The aquatic stage of biphasic amphibians typically represents a phase of rapid growth and development ^[1][50]. For many amphibians the aquatic life cycle starts when females deposit gelatinous eggs in the water, which are externally fertilized by a male during amplexus. After a period of development, the embryos hatch and enter the larval stage ^[2][51]. The larval stage is mainly free-living and non-reproductive, and goes through metamorphosis to reach the terrestrial stage ^[3][52]. The evolutionary persistence of the biphasic life cycle and free-living larval stage suggest a strong adaptive significance to resource acquisition and growth during this phase ^[3][52]. The life histories of biphasic amphibians generally yield high numbers of progeny with very little parental care, and only a small percentage of individuals from the larval stage make it through metamorphosis ^[4][5][6][26,27,53]. As a result, human-induced pressures that further reduce the survival of the amphibian aquatic stage can have compounding catastrophic results on the viability of a population ^[7][8][9][54,55,56]. Threats include reduced hydroperiods and increased temperatures caused by climate change ^{[7][10][11][12][13][14]}[54,57,58,59,60,61], predation or competition by invasive species ^[15][16][17][62,63,64], disease ^[18][39], and pollution ^[19][20][65,66]. Considering the natural low survival rates in the aquatic stage ^[4][5][6][26,27,53] and the emergence of compounding anthropogenic threats, the occurrence of failed recruitment has real implications for the continued persistence of wild populations ^[7][20][54,66].

The embryos and larvae of amphibians are confined to the aquatic habitat they are deposited into, and while many species, such as Bufo gargarizans ^[15][62], demonstrate evolutionary adaptation to speed-up development to escape sub-optimal environments ^[21][22][23][67,68,69], others, such as the natterjack toad (Bufo calamita), do not ^[24][25][26][70,71,72]. Extreme temperature changes influenced by a shifting climate can create unfavourable conditions within ponds, leading to a reduced hydroperiod and an altered pond water chemistry ^[27][28][73,74]. An increased frequency of drought and extended above-average temperatures can affect the survival of embryos and larval development, but this is species-dependent as a result of thermo-tolerance thresholds and certain life history characteristics specific to certain habitats ^{[29][30][31][32]}[75,76,77,78]. The plasticity of the developmental period of larval amphibians in response to altered hydroperiods and water chemistry plays an important part in the long-term viability of amphibian populations ^[14][61].

A reduced hydroperiod is a key driver of recruitment failure in pond-breeding amphibians ^[33][34][79,80]. Studies show that the timing and length of the hydroperiod in ephemeral ponds can have an impact on the reproductive success of many amphibians ^[35][36][37][81,82,83]. When the hydroperiod in ephemeral ponds is shorter than the required development time of larvae, then reproductive success is not reached, as the larvae desiccate as the waterbody, dries ^[35][81]. As the hydroperiod of more ephemeral ponds throughout the landscape reduces, the remaining permanent ponds and their connectivity becomes increasingly important to amphibian reproduction ^[38][39][84,85]. However, permanent ponds are often characterised by different water chemistry, competition, predation, and resource profiles compared to ephemeral ponds ^{[40][41][42][43]}[86,87,88,89].

An increased water temperature can cause mass-mortality events at the larval and embryo stages of amphibian life cycles ^[44][90]. Amphibians exposed to increased temperature profiles, consistent with current and predicted climate trends during early larval development, may have an increased rate of mortality ^[45][91], with one study demonstrating a 100% mortality rate of the common hourglass tree frog (Polypedates cruciger) larvae at temperatures around 34 °C, and death at metamorphosis in larvae kept at 32 °C ^[29][75]. Although some amphibian species demonstrate plasticity in their thermal tolerance, enabling them to cope with extreme temperatures, thermal tolerance and acclimation capacity vary with life stage ^[14][61]. In newly emerged larvae, acclimation capacity is low and the risk of ongoing negative effects of temperature change is high ^[23][69]. Extreme temperatures can also cause sublethal negative effects at the aquatic stage by disrupting the time to and size at metamorphosis ^{[23][29][44][46]}[69,75,90,92]. The rate of development and body size at metamorphosis are vital components of amphibian fitness ^[44][90] and are species-specific. Manasee et al. ^[29][75] found that elevated temperatures delayed tadpole development time and reduced body growth in the common hourglass tree frog; however, in the Asiatic toad (Bufo gargarizans), warmer temperatures resulted in a shorter larval period, and a reduced body size and hind limb length ^[46][92]. Thus, the thermal landscape influences the plastic developmental traits of many ephemeral pond-breeding amphibians, and therefore shapes the growth and development ^[23][69].

Rising temperatures also negatively affect other environmental conditions, such as the decreasing dissolved oxygen levels in ponds ^[47][93], which can cause the added stress of hypoxia at the larval stage ^[48][94]. Hypoxia has been shown to cause serious abnormalities in the central nervous system of bullfrog (Lithobates catesbeianus) larvae, and reduce body mass and length in exposed individuals ^[49][95]. Extreme temperatures also increase bacterial blooms in ponds, giving rise to deleterious pathogens, such as heterotrophic bacteria (Cyanobacterial lipopolysaccharide) and cyanobacterial toxins (microcystins) that can affect embryo masses and cause significant liver and intestinal toxicity in larvae ^[44][50][90,96]. Such changes in temperature can also compromise immunity at the larvae stage, leading to increased susceptibility to infections ^[51][52][97,98]. The combination of these potential impacts makes climate change a significant threat to the survival of larval amphibians.

Invasive species cause substantial environmental damage through both direct and indirect impacts on species and populations ^[53][54][99,100]. At the aquatic stage in the amphibian life cycle, impacts can be direct through predation or competition, as well as indirect through the modification of habitat or alteration of larval behaviour ^{[15][55][56][57]}[2,62,101,102]. Invasive plants reduce the quality of amphibian aquatic habitat by altering the physical structure of aquatic vegetation. This shift in vegetation can directly and indirectly affect the aquatic stage in amphibians by disrupting food webs, changing the chemical composition of pond water, and impacting on egg deposition and clutch structure ^[15][57][58][62,102,103]. Brown et al. ^[15][62] found that the invasive plant, Lythrum salicaria, impacted the larvae of the American toad (Bufo americanus) by direct toxicity of leached tannins. Indirect negative impacts on food webs were also observed through a tadpole gut analysis, which found reduced algal communities in ponds that supported invasives compared to non-invasive plant communities ^[15][62]. Similarly, Pinero-Rodríguez et al. ^[58][103] found that the invasive floating plant (Azolla filiculoides) altered the chemical and physical structure of Mediterranean temporary ponds by forming a dense mat over the water surface, which decreased the pH and oxygen concentration, and increased nutrients, nitrogen, and phosphorus compounds, negatively influencing tadpole survival rates in the slow-developing western spadefoot toad (Pelobates cultripes).

The effects of invasive predators on amphibian populations are well documented ^[16][57][59][63,102,104]. In the presence of the predatory invasive fish, bluegill (Lepomis macrochirus) and largemouth bass (Micropterus salmoides), Rowe and Garcia ^[57][102] found a strong negative relationship with native amphibian counts, suggesting direct predation across the embryo and larval stage. Studies by Hamer ^[60][105] and Klop-Toker et al. ^[61][106] additionally found a negative relationship between the invasive mosquitofish (Gambusia holbrooki) and the reproduction probability of seven different frog species, including the endangered green and gold bell frog (Litoria aurea). Many species have defensive traits to help protect them from predators, such as increased tail fin depth or chemical recognition of predators. However, some species respond differently depending on whether they are exposed to a native predator or invasive predator that they do not have an evolutionary history with ^[59][104]. For the Iberian green frog, Pelophylax perezi, tadpoles detect chemical cues from native predators (dragonfly nymphs), but not invasive red swamp crayfish (Procambarus clarkia), demonstrating a lack of evolved predator perception to the crayfish.

Invasive anurans also negatively impact larval development rates and survivorship through exploitative competition ^[16][17][63,64]. Bullfrogs decreased the size of R. boylii metamorphs through resource competition during the larvae stage ^[17][64], and the presence of bullfrog larvae indirectly impacted the native red-legged frog (Rana aurora) by reducing activity levels and increasing refuge-seeking behaviour ^[16][63]. Kupferberg ^[17][64] found that predation by introduced bullfrogs (Rana catesbeiana) reduced the abundance of native yellow-legged frog (Rana boylii) larvae.

The aquatic habitat of embryos and larvae also harbours disease-causing pathogens. Most notably, the fungal pathogen Batrachochytrium dendrobatidis (Bd), which causes the disease chytridiomycosis, was first linked to amphibian population declines in 1992 ^[62][63][107,108]. Chytridiomycosis is now considered responsible for the decline in hundreds of frog species around the world ^[64][65][109,110]. Bd occurs in the water and moist soil of temperate freshwater environments, and larvae become infected with its motile zoospores that penetrate the keratinised mouthparts (tooth rows and jaws) ^[66][111]. Because the area of infection is lower in larvae than the later terrestrial stages where the pathogen infects the skin ^[66][67][111,112], it is rarely lethal in larvae; however, it has been associated with mouthpart loss ^[68][113], and decreased activity and reduced foraging performance, causing nutrient disruptions which impact the growth and development rates ^[69][114]. Parris and Baud ^[70][115] found that exposure to Bd significantly reduced the growth and development of larvae; however, no effect was observed on survival. When larvae go through metamorphosis and start depositing keratin in other areas such as the epidermis, Bd can spread, impacting the infected metamorphosing individuals ^[18][71][39,116]. Metamorphosis has been identified as a highly vulnerable stage in the life cycle as immune function is reduced during this period of extreme physiological change ^[72][73][117,118]. As such, juveniles with immature immune systems may have a compounded susceptibility to Bd and demonstrate higher mortality rates following exposure at this stage ^[73][74][75][118,119,120].

Altered water chemistry, caused by mining, industrial, and agriculture practices, can negatively impact the aquatic stages of amphibians. Changes in pH levels and the release of coal combustion and heavy metals, such as iron, manganese, and copper, into aquatic environments have been shown to cause acute negative impacts on amphibian larvae and embryos ^{[19][20][76][77]}[65,66,121,122]. Salice et al. ^[20][66] examined the population-level impacts of aquatic coal combustion residue (CCR) on the different life stages of the eastern narrow-mouth toad, (Gastrophryne carolinensis). Population models indicated that toads exposed to CCRs were more susceptible to decline and extinction compared to non-exposed toad populations ^[20][66]. Acidic rain and emissions of sulphur dioxide caused by industrial pollution have been found to cause impacts on the development and survival of amphibian larvae ^{[76][78][79][80]}[121,123,124,125]. Farquharson et al. ^[76][121] found that prolonged exposure to decreased pH levels (increased acidity) resulted in decreased tadpole size and increased tadpole deformities. Increasing acidity was also found to delay metamorphosis in tadpoles. Mining practices and the run-off from tailing dams can release high levels of heavy metals, such as iron, into waterways ^[81][82][126,127], which can result in larval fatalities ^[77][83][122,128]. In agricultural fields, pesticides and fertilisers accumulate, and can become a source of contamination for nearby environments ^[84][129]. These chemicals are washed into rivers, lakes, and other waterways from the land ^[85][130]. Pesticides, such as glyphosate and atrazine, can cause acute and chronic effects on amphibians, including developmental effects and disruptions of the nervous system ^[86][87][131,132]. Due to the many potential anthropogenic sources of water pollution, this threat has the potential to impact many species that persist in non-protected areas.