East African River Hippopotamus
Hippopotamus amphibius kiboko
Pygmy Hippopotamus
Choeropsis (Hexaprotodon) liberiensis

Order: Artiodactyla
Family: Hippopotamidae

1) General Zoological Data

Hippopotamuses belong to the suborder Suina, a varied group of artiodactyls that were summarized aptly by Macdonald and Bosma (1985) thus: "Apart from a number of extinct groups, the artiodactyl suborder Suina comprises the families Suidae (pigs), Tayassuidae (peccaries) and Hippopotamidae (hippopotami) (Romer, 1966). The species which make up these families have several tens of millions of years of separate evolutionary history and demonstrate wide variation in body size, external and internal structure, behaviour, environmental adaptation, and geographical distribution. Nevertheless, they are brought together in one suborder, mainly because they share a primitive structure of the limbs and dentition. In addition, they have no horns or antlers. With regard to the soft parts, they have a relatively simple, non-ruminating stomach (Langer, 1973)." One can add to this that they also share very similar placentas.

The term hippopotamus derives from hippos=horse and potamos=river (Gotch, 1979), while amphibius makes reference to living on land and water. Choeropsis refers to "belonging to a pig". "Kiboko" is the Swahili name for the animal. "Hexaprotodon" refers to having 6 front teeth. The plural of hippopotamus is hippopotamuses or hippopotami; the former is employed here.

Numerous skeletal remains of extinct species indicate that hippos were widely distributed over Africa and were also present in Cyprus, Madagascar and reached into Palestine. Since hippos depend on river and water systems, the desertification of Northern Africa has widely eliminated these species, and human intervention has led to the elimination from Palestine, and Cyprus. These finds have suggested to some authors that doubt exists for the need to specify two genera for Hippopotamidae, an aspect that finds further support from their essentially identical karyotypes. A comprehensive review of hippos may be found in Eltringham (1999) and, for the Pygmy Hippopotamus, by Lang (1975). Various authors have presented the action plans to assure survival of these species into the future and the volume also gives the current distribution of both species in Africa (Oliver, ed., 1993).

A number of genetic studies have recently been performed in attempts to delineate the relationships of various artiodactyl taxa to cetacean and other mammalian forms. Thus, a comprehensive analysis by Nikaido et al. (1999) affirms the close relationship from their study of SINE in certain certiodactyls. Ursing & Arnason (1998), studying the mitochondrial genomes, had found support for a hippopotamus-cetacean clade, with a 54 MY separation. Hasegawa & Adachi (1996) were critical of this analogy, and Nomura & Yasue (1999) further pursued the taxonomic relations. Kleineidam et al. (1999) studied pancreatic ribonuclease genes and also supported the relationship of hippopotamidae to odontocetes. In a new study of the mitochondrial genome of various presumably related taxa, Arnason et al. (2000) again confirmed the cetacean-hippo clade and further examined the split of odontocetes and mysticetes. Madsen et al. (2002) most recently examined the genetics of adrenergic receptor molecules and found nesting of whales as a sister group of hippos.

Hippopotamus amphibius:

The large hippopotamus now is confined to the large river systems of Africa. The animal is sometimes referred to as "Nile or River Hippo" and weighs between 1,000 and 4,500 kg (Nowak, 1999). Gestation lasts 227-240 days and results in a single birth of 25-55 kg. The maximal lifespan is given by Jones (in Nowak, 1999) as being 61 years. The gregarious animals are very aggressive when disturbed during foraging and child rearing; frequent deaths have been described (Pickles, 1987; Eltringham, 1999). They are maintained in many zoological parks. The usage of subspecies is not usually practiced any longer in designating origin of members of this species.

Pygmy hippopotamus:

This more terrestrial Western African species (mainly Liberia) is now rare and in need of protection (Robinson, 1971). The subspecies Hexaprotodon liberiensis heslopi has presumably become extinct (Antonius, 2003). It is a solitary animal that is also frequently found in Zoos. It is long-lived; Jones (1982) found a maximal lifespan of 43 years and 10 months. Adults weigh 150-270 kg and neonates are 3-6 kg. The length of gestation is 184-204 days according to Nowak (1999), 200 days according to Lang (1975). The latter author compiled the most extensive observations on the captive management of pygmy hippopotamuses while director at Basel Zoo and observed one set of stillborn twins (male/female) and he knew of one other set of twins. He stated that animals with birth weights under 5 kg need special attention.

2) General Gestational Data

The Nile hippopotamus has singleton births after a gestation period of 227-240 days. One placenta weighed 4,010 g. The pygmy hippopotamus also has singleton births although Lang (1975) observed nonviable twins twice. Gestation lasts 200 days and the placentas obtained by me weighed 600, 950 and 950 g.

3) Implantation

Really early stages of placentation have not been described. The only young specimen studied in situ is that by Amoroso et al. (1958) of a gestation estimated to be 60 days old. They found bare spots of membranes at the peripheral tips and over the endocervical os. Moreover, the antimesometrial villous covers were less profuse. That immature fetus was contained in the left uterine horn, on which side the corpus luteum also was found.

4) General Characterization of the Placenta

Two placentas of a Nile hippopotamus and four placentas from Pygmy hippopotamuses were available to me. Because of their similarity, both species will be presented in this chapter rather than separating them.

Nile hippopotamus placenta:

The first placenta came from a gestation in which the father had killed the neonate. The neonate weighed 32,500 g. The placenta was a huge sac, covered diffusely with villi on its outside. It measured 104 cm in length and 25 cm in width. It was 0.3 cm in thickness and had some non-villous regions at each end. The umbilical cord inserted in the middle and measured 109 cm in length. It contained only three large blood vessels and an allantoic duct. The placenta weighed 4,010 g. The fetal surface was covered with a delicate amnion on which there were numerous plaques of squamous metaplasia (pustules) that also extended over the entire length of the umbilical cord.

The second placenta also comes from a term pregnancy in which the fetus survived. It was accidentally born on land and, therefore, a good placenta was available. It weighed 4,800 g, measured 130 cm in length and had a four-vessel cord attached in the middle that was 31 cm long. From its torn fetal end, however, extruded 51 cm torn vessels, much as they were already depicted in the excellent description by Teuscher (1937).

	Freshly delivered placenta of Nile river hippopotamus. Note the isolated blood vessels (horizontal) that emanate from the cord.
	Numerous foci of squamous metaplasia are found on the cord's surface and extend onto the amnion.
	Maternal surface with diffuse vili extending 1-2 mm.

Pygmy hippopotamus placenta:

The fetal membranes of a pygmy hippopotamus have previously been described by Teuscher (1937). I had four placentas available for study, three from term, surviving neonates and one from a neonatal death at San Diego Zoo. The male neonate that died weighed 5,430 g. Its placenta was 600 g; another placenta weighed 950 g, a fourth weighed 1,125 kg. It was 80 cm long and 0.3 cm in thickness. Its cord inserted centrally and was 100 cm long. The surface was uniformly studded with villi. The amnion had numerous areas of prominent squamous metaplasia.

5) Details of fetal/maternal barrier

This is an epitheliochorial placenta with diffuse villous coverage over nearly the entire chorion; only the lateral tips are free of villi. The villi are short and only some of them are slightly branched. They are covered by cuboidal trophoblast which contains no binucleated cells, as seen in other artiodactyla. The subchorionic trophoblast between the villi is rather more cylindrical and has a paler, empty-appearing cytoplasm. Macdonald & Bosma (1985) described PAS-positive granules in their cytoplasm. Syncytial cells are not present. Both species have yellow-brown pigment accumulations in some trophoblastic cells, especially beneath the chorionic plate. While this pigment is more frequently present between the villi in the subchorionic trophoblast, it is worth noting that it was also found in trophoblast of some of the villous tips. This is distinctly different from the pigmented cells in the placentas of most other artiodactyla. Moreover, it was not associated with any degenerative changes or areas of hemorrhage. It is also iron-stain negative, as it is in other ungulates. The second Nile hippo placenta had absolutely no yellow pigment in the trophoblast. The villi contain no obvious Hofbauer cells, and the thick, muscular nature of the chorionic vessels is remarkable.

Two types of villi are distinguished since their first descriptions by Eschricht (1837) and von Baer (1837). They were further delineated by Turner (1876), Ludwig (1968) and, again, by Macdonald & Bosma (1985). They are the "areolar" villi that lie over the mouths of endometrial glands, and the inter-areolar villi. The latter have more prominent interdigitations of the fetal capillaries with the trophoblast than the former. The former are less branched and broader, and Macdonald & Bosma described "plumes" originating from some cells. Ludwig (1968) made a special study of these villous differences in a variety of mammalian placentas and referred to the former type as being "nephropneumoid", the other being the "enteroid" villi. Turner (1876) had described in some detail the surface structure of villi and endometrium, thus delineating the areolae as regions of endometrial secretion that would subservice different exchange functions than the more abundant nephropneumoid areas.

One remarkable feature is the intense interdigitation of the cuboidal trophoblast with villous capillaries. This aspect has been specially considered by Macdonald & Bosma (1985) who reviewed the literature and added observations on two pygmy and one Nile hippopotamus placentas. Two of these were of immature specimens. They make special reference to the different appearance of some villi - those overlying the areolae (basal capillaries) and those subserving a respiratory exchange (interdigitation of capillaries with trophoblast). The latter produce the "epithelial plates", regions of maximal thinness and, therefore, deemed to be used for gas exchange.

6) Umbilical cord

The umbilical cord of the hippopotamus has excited many earlier observers because of their pronounced, large areas of squamous metaplasia, most beautifully depicted by Hediger (1962). They are already present as foci of squamous epithelium in very young specimens (Keibel, 1893). That author described two young embryonic specimens, observed the areas of squamous metaplasia, found the cord to be already quite long and lightly left spiraled, observed the allantoic duct and remnants of the omphalomesenteric duct. He found two arteries and two veins, aside from smaller vessels. The veins joined immediately after entering the abdomen to form the single umbilical vein. Amoroso et al. (1958), in their comprehensive description of placenta and cord of an immature Nile hippopotamus placenta also observed 4 blood vessels. It is thus surprising, that the umbilical cords of one of the one specimens of Nile hippopotamus available to me and two of the pygmy hippopotamuses all had only three blood vessels, in addition to the small number of small blood vessels as are found in other artiodactyla. They were complete cross sections of the midportions of the cords. This contrasts with the earlier statements above and those of Macdonald and Bosma (1985) who also described four vessels in both species of hippopotamus; they contrasted this to the three vessels found in pigs and peccaries. One of the three pygmy hippopotamuses available to me had four blood vessels. It is the neonate with funisitis (see below). The other striking difference of hippos from suids, of course, is their large number of "pustules" (squamous metaplasia) on cord and amnion. Macdonald & Bosma (1985) referred to them as "pustule-encrusted" cords. The pictures of these cords shown by Hediger (1962) are impressive. It is probable that one blood vessel had already branched into the allantoic sac in those cases where only three vessels were found. This is not too surprising as the large vessels enter the allantoic sac and split off the Wharton's jelly quite early. Thus, the isolated long vessels sticking out from the end of the cord shown by Teuscher (1937) and in the Nile hippo placenta shown above apparently lie more or less free until they find the allantoic sac surface. The large areas of squamous metaplasia on the cord that extend also for a short distance over the amnion are surprising because they often contain clear fluid. Teuscher (1937) even found some bone in one such cyst.

All observers also described the presence of numerous small blood vessels, and Amoroso et al. (1958) quoted Wislocki as having suggested that these contribute to the composition of amnionic fluid. It is true, however, that there is no resolution as to nature and raison d'être for their existence. While some accompany the allantoic duct, this is only the minority and it will be seen in the section of the Nile hippo cord below that there are relatively few blood vessels only in the cord, and that is equally true of the allantoic sac.

The umbilical cord of hippos is quite long; thus, one of the pygmy hippopotamuses had a 100 cm long cord and, grossly as well as histologically, it had only three blood vessels. Another complete cord I observed also had only three vessels and the allantoic duct with hippomanes, while a third had the normal complement of four large vessels.

7) Uteroplacental circulation

This has not been studied except for the cursory description of the one pregnant uterus studied by Amoroso et al. (1958).

8) Extraplacental membranes

The amnions of both species are thin and characterized by a very flat epithelium that has a large number of "pearls" consisting of squamous metaplasia with much keratin production. These extend also over the entire umbilical cord. Brown hippomanes with birefringent crystalline content are found in both species, in the allantoic duct as well as in the allantoic sac. Keibel (1893) stated that the crystals were composed of ammonium-magnesium phosphate. The hippomanes were the special topic of a contribution by Hediger (1962) from his experience at Basel Zoo. This is an interesting paper, but it confuses the squamous metaplasia of cord and amnion with hippomanes, allantoic debris that is a concentrate of urinary substances. I found no hippomanes in the second specimen of Nile hippo.

There is a large allantoic sac that partially separates amnion from chorion. It has cuboidal epithelium that was mostly eroded in my specimen. Remarkably, there are very sparse vessels in the allantoic membrane. Amoroso et al. (1958) described it as being bilobed and entered centrally by the allantoic duct from the umbilical cord. It contained hippomanes even in the young specimens observed by earlier observers. The lining is a flat squamoid epithelium. Near term, the allantoic sac is smaller and gives way to a larger amnion.

9) Trophoblast external to barrier

In the absence of an implanted placenta for study, it can only be conjectured that, analogous to other artiodactyla and cetacea, trophoblastic invasion into endometrium does not occur.

10) Endometrium

Again, in the absence of an implanted placenta, the endometrium and possible decidualization cannot be described. Obviously, there is no decidua capsularis as the entire sac is covered by villi. The endometrium underlying the villi was intact and covered by cuboidal epithelium in the specimen described by Amoroso et al. (1958). They also reported on a significant infiltration by lymphocytes and plasma cells of the endometrium.

The neonatal endometrium is not stimulated and has prominent ridges, but no endometrial glands.

11) Various features

Both species have a peculiar skin secretion that has often been referred to as "blood sweat" because it appears sometimes similar to blood when the light shines appropriately (Nowak , 1999). It is a clear, mucoid secretion that makes the skin slippery and only appears red on occasion. We have attempted to identify specific pigments (the fluid turns dark after standing), without success. It is conjectured that this "sweat" is produced to allow the skin to remain moist, thus avoiding cracking of the epidermis when the animal is out of the water. In sections taken from injuries, rare, deeply-seated glands were found in a Nile river hippopotamus of one year age. It also contained a thin duct. In a 39 y.o. pygmy hippopotamus one hair shaft were found. The deeply-seated glands have minute PAS-positive granules in occasional cells and more in the ducts, whose content is also PAS-positive. They are mucin-negative and did not stain with BRST and SMA (smooth muscle antigen) antibodies either. There were additional sebaceous glands surrounding the hair shafts of the pygmy hippopotamus. None of the contents were discolored as "blood sweat" is. Photographs of these findings are shown next. Oliver (1975) has written about these gland and referred to earlier literature. They were shown to produce hyperosmolar, alkaline secretion whose role "remains obscure". He observed that the physiology and structure of these glands is essentially similar in both species. Lochte (1951) described in great detail the structure of hippopotamus hair. The tusks of both species grow continuously and have been used as "ivory". Nursing of neonates takes place under water. Ears are moved independently, and defecation is dispersed by intense movement of the tail. The nature of the pigment in “blood sweat” has now been elucidated by Saikawa et al. (2004) in a detailed chemical analysis. They identified an orange and red pigment. The labile red pigment being hipposiduric acid (so named by the authors), the orange pigment is norhipposudric acid, three-phenolic rings structures. The red pigment has some antibiotic property and the possibility of the pigments acting as sunscreens was proposed by the authors. They are quite labile and are under further study by the authors.

	Left hair shaft & gland of 39 y.o. Pygmy hippopotamus Right gland a duct of 1 y.o. Nile river hippopotamus.
	PAS stained gland and duct of Nile river hippopotamus.
	Sebaceous-mixed glands adjacent to the hair shaft of a 39 y.o. male pygmy hippopotamus.

13) Genetics

The Nile hippopotamus and the pygmy hippopotamus, both have 36 chromosomes, all metacentrics (Gerneke, 1965; Hsu & Benirschke, 1977). Only the Y-chromosome is acrocentric and it is larger in the pygmy hippopotamus. The two karyotypes are otherwise essentially similar. Hybrids are unknown.

14) Immunology

I am not aware that any studies have been done of the hippopotamus immune system other than those described in the paper by Tsuji et al. (1984). These authors studied the molecular weights of transferrins by rabbit antibodies in a variety of artiodactyl mammals.

15) Pathological features

Bush et al. (1972) described uterine prolapse in a pygmy hippopotamus. Schistosoma edwardiensis was found by Pitchford & Visser (1981) at Kruger National Park in the large hippopotamus, and they discussed the synonymy of different schistosoma species. Dauth et al. (1988) studied blood levels in river hippopotamuses of the Kruger National Park and found them to be lower to that of the population. Copper concentrations (and other trace elements) in liver and kidney were studied by Mwase et al. (2002) at Kafue National Park. No elevation was found, irrespective of the proximity to the "copperbelt". Fatal encephalomyocarditis virus infection of a pygmy hippopotamus (and other species) was described by Reddacliff et al. (1997) from Australia. Kuttin et al. (1982) described "coccidiosis of the placenta" in a Nile hippopotamus, but the paper was not available to me. Griner (1983) examined only neonates of Nile hippopotamuses that had drowned because of management errors. Two neonatal pygmy hippopotamuses (one stillborn) had unknown causes of death. A variety of parasites and diseases of hippopotamuses is discussed by Eltringham (1999

One of our pygmy hippopotamus specimens had marked funisitis (inflammation of arteries and veins) of the umbilical cord, in the absence of chorioamnionitis, as would be usually seen in human specimens. Organisms were not seen histologically, but the neonate had intrauterine aspiration pneumonia. A similar situation has been observed in ascending infection of the horse.

16) Physiologic data

Thermoregulation of the hippopotamus was studied by Cena (1964). Macdonald & Hartman (1983) dissected the stomachs of adult and neonatal pygmy hippos, finding them to be four-chambered. Griner (1983) specified only three chambers. Nevertheless, the animals do not ruminate. The renal anatomy of adult and neonatal pygmy hippopotamus was described in great detail by Maluf (1994). Rouille et al. (1988) studied the occurrence of vasopressin in river hippopotamus and in collared peccary. They found arginine vasopressin but no lysine vasopressine in these species, contrary to the finding in Suidae. In a molecular study of milk casein, Gatesy et al. (1996) found the structure to be closer to cetacea than to other artiodactyl species, confirming the close relationship of whales and hippos. From these data, the authors suggested that an intermediate fossil may so far be "missing". The means of immobilization of the Nile hippopotamus (9 animals) was described by Ramsay et al. (1998). One death occurred from apnea. Flach et al. (1998) removed a dead fetus from a pygmy hippo by Cesarean section with recovery of the dam. Using specific antibodies to retinal cone visual pigments, Peichl et al. (2001) demonstrated the absence of S-cones in odontocetes and five species of seals, but affirmed their presence in pygmy hippopotamus (and other terrestrial animals). This indicated to the authors an absence of color vision in marine mammals. Eltringham (1999) has most comprehensively detailed the anatomy, known physiology, distribution, and general problems affecting hippopotamuses.

17) Other resources

Cell strains of fibroblast are available of both species from CRES at the San Diego Zoo by contacting Dr. Oliver Ryder at oryder@ucsd.edu.

18) Other remarks - What additional Information is needed?

The discrepancy of the number of umbilical cord blood vessels is troubling and needs further study. Also, early specimens, preferably implanted placentas, should be described. There is too little endocrine information and the absence of binucleated trophoblastic cells is of interest. Thus, is there any placental lactogen produced during pregnancy, as in other ungulates? Ascending infection, as seen in equines, needs to be paid attention to.

Acknowledgement

The animal photographs in this chapter come from the Zoological Society of San Diego.

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