Mouthbrooding Females Research Articles

Opsin Expression Varies with Reproductive State in the Cichlid Fish Astatotilapia burtoni.

Animals use visual communication to convey crucial information about their identity, reproductive status, and sex. Plasticity in the auditory and olfactory systems has been well-documented, however, fewer studies have tested for plasticity in the visual system, a surprising detail since courtship and mate choice are largely dependent on visual signals across taxa. We previously found reproductive state-dependent plasticity in the eye of the highly social cichlid fish Astatotilapia burtoni. Male A. burtoni increase their courtship, including multicomponent visual displays, when around ovulated females, and ovulated females are more responsive to male visual courtship displays than non-ovulated females. Based on this, we hypothesized that ovulation status impacts visual capabilities in A. burtoni females. Using electroretinograms, we found that ovulated females had greater visual sensitivity at wavelengths corresponding to male courtship coloration compared with non-reproductively-receptive females. In addition, ovulated females had higher neural activation in the retina and higher mRNA expression levels of neuromodulatory receptors (e.g., sex-steroids; gonadotropins) in the eye than non-ovulated females. Here, we add to this body of work by testing the hypothesis that cone opsin expression changes with female reproductive state. Ovulated females had higher expression of short wavelength sensitive opsins (sws1, sws2a, sws2b) compared with mouthbrooding females. Further, expression of sws2a, the most abundant opsin in the A. burtoni eye, positively correlated with levels of circulating 11-ketotestosterone and estradiol and estrogen, androgen, and gonadotropin system receptor expression in the eye in females. These data indicate that reproductive state-dependent plasticity also occurs at the level of photoreceptors, not just through modulation of visual signals at downstream retinal layers. Collectively, these data provide crucial evidence linking endocrine modulation of visual plasticity to mate choice behaviors in females.

Noise during mouthbrooding impairs maternal care behaviors and juvenile development and alters brain transcriptomes in the African cichlid fish Astatotilapia burtoni.

Anthropogenic noise has increased underwater ambient sound levels in the range in which most fishes detect and produce acoustic signals. Although the impacts of increased background noise on fish development have been studied in a variety of species, there is a paucity of information on how noise affects parental care. Mouthbrooding is an energetically costly form of parental care in which the brooding fish carries developing larvae in the buccal cavity for the duration of development. In the African cichlid Astatotilapia burtoni, females carry their brood for ~2 weeks during which time they do not eat. To test the hypothesis that increased background noise impacts maternal care behaviors and brood development, we exposed brooding females to a 3-h period of excess noise (~140 dB) played through an underwater speaker. Over half of noise-exposed brooding females cannibalized or pre-maturely released their brood, but 90% of control females exhibited normal brooding behaviors. RNA-seq analysis revealed that transcripts related to feeding and parental care were differentially expressed in the brains of noise-exposed females. Juveniles that were exposed to noise during their brood period within the mother's mouth had lower body condition factors, higher mortality and altered head transcriptomes compared with control broods. Furthermore, onset of adult-typical coloration and behaviors was delayed compared with control fish. Together, these data indicate that noise has severe impacts on reproductive fitness in mouthbrooding females. Our results, combined with past studies, indicate that parental care stages are extremely susceptible to noise-induced perturbations with detrimental effects on species persistence.

Distribution of aromatase in the brain of the African cichlid fish Astatotilapia burtoni: Aromatase expression, but not estrogen receptors, varies with female reproductive‐state

Estrogen synthesis and signaling in the brains of vertebrates has pleotropic effects ranging from neurogenesis to modulation of behaviors. The majority of studies on brain-derived estrogens focus on males, but estrogenic signaling in females likely plays important roles in regulation of reproductive cycling and social behaviors. We used females of the mouth brooding African cichlid fish, Astatotilapia burtoni, to test for reproductive state-dependent changes in estrogenic signaling capacity within microdissected brain nuclei that are important for social behaviors. Expression levels of the rate-limiting enzyme aromatase, but not estrogen receptors, measured by qPCR changes across the reproductive cycle. Gravid females that are close to spawning had higher aromatase levels in all brain regions compared to females with lower reproductive potential. This brain aromatase expression was positively correlated with circulating estradiol levels and ovarian readiness. Using chromogenic in situ hybridization we localized aromatase-expressing cells to ependymal regions bordering the ventricles from the forebrain to the hindbrain, and observed more abundant staining in gravid compared to mouth brooding females in most regions. Staining was most prominent in subpallial telencephalic regions, and diencephalic regions of the preoptic area, thalamus, and hypothalamus, but was also observed in sensory and sensorimotor areas of the midbrain and hindbrain. Aromatase expression was observed in radial glial cells, revealed by co-localization with the glial marker GFAP and absence of co-localization with the neuronal marker HuC/D. Collectively these results support the idea that brain-derived estradiol in females may serve important functions in reproductive state-dependent physiological and behavioral processes across vertebrates.

Parental investment matters for maternal and offspring immune defense in the mouthbrooding cichlid Astatotilapia burtoni

BackgroundParental care, while increasing parental fitness through offspring survival, also bears cost to the care-giving parent. Consequentially, trade offs between parental care and other vitally important traits, such as the immune system seem evident. In co-occurring phases of parental care and immunological challenges negative consequences through a resource allocation trade off on both the parental and the offspring conditions can be predicted. While the immune system reflects parental stress conditions, parental immunological investments also boost offspring survival via the transfer of immunological substances (trans-generational immune priming).We investigated this relationship in the mouthbrooding East African cichlid Astotatilapia burtoni. Prior to mating, females were exposed to an immunological activation, while others remained immunologically naïve. Correspondingly, the immunological status of females was either examined directly after reproduction or after mouthbrooding had ceased. Offspring from both groups were exposed to immunological challenges to assess the extent of trans-generational immune priming. As proxy for immune status, cellular immunological activity and gene expression were determined.ResultsBoth reproducing and mouthbrooding females allocate their resources towards reproduction. While upon reproduction the innate immune system was impeded, mouthbrooding females showed an attenuation of inflammatory components. Juveniles from immune challenged mouthbrooding females showed downregulation of immune and life history candidate genes, implying a limitation of trans-generational plasticity when parents experience stress during the costly reproductive phase.ConclusionOur results provide evidence that both parental investment via mouthbrooding and the rise of the immunological activity upon an immune challenge are costly traits. If applied simultaneously, not only mothers seem to be impacted in their performance, but also offspring are impeded in their ability to react upon a potentially virulent pathogen exposure.

Context-dependent chemosensory signaling, aggression, and neural activation patterns in gravid female African cichlid fish

Social animals must constantly assess their environment to make appropriate behavioral decisions. The use of various sensory modalities is imperative in this process and it is hypothesized that the highly conserved brain nuclei comprising the social decision-making network (SDMN) integrates social information with an animal's internal state to elicit behavioral responses. Here, we used the highly social African cichlid fish, Astatotilapia burtoni, to investigate whether reproductively receptive (gravid) females show contextual chemosensory signaling, social behaviors and neural activation patterns within the SDMN. We exposed gravid females to different social contexts: (1) dominant male (inter-sexual reproductive); (2) mouth brooding (non-receptive) female; (3) gravid female (intra-sexual aggressive); (4) juvenile fish (low social salience); and (5) empty compartment (control). By injecting females with a blue dye to visualize urine pulses, we found that gravid females show context-dependent urination, exhibiting higher urination rates in the presence of dominant males (reproductive context) and mouth brooding females (aggressive contexts). Further, gravid females show contextual aggression with increased aggressive displays toward mouth brooding females compared with other gravid females. Using in situ hybridization to quantify cells expressing the immediate early gene cfos as a measure of neural activation, we also show that certain regions of the SDMN in gravid females are differentially activated after exposure to high compared with low social salience contexts. Coupled with previous reports, these results demonstrate true chemosensory communication in both sexes of a single fish species, as well as reveal the neural substrates mediating intra- and inter-sexual social behaviors in females.

Reproductive seasonality of Geophagus steindachneri Eigenmann & Hildebrand, 1922 (Perciformes: Cichlidae) in a tropical mountain river

Reproductive seasonality in tropical freshwater fishes is strongly influenced by rainfall. In lowlands, floods spill laterally to floodplains and fishes usually breed during the flooding season. In mountain rivers, floods are sudden and flush out aquatic organisms. Fishes in mountain rivers usually breed during dry seasons, what has been hypothesized as a strategy to reduce mortality due to strong floods. If that is the case, mouth-brooding fishes should suffer less from strong floods and should have more prolonged breeding seasons in mountain rivers. Here we investigated the breeding activity of a mouth-brooding cichlid (Geophagus steindachneri) in a mountain river in Colombia using three kinds of evidence: monthly variation of gonad weight, macroscopic and histological observations of the gonads, and occurrence of mouth-brooding females. Analysis was made on adults captured monthly throughout a year. The results indicate that G. steindachneri breeds during the dry season in the mountain river studied. Female mouth brooding was related with a halt in the maturation of their ovaries. Other factors than the flushing-out effect of floods on offspring may be determining dry-season breeding of fishes in tropical mountain rivers.

Food deprivation explains effects of mouthbrooding on ovaries and steroid hormones, but not brain neuropeptide and receptor mRNAs, in an African cichlid fish

Feeding behavior and reproduction are coordinately regulated by the brain via neurotransmitters, circulating hormones, and neuropeptides. Reduced feeding allows animals to engage in other behaviors important for fitness, including mating and parental care. Some fishes cease feeding for weeks at a time in order to provide care to their young by brooding them inside the male or female parent's mouth. Maternal mouthbrooding is known to impact circulating hormones and subsequent reproductive cycles, but neither the full effects of food deprivation nor the neural mechanisms are known. Here we ask what effects mouthbrooding has on several physiological processes including gonad and body mass, brain neuropeptide and receptor gene expression, and circulating steroid hormones in a mouthbrooding cichlid species, Astatotilapia burtoni. We ask whether any observed changes can be explained by food deprivation, and show that during mouthbrooding, ovary size and circulating levels of androgens and estrogens match those seen during food deprivation. Levels of gonadotropin-releasing hormone 1 (GnRH1) mRNA in the brain were low in food-deprived females compared to controls and in mouthbrooding females compared to gravid females. Levels of mRNA encoding two peptides involved in regulating feeding, hypocretin and cholecystokinin, were increased in the brains of food-deprived females. Brain mRNA levels of two receptors, GnRH receptor 2 and NPY receptor Y8c, were elevated in mouthbrooding females compared to the fed condition, but NPY receptor Y8b mRNA was differently regulated by mouthbrooding. These results suggest that many, but not all, of the characteristic physiological changes that occur during mouthbrooding are consequences of food deprivation.

Contextual chemosensory urine signaling in an African cichlid fish

Chemosensory signaling is crucial for communication in many fish species, but little is known about how signalers modulate chemical output in response to sensory information and social context. Here, we tested the hypothesis that dominant male African cichlid fish (Astatotilapia burtoni) use urine signals during social interactions, and demonstrate that this signaling depends on social context (reproductive; territorial) and on available sensory information (visual cues; full interaction). We injected males with dye to visualize urine pulses and exposed them to full sensory information or visual cues alone of four types: (1) dominant male; (2) gravid (reproductively receptive) females; (3) mouth-brooding (non-receptive) females; or (4) control (no fish). We found that males released urine sooner and increased their urination frequency when visually exposed to gravid females as compared with mouth-brooding females and or no-fish controls. While males could distinguish female reproductive states using visual cues alone, courtship behavior rates were ∼10-fold higher when they fully interacted with gravid females compared with receiving visual cues alone. Males also increased their urination and territorial behaviors when exposed to another male, suggesting that chemical signals may convey information on dominance status. These data support the hypothesis that dominant males use urine as a chemical signal and adjust the frequency of their urine output based on contextual information.

Invisible pair bonds detected by molecular analyses

A focus on pair bonds between males and females is fundamental to study the evolution of social organization. Because pair bonds are generally identified from direct observations of pairs that maintain physical proximity, pair bonds may have been overlooked in animals that do not exhibit such visible pairs. The Lake Tanganyika cichlid fish Xenotilapia rotundiventralis forms schools that consist of mouthbrooding and non-brooding adults in mid-water, and visible pairs are not recognized. A previous study suggested that mouthbrooding females transfer fractions of the young to males when the young become large. However, it remains a mystery whether the mating pairs maintain pair bonds so that the females can transfer the young to their mates. To answer this question, we conducted a parentage analysis using 10 microsatellite markers. The analysis showed that the mouthbrooding adults were most likely genetic fathers and mothers of the young in their mouths. This finding suggests that the female-to-male shift of young takes place between mating partners, and thus the mating pairs maintain pair bonds at least until the shift of young. The present study is the first to detect pair bonds in animals in which physical proximity has not been observed.

Heritability and adaptive significance of the number of egg-dummies in the cichlid fish Astatotilapia burtoni

Cichlid fishes are a textbook example of rapid speciation and exuberant diversity--this applies especially to haplochromines, a lineage with approximately 1800 species. Haplochromine males uniquely possess oval, bright spots on their anal fin, called 'egg-spots' or 'egg-dummies'. These are presumed to be an evolutionary key innovation that contributed to the tribe's evolutionary success. Egg-spots have been proposed to mimic the ova of the mouthbrooding females of the corresponding species, contribute to fertilization success and even facilitate species recognition. Interestingly, egg-spot number varies extensively not only between species, but also within some populations. This high degree of intraspecific variation may appear to be counterintuitive since selection might be expected to act to stabilize traits that are correlated with fitness measures. We addressed this 'paradox' experimentally, and found that in the haplochromine cichlid Astatotilapia burtoni, the number of egg-spots was related to male age, body condition and dominance status. Intriguingly, the egg-spot number also had a high heritable component (narrow sense heritability of 0.5). These results suggest that the function of egg-spots might have less to do with fertilization success or species recognition, but rather relate to mate choice and/or male-male competition, helping to explain the high variability in this important trait.

The functional significance of buccal feeding in the mouthbrooding cichlid Tropheus moorii

The mouthbrooding cichlid Tropheus moorii exhibits an exceptional degree of maternal investment. Females produce very large eggs, which are incubated for a period of six weeks, and they feed their young in the mouth while starving themselves. Only very few fish species are known to feed their young. We hypothesized that feeding may either (1) benefit females directly if young develop faster, or (2) provide a size advantage to young. To distinguish between these mutually non-exclusive hypotheses, we measured costs and benefits arising for mouthbrooding females when feeding their young. Field observations revealed that mouthbrooding females reduced bite rates, locomotion and territorial defence compared to non-brooding adults. Feeding rates correlated positively with locomotion, as fish moving around more also spent more time with territory defence and other social interactions. This suggests that buccal feeding is costly when compared to mere incubation without feeding the young. In an experiment, in which we controlled the access of females to food, we showed that these costs are apparently not counterbalanced by a benefit to females through a shorter incubation duration. Rather, fed young were larger, heavier and had higher burst-swimming speeds. The extreme maternal investment of T. moorii appears to yield fitness benefits to females by producing larger and stronger young, which consequently should have better survival chances in their natural environment.

Use of Human Chorionic Gonadotrophin (hCG) for Induced Spawning in Tilapia Under Laboratory Conditions

A study was conducted to determine the optimum dosage of Chorulon®, a commercial preparation of human chorionic gonadotrophin (hCG), for increasing spawning rates in the Nile tilapia, Oreochromis niloticus, under laboratory conditions. Dosages of 500-3,500 IU·kg·1 body weight hCG were injected intramuscularly into sexually-mature females reared in fiberglass aquaria. Spermiating males were provided at 1 :1 sex ratio. The number of ovulated and mouthbrooding females were determined 24 h after injection. The eggs from ovulated females were stripped and fertilized artificially while those from mouthbrooding females were removed from the oral cavity of the fish. All eggs were transferred to individual zuger jars for determination of pigmentation rates as an indicator of fertilization rates. Results showed that dosages of 2,000-3,500 tu·kg·1 hCG were effective in significantly increasing the percentage of females either ovulating or mouthbrooding 24 h after injection (25-75%), with pigmentation rates ranging from 36 to 83%. The optimum dosage of the hormone was 3,500 IU·kg

Food intake by mouthbrooding females ofCyphotilapia frontosa (Cichlidae) to feed both themselves and their young

Feeding behaviour of mouthbrooding females ofCyphotilapia frontosa was observed in their natural habitat, and specimens of mouthbrooding females and the young in their mouths were examined in the laboratory. Mouthbrooding females exhibited feeding actions and their guts contained about one quarter as much food as those of nonbrooding adults. A substantial amount of food was found in young 12.5 mm TL who retained a large quantity of yolk, and gut fullness of young increased as they grew. Weight changes of the young suggested that the buccal feeding augmented their growth.

Active browsing by mouthbrooding females of Tropheus duboisi and Tropheus moorii (Cichlidae) to feed the young and/or themselves

Fishes usually do not eat while brooding offspring in their mouths. In two epilithic algal eaters Tropheus duboisi and T. moorii in Lake Tanganyika, however, mouthbrooding females exhibited feeding actions. In T. duboisi, the feeding rate of mouthbrooding females was 80 percent of that of males and non brooding females irrespective of the developmental state of their offspring. In T. moori, females brooding early embryos rarely fed but their feeding rate increased with development of offspring. An examination of specimens revealed that such females took food for nourishment of themselves and the young in the former species but for nourishment of only the young in the latter.

Effects of Frequency of Egg and Fry Removal on Spawning by Tilapia nilotica in Hapas

Abstract Tilapia nilotica were spawned in small-mesh net enclosures called hapas. During a 5-week period, eggs and fry were removed from mouth-brooding females at examination intervals of 2, 4, 7, or 10 d. A decrease in time between examination intervals increased the total number of spawns collected (P < 0.05), but more spawns were obtained per examination for the 7- and 10-d intervals (P < 0.05). For artificial incubation of eggs, the 7-d interval was considered optimal.

Salinity tolerance of Nile tilapia fry ( Oreochromis niloticus), spawned and hatched at various salinities

Fertilized eggs of the Nile tilapia ( Oreochromis niloticus L.) spawned in freshwater, were removed from mouthbrooding females, 1 day post-spawning and artificially incubated at elevated salinities. At 6 days post-hatching, mean survivals of 85.5, 84.4, 82.5, 56.3, 37.9, 20.0 and 0% were recorded for broods incubated at salinities of 0, 5, 10, 15, 20, 25 and 32 ppt, respectively. Fertilized eggs exhibited a 96-h median lethal salinity (MLS-96) of 18.9 ppt, a value identical to that of 7- to 120-day-old fry and fingerlings. Fertilized eggs exhibited a higher median survival time (ST 50 = 978 min) than 7- to 395-day-old fry and fingerlings (ST 50 = 28.8–179.0 min). The salinity tolerances of fry spawned at various salinities and fry spawned in freshwater but hatched at various salinities, were determined using the median survival time (ST 50), mean survival time (MST) and 96 h-median lethal salinity (MLS-96) indices. For comparative purposes, fry spawned and hatched in freshwater were acclimatized to various salinities and their salinity tolerance determined. Fry salinity tolerance progressively increased with increasing salinity of spawning, hatching, or acclimatization. However, at equivalent salinity, early exposure (spawning) produced progeny of comparatively higher salinity tolerance than those spawned in freshwater and hatched at elevated salinity. Similarly, at equivalent salinity, progeny spawned in freshwater but hatched at elevated salinity exhibited higher salinity tolerance than those spawned and hatched in freshwater, then acclimatized to an elevated salinity. The utility of these methods of early salinity exposure toward the saltwater culture of tilapias is discussed.

Brood care motivation and hunger in the mouthbrooding cichlid [formula omitted

The interrelationship of hunger and brood care motivation was investigated in the mouthbrooding cichlid Pseudocrenilabrus multicolor from East Africa. In this species the female takes up her eggs during spawning and carries them continuously in her buccal cavity for approximately 10 days (at 27°C). Females with brood in their mouths do not eat. If the brood is removed from the female's mouth at varied times after spawning, brood care motivation (BM) - defined as the readiness to keep brood in the mouth and not to eat it - is maintained for several days (as in most first spawners) or disappears within a few hours (as in most non-first spawners). Hunger - as measured by the amount of Tubifex ingested per unit time - depends on whether the brood-deprived female exhibits BM or not: females without BM eat as much as non-brooding control females. Females with BM eat only approximately half as much. This is true for first spawners as well as for non-first spawners. It can be concluded that a complete physiological anorexia is not needed to prevent brood-eating in mouthbrooding females. A substantial lowering of the general satiation level is sufficient. In addition, stimuli from the brood in the female's mouth totally inhibit food ingestion from outside the mouth during mouthbrooding.