The World Wide Fund for Nature (WWF) subdivides the realm into bioregions, defined as "geographic clusters of ecoregions that may span several habitat types, but have strong biogeographic affinities, particularly at taxonomic levels higher than the species level (genus, family)."

Laurel forest and other cloud forest are subtropical and mild temperate forest, found in areas with high humidity and relatively stable and mild temperatures. Tropical rainforest, tropical and subtropical moist broadleaf forests are highlight in Southern North America, Amazonia, Caribbean, Central America, Northern Andes and Central Andes.
The Amazonia bioregion is mostly covered by tropical moist broadleaf forest, including the vast Amazon rainforest, which stretches from the Andes Mountains to the Atlantic Ocean, and the lowland forests of the Guianas. The bioregion also includes tropical savanna and tropical dry forest ecoregions.
The Central Andes lie between the gulfs of Guayaquil and Penas and thus encompass southern Ecuador, Chile, Peru, western Bolivia, and northwest and western Argentina.
Eastern South America includes the Caatinga xeric shrublands of northeastern Brazil, the broad Cerrado grasslands and savannas of the Brazilian Plateau, and the Pantanal and Chaco grasslands. The diverse Atlantic forests of eastern Brazil are separated from the forests of Amazonia by the Caatinga and Cerrado, and are home to a distinct flora and fauna.
North of the Gulf of Guayaquil in Ecuador and Colombia, a series of accreted oceanic terranes (discrete allochthonous fragments) have developed that constitute the Baudo, or Coastal, Mountains and the Cordillera Occidental.
The Orinoco is a region of humid forested broadleaf forest and wetland primarily comprising the drainage basin for the Orinoco River and other adjacent lowland forested areas. This region includes most of Venezuela and parts of Colombia, as well as Trinidad and Tobago.
The temperate forest ecoregions of southwestern South America, including the temperate rain forests of the Valdivian temperate rain forests and Magellanic subpolar forests ecoregions, and the Juan Fernández Islands and Desventuradas Islands, are a refuge for the ancient Antarctic flora, which includes trees like the southern beech (Nothofagus), podocarps, the alerce (Fitzroya cupressoides), and Araucaria pines like the monkey-puzzle tree (Araucaria araucana). These rainforests are endangered by extensive logging and their replacement by fast-growing non-native pines and eucalyptus.

South America was originally part of the supercontinent of Gondwana, which included Africa, Australia, India, New Zealand, and Antarctica, and the Neotropic shares many plant and animal lineages with these other continents, including marsupial mammals and the Antarctic flora.

After the final breakup of the Gondwana about 110 million years ago, South America was separated from Africa and drifted north and west. 66 million years ago, the Cretaceous–Paleogene extinction event altered local flora and fauna. Much later, about two to three million years ago, South America was joined with North America by the formation of the Isthmus of Panama, which allowed a biotic exchange between the two continents, the Great American Interchange. South American species like the ancestors of the Virginia opossum (Didelphis virginiana) and the armadillo moved into North America, and North Americans like the ancestors of South America's camelids, including the llama (Lama glama), moved south. The long-term effect of the exchange was the extinction of many South American species, mostly by outcompetition by northern species.

The Neotropical realm has 31 endemic bird families, which is over twice the number of any other realm. They include tanagers, rheas, tinamous, curassows, antbirds, ovenbirds, toucans, and seriemas. Bird families originally unique to the Neotropics include hummingbirds (family Trochilidae) and wrens (family Troglodytidae).

Mammal groups originally unique to the Neotropics include:
• Order Xenarthra: anteaters, sloths, and armadillos
• New World monkeys
• Solenodontidae, the solenodons
• Caviomorpha rodents, including capybaras, guinea pigs, hutias, and chinchillas
• American opossums (order Didelphimorphia) and shrew opossums (order Paucituberculata)
The Neotropical realm has 63 endemic fish families and subfamilies, which is more than any other realm. Neotropical fishes include more than 5,700 species, and represent at least 66 distinct lineages in continental freshwaters (Albert and Reis, 2011). The well-known red-bellied piranha is endemic to the Neotropic realm, occupying a larger geographic area than any other piranha species. Some fish groups originally unique to the Neotropics include:
• Order Gymnotiformes: Neotropical electric fishes
• Family Characidae: tetras and allies
• Family Loricariidae: armoured catfishes
• Subfamily Cichlinae: Neotropical cichlids
• Subfamily Poeciliinae: guppies and relatives
Examples of other animal groups that are entirely or mainly restricted to the Neotropical region include:
• Caimans
• New World coral snakes
• Poison dart frogs
• Dactyloidae ("anoles")
• Rock iguanas (Cyclura)
• Preponini and Anaeini butterflies (including Agrias)
• Brassolini and Morphini butterflies (including Caligo and Morpho)
• Callicorini butterflies
• Heliconiini butterflies
• Ithomiini butterflies
• Riodininae butterflies
• Eumaeini butterflies
• Firetips or firetail skipper butterflies
• Euglossini bees
• Augochlorini bees

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