reproductive proteins for species in which males transfer ejaculate to females using a spermatophore package. By combining RNA sequencing and proteomics, we characterize putative SFPs, identify proteins transferred in the male spermatophore and identify candidate genes contributing to a one‐way gametic incompatibility between Z and E strains of the European corn borer moth Ostrinia nubilalis. We find that the accessory glands and ejaculatory duct secrete over 200 highly expressed gene products, including peptidases, peptidase regulators and odourant‐binding proteins. A comparison between Ostrinia strains reveals that accessory gland and ejaculatory duct sequences with hormone degradation and peptidase activity are among the most extremely differentially expressed. However, most spermatophore peptides lack reproductive tissue bias or canonical secretory signal motifs and aproximately one‐quarter may be produced elsewhere before being sequestered by the male accessory glands during spermatophore production. In addition, most potential gene candidates for postmating reproductive isolation do not meet standard criteria for predicted SFPs and almost three‐quarters are novel, suggesting that both postmating sexual interactions and gametic isolation likely involve molecular products beyond traditionally recognized SFPs.