Giant Anteater

Giant anteater at San Diego Zoo.

3) Implantation

No early implantation stages of giant anteaters have been available to me. Walls (1939) described a placenta and fetus in the first trimester. The embryo was 10 mm long. There was rather more trophoblast covering the villi than at term. As is often true in early gestation, it was a two-layered structure, composed of cytotrophoblast and "plasmoditrophoblast" (syncytium). In his early specimen, the fetal capillaries had just begun to become visible within the villi. He found no yolk sac, nor did he describe a yolk sac membrane. Walls (1939) only found that the distal intestine protruded slightly into the umbilical cord. The allantoic "canal" (duct) had already nearly closed. What is difficult for me to understand is that he, and perhaps Wislocki (1928 - with a different anteater species, to be sure) interpreted the sinusoidal channels to be solid peripheral trophoblast. I interpret them to be composed of connective tissue with trophoblastic cover-i.e. peripheral extensions of the villi. Becher (1931) made the same observation as Walls and Wislocki. Walls described the uterine artery and vein pass through the center of the placenta, and also showed the "fusion" of villi with these peripheral sinusoidal trabeculae. Electronmicroscopy is needed to resolve the question.

Because of these findings and those by Enders (1960) on nine-banded armadillo, it may perhaps be justified to draw some analogies in interpreting the early anteater placental development. There are certain homologies to be found in these placentas: 1) the animals' classification (Xenarthra); 2) the uterine morphology (simplex uterus); 3) the fundal implantation; 4) and the similarities in the histologic appearance of the final placenta, especially the sinusoidal base. Please refer to the chapter on nine-banded armadillo. Even more important for a comparison, however, are the very detailed observations made by Becher (1931) on a large material of tamanduas (Tamandua tetradactyla), a more closely related species. There are still the same unresolved issues as to the nature and origin of the trabecular periphery of the placenta and its relation to the uterine wall. That is one reason for expanding on the topic slightly.

Enders (1960) summarized publications on armadillo placentation and added his own studies of various phases of the development of Dasypus novemcinctus. He divided these into three stages, the earliest of which is the formation of an inverted yolk sac placenta. Nothing of this structure is left at term (and it is doubtful, at least to me, that this exists in anteaters). He also emphasized the implantation of armadillos over the modified fundal endometrium with its unique "blood sinuses". These sinuses are also apparently present in the giant anteater placenta. The peripheral trabeculae, once interpreted to be solid trophoblast by Walls (1939), possess an internal connective tissue core with trophoblast on the surface. Wislocki (1928) in his description of a Cyclopes didactylus specimen (this is the "silky anteater") makes reference to this layer as having "…maternal tissue ….undergone a symplasmic transformation". Enders then described the broad trophoblastic cell columns and red cell precursors in the further placental development, all of which are also seen in the anteater. Moreover, as in the armadillo, the anteater lacks visible "Langhans' cells" (cytotrophoblast) on the surface of villi, and its placenta has the same very thin syncytial villous covers at term. It may be necessary to do electronmicroscopy to ascertain whether truly cytotrophoblast is absent at term. Cytotrophoblast, however, was shown to be present in young specimens, as derives from Walls' study (1939). I have reservations, however, when Enders makes reference to his interpretation that the connective tissue of chorion and villi derives from cytotrophoblast (and must state here that, in letter after reading this statement, Enders [2002] indicated that the paragraph suggesting this origin was badly written. What he meant to express is that "the chorionic plate was derived from what was originally the central region of the implantation site, not that its mesenchymal layer was derived from central trophoblast"). Also, the notion of the existence of "anastomosis" of villi cannot be confirmed (again Enders, in letters [2002], writes that he actually meant that the villi do not anastomose). I will make some more detailed remarks below of what connects villi that are seen only on flat sections.

4) General Characterization of the Placenta

Gross appearance of term placenta from a giant anteater.

Another term giant anteater placenta

Term placenta of giant anteater, fetal surface.

Maternal surface of giant anteater placenta.

I have had five placentas of giant anteaters available for study. One weighed 250 g, two of these weighed 270 g, another 200 g and the other 296 g without cord and membranes. They are shown above. They were disk-shaped, two having the appearance of a duplex organ, while the others had single disks measuring 16 x 14 x 1.5 cm. The maternal surface has a multicotyledonary appearance with little decidua covering the villous tissue. The latter was pale and very friable in consistency. There was no calcification. The membranes attached at the placental margin and the umbilical cords inserted near the center of the disk. The chorionic surface vessels were unusually prominent. The uterine attachment site is probably fundal in the unicornuate uterus of anteaters. The specimens available to me, however, did not show the "diffuse" placentation suggested by other authors. Rather, the anteater has a good chorioallantoic membrane that is not dissimilar to that of primates.

The maternal aspect of these placentas was similar to that of the nine-banded armadillos, in that a network of thin bundles of basal "sinusoidal network" was anchored to the villi, without intervening endometrial glands or decidua. Thus, the uterus must have similar implantational characteristics to that of armadillos. Two large maternal vessels penetrated deep into the villous tissue of my specimens, as was shown already by Walls (1939).

Complete cross section of term giant anteater placenta.

Because of the unusual complexity of the basal portions of the villous tissue, it is best to illustrate this region next. This portion of the placenta is basal and peripheral, but is also interspersed with villous tissue. Once interpreted as solid (cyto-) trophoblast, I view it more as peripheral placental tissue into which the villi may expand. This region is often described, and quite different interpretations are made of its origin. Is it "solid trophoblast", "syncytial formation of endometrial epithelium", "corpus cavernosum of uterus", and perhaps a mixture of all? It is clear from this diversity of opinions that the trabecular region is still poorly understood. What is needed now is a specific genetic analysis of this trabeculae, employing cytogenetic identification of sex chromosomes in male fetuses, or specific trophoblast/endothelium staining with antibodies. It is very important, however, to be cognizant of the fact that this layer is shed with the placenta at delivery. Thus, an origin from endo- myometrium seems to me is unlikely. Sonographic study of the pregnant uterus in any of these species would also be of interest. It may better reveal the pattern of blood flow.

To be in a better position of understanding this trabecular tissue, the following sections were stained with Masson's trichrome stain. They show that these trabeculae have a central (blue) connective tissue core, upon which rests the presumably single-layered trophoblast. These trophoblastic cells are much more plump than those covering the villi. In the regions shown with arrows, the peripheral villi including fetal capillaries infiltrate the trabeculae and then expand into them.

Trichrome stain of trabecular region at base of placenta. At the arrow points, one can see the beginning of villous expansion into the solid trabeculae.

Higher magnification of similar areas with fetal capillaries (F.V.), solid trabeculae (T) and transition of villus into trabeculae at arrow. (V=villus).

Higher magnification of similar areas with fetal capillaries (F.V.), solid trabeculae (T) and transition of villus into trabeculae at arrow. (V=villus).

5) Details of fetal/maternal barrier

This is a hemochorial, villous placenta. The villous syncytial trophoblast is in direct contact with the maternal blood. Mossman (1987) considered this placenta to be intermediate between labyrinthine and villous types. I believe it to be more typical of a hemochorial villous placenta, but with an unusual basal portion of sinusoidal tissue, much as is found in armadillos. The labyrinthine notion probably stems from the description of villous anastomoses (see Wislocki, 1928). But Wislocki was also quite adamant that this labyrinth is very dissimilar from the placental labyrinth of Callitrichids. The villi are anchored to the thin lamellae of the spongy sinusoids, which do not lose their probably trophoblastic surfaces, as Enders pointed out, although he called them endothelial cells. This is also shown in the sections here presented. Villi also intermingle diffusely with this spongy region of the placenta.

I found no direct "anastomoses" between villi, as were described by Enders (1960, see comment by Enders [2002] above) in armadillo placentas. The trabeculae connect some villi, as shown in flat sections through the placenta. No cytotrophoblast is apparent in the term placentas, but it is present in the first trimester. A similar misinterpretation of the absence of cytotrophoblast was made for the term human placenta in the past, until electromicroscopy showed its presence.

The surface of the term gestation villi has a very thin layer of syncytium. It is striking to observe, how close the fetal capillaries come here to the trophoblastic epithelium, a feature upon which Wislocki already commented. He also drew attention to the unusual, perhaps unique, large cells that cluster around the larger fetal villous blood vessels. He likened their appearance to that of decidual cells but was uncertain of their function. They are certainly similarly prominent in the current species under consideration.

The villous tissue is thicker and much more pronounced in the central part of the placenta. At its edges, it is thinner and is eventually completely replaced by the sinusoidal tissue.

Complete thickness of mature placenta from a giant anteater gestation. Note the basal sinusoidal region with persistent epithelial covers. Villi attach and intersperse with these lamellae. They were once interpreted to be solid trophoblast.

Higher magnification of villi to show the thin syncytium of a term placenta. Note that the fetal capillaries nearly reach the villous surfaces. IVS=intervillous space; NRBC=nucleated red blood cell. Note also the large perivascualr cells of the villous stroma.

Higher magnification of villi to show the thin syncytium of a term placenta. Note that the fetal capillaries nearly reach the villous surfaces. IVS=intervillous space; NRBC=nucleated red blood cell. Note also the large perivascualr cells of the villous stroma.

6) Umbilical cord

The umbilical cords of our placentas inserted near the centers of the disks and had very few twists. They measured 10, 16, 20, 21.5 and 27 cm in length of these delivered placentas. It is unknown how much cord remained on the neonates, all of whom survived for several days. There are two umbilical arteries and one vein in the umbilical cord. There are no ducts; only a minute remnant of a former allantoic duct was seen in one cord. The surface is a single-layered flat squamous amnionic epithelium. Walls (1939) commented upon the anteater cord as being very similar to that of human placentas, with which I concur.

7) Uteroplacental circulation

No data are known to me of the manner of entry of two large maternal blood vessels into the center of the placenta. This unusual vascularization is supported by the description of an early anteater pregnancy reported by Walls (1939). On the other hand, the congruence of the sinusoidal spaces with the intervillous space is often commented upon.

Section taken from near the margin of the anteater placenta, as the villous tissue (left) becomes less prominent and the sinusoids (right) take over. FV=fetal vessel; IVS=intervillous space.

8) Extraplacental membranes

These are composed of amnion, with an occasionally tall epithelium, and the chorion. The two layers are fused; there is no allantoic sac. No yolk sac remnants are found. The amount of decidua capsularis is extremely variable. Walls (1939) found it to be absent in most areas. On macroscopic examination, the membranes of my recent placenta had a gelatinous consistency.

Membranes of term placenta. The left photograph merely shows a very dense, cellular chorion, little decidua capsularis.

Membranes of term placenta. There is some decidua capsularis attached to the chorion.

9) Trophoblast external to barrier

There probably is very little apparent trophoblastic invasion, as the villi implant on modified basal sinusoids. This region is shed with the placenta. But no intact uterus with an implanted placenta has been available and judgment must be withheld about the possible uterine trophoblastic invasion.

At the site of attachment, there is little decidua basalis next to the sinusoidal region. The basal decidua is intermingled with some trophoblast. Rare glands are present. There are also some hyalinized villi.

10) Endometrium

There is little decidua shed with the delivered placenta. Moreover, Walls (1939) found virtually no decidua vera, and only very little deciduas capsularis in his first trimester specimen. Little decidual tissue, intermixed with some trophoblast, is found in the delivered placenta.

11) Various features

No other significant observations were made.

12) Endocrinology

Ovarian cycles and a pregnancy were examined by fecal steroid analysis only once (Patzl, et al., 1998). These investigators found the onset of ovarian cyclic activity after pregnancy to occur within 4-11 weeks. The pregnancy lasted 184 days. The average length of ovarian cycles was 51 days. Progestogens rose above the cyclic luteal activity in the second half of pregnancy and were 20 times higher during the week before parturition. Estrogens rose significantly in the last third of gestation and, especially, shortly before birth.

A detailed hormonal study of cycles in six animals and study of pregnancy diagnosis comes from the doctoral work of Nicole Schauerte (2005). Cycles were not seasonal and polyestrous with slight vaginal bleeding occurring in the pro-estrus. Ultrasonic determination of pregnancy was also documented.

Additional observations on the endometrium by Becher (1931) are useful. In his large study of pregnant and nonpregnant uteri of four-toed tamanduas, Becher described the cyclical change of the endometrium. It had many similarities with human endometrium, and periods of estrus were described with marked secretion of mucus from the vagina. In the later stages of the endometrial cycle, the photographs suggest that there is a decidua-like change of the endometrial stroma. Much extravasation was reported in later stages of the cycle and in early pregnancy, Becher described a thick decidua. He also correlated the endometrial changes with the state of ovarian activity and described the appearance of the corpus luteum.

13) Genetics

Giant anteaters have 60 chromosomes (Hsu, 1965). We can confirm this from our own study of four animals. No other chromosomal studies have been published. Garcia et al. (2005) studied and characterized six microsatellite markers that may become useful for studies of population and paternity.

14) Immunology

There are no published reports.

15) Pathological features

A small number of giant anteaters have been autopsied and only a few specific lesions were identified. Baskin et al. (1977) found infection with Yersinia pseudotuberculosis, presumably from contamination of feed by rats or pigeons. Diniz et al. (1995) identified digestive disorders as most prominent features, with many other systemic diseases being less common (infection, nutritional, respiratory, etc.). Parasites of one kind or another were found in 48% of fecal samples. Griner (1983) listed trauma as a major problem in captivity (tapir-induced neonatal mortality), and arteriosclerosis.

RNA viruses (Picobirnavirus) were found in the fecal study by Haga et al. (1999). Labruna et al. (2002) found numerous ticks (Amblyomma sp. etc.) on these animals as well as many other species from Parana and Mato Grosso. Freitas et al. (2006) recovered Eimeria species in giant anteaters.

Coke et al. (2002) documented gastric perforation (Entameba and Acanthamoeba spp.) in an animal with peritonitis (after perforation) and cardiac myopathy (dilatation).

16) Physiologic data

There are no reports to my knowledge.

17) Other resources

Fibroblast cell strains of four animals are available from the "Frozen Zoo" at CRES of the San Diego Zoo by contacting Dr. Oliver Ryder at oryder@ucsd.edu.

18) Additional remarks and needs for future studies

The contribution by Becher (1931) on Tamandua tetradactyla is important in this context. In the first place, Becher had an unusually large material of pregnant and nonpregnant uteri available. But, more importantly, he described the material as having been well fixed for histologic studies. In addition, he had several stages of fetal/placental development available that provide more insight than the study of one particular stage. He pointed out that this is perhaps the reason why his findings differ in many ways from the interpretation of Wislocki's single specimen of a two-toed anteater (1928).

I will summarize some salient conclusions from this related species of anteater that was studied by Becher. These remarks are probably equally applicable to the placenta of the giant anteater. The placenta was always implanted at the fundus and, with advancing gestation, the uterine wall became very thin in the region of nidation. In the fundus, a vascular network developed in the musculature that was visible externally. Becher spoke of a "corpus cavernosum". The maternal surface of the placenta had a finely lobulated appearance. No yolk sac and no allantois were observed, even in the earliest stages of pregnancy (that is quite different from what Enders described for the armadillo). The umbilical cord inserted centrally and the lengths of some cords were given for immature stages. Becher and other observers reported that the periphery of the placental disk has a trough. Perhaps this led to the suggestion that it has a bell shape. The study presented an interesting flat section through the organ. This displayed the "mesodermal villi" (his choice of words) to be connected to one another by the solid trophoblastic trabeculae. These, Becher found (in contrast to Wislocki) are composed of large cytotrophoblast and contain no connective tissue. He found these sheets of trabeculae to become hyalinized toward term and interpreted them not to become zones of possible future expansion of the villous structures. Their plump cells have no microvillous surface. Most interesting to me is his observation that the placenta shed in the "muscular" (!) cavernous zone (not in the decidua).

There is much work to be done on the nature of this peripheral zone of the xenarthran placentas. Most important will be the identification (maternal vs. fetal) of the individual trabecular elements by genetic means. Electronmicroscopy will be mandatory also to better understand the contribution of the trabeculae to the placental development and its function.

Acknowledgement

Most animal photographs of this book come from the Zoological Society of San Diego. I appreciate their help and that of the pathologists at the San Diego Zoo.

References

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Cell strains from CRES at: www.sandiegozoo.org

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