Marsupial Pouch Research Articles

Bioactive components in the marsupial pouch and milk

Abstract Marsupials give birth to immunologically naïve young after a relatively short gestation period compared to eutherians. Consequently, the joey significantly relies on maternal protection, which is the focus of the present review. The milk and the pouch environment are essential contributors to maternal protection for the healthy development of joeys. In this review, we discuss bioactive components found in the marsupial pouch and milk that form cornerstones of maternal protection. These bioactive components include immune cells, immunoglobulins, the s100 family of calcium-binding proteins, lysozymes, whey proteins, antimicrobial peptides and other immune proteins. Furthermore, we investigated the possibility of the presence of plurifunctional components in milk and pouches that are potentially bioactive. These compounds include caseins, vitamins and minerals, oligosaccharides, lipids, and microRNAs. Where applicable, this review addresses variability in bioactive components during different phases of lactation, designed to fulfil the immunological needs of the growing pouch young. Yet there are numerous additional research opportunities to pursue, including uncovering novel bioactive components, investigating their mode of action, dynamics, stability, and ability to penetrate the gut epithelium ensuring systemic actions.

Dynamical study of three African Easterly Waves in September 2021

AbstractThree convectively active African easterly waves (AEWs) that propagated south of the African easterly jet were observed over the northeast Atlantic Ocean in September 2021. Their evolution is studied using a suite of theoretical frameworks, as well as the European Centre for Medium‐range Weather Forecast reanalyses and satellite‐derived brightness temperature observations. The environment of these AEWs was sampled during the Cloud–Atmospheric Dynamics–Dust Interactions in West Africa campaign near Cape Verde with the goal to assess their potential for developing into tropical cyclones. We highlight the processes that inhibited the development of the first AEW (which evolved into tropical disturbance Pierre‐Henri) and that played a role in the development of the later two into tropical storms Rose and Peter on September 19, 2021. The three AEWs developed a so‐called “marsupial protective” pouch. For Peter and Rose, the pouch was associated with a vertically aligned vortex at low levels and efficiently protected the convective systems inside from dry and dusty air intrusion. The development of this low‐level vortex is associated with an interaction with the monsoon trough for Rose and with a vorticity center associated with a wave propagating north of the African easterly jet (AEJ) in the case of Peter. The presence of a dust flux toward the convective core near the surface is highlighted for Rose and Peter in spite of the presence of the protective marsupial pouch. On the other hand, Pierre‐Henri interacted positively with both the monsoon trough and an AEW north of the AEJ but failed to develop into a tropical cyclone. The wave north of the AEJ brought Saharan air layer air masses inside the pouch that led to a drying of the circulation that may explain the decrease in convective activity.

Tropical cyclogenesis: Controlling factors and physical mechanisms

In this review, advances in the understanding of the controlling factors and physical mechanisms of tropical cyclogenesis (TCG) are summarized from recent (2018–2022) research on TCG, as presented in the Tenth International Workshop on Tropical Cyclones (IWTC-10). Observational, theoretical, and numerical modeling studies published in recent years have advanced our knowledge on the influence of large-scale environmental factors on TCG. Furthermore, studies have shown clearly that appropriate convective coupling with tropical equatorial waves enhances the development chances of TCG. More recently, illuminating research has been carried out on analyzing the mechanisms by which oscillations and teleconnections (El Niño Southern Oscillation (ENSO) in particular) modulate TCG globally, in association with changes in the sea surface temperature (SST). In addition to this, recent research has diligently addressed different aspects of TCG. Multiple studies have reported the applicability of unified theories and physical mechanisms of TCG in different ocean basins. Recently, research has been carried out on TCG under different flow pattern regimes, dry air intrusion, importance of marsupial pouch, genesis of Medicanes, wind shear, convection and vertical structure. Furthermore, studies have discussed the possibility of near equatorial TCG provided that there is enough supply of background vertical vorticity and relatively low vertical wind shear. Progress has been made to understand the role of climate change on global and regional TCG. However, there are still significant gaps which need to be addressed in order to better understand TCG prediction.

Pouch bacteria: an understudied and potentially important facet of marsupial reproduction

Australia is home to a rich biodiversity of marsupials that are found nowhere else. Unfortunately, many of these species are currently threatened with extinction due to introduced feral predators and other anthropogenic factors. There is growing recognition that host-associated microorganisms can play important roles for animal health, with billions of dollars currently being invested into human gut microbiome research and the development of microbiome-based therapeutics to improve human health. Can microorganisms also be harnessed to stem the tide of marsupial extinctions? In this review, we provide an overview of some of the challenges facing Australia’s marsupials, and our current understanding of the microbiology of the marsupial pouch. We also propose outstanding research questions pertaining to the marsupial pouch, which, if addressed, may provide actionable knowledge and novel microbial therapies that could help stem the tide of marsupial extinctions in Australia.

Host reproductive cycle influences the pouch microbiota of wild southern hairy-nosed wombats (Lasiorhinus latifrons)

BackgroundMarsupials are born much earlier than placental mammals, with most crawling from the birth canal to the protective marsupium (pouch) to further their development. However, little is known about the microbiology of the pouch and how it changes throughout a marsupial’s reproductive cycle. Here, using stringent controls, we characterized the microbial composition of multiple body sites from 26 wild Southern Hairy-nosed Wombats (SHNWs), including pouch samples from animals at different reproductive stages.ResultsUsing qPCR of the 16S rRNA gene we detected a microbial community in the SHNW pouch. We observed significant differences in microbial composition and diversity between the body sites tested, as well as between pouch samples from different reproductive stages. The pouches of reproductively active females had drastically lower microbial diversity (mean ASV richness 19 ± 8) compared to reproductively inactive females (mean ASV richness 941 ± 393) and were dominated by gram positive bacteria from the Actinobacteriota phylum (81.7–90.6%), with the dominant families classified as Brevibacteriaceae, Corynebacteriaceae, Microbacteriaceae, and Dietziaceae. Three of the five most abundant sequences identified in reproductively active pouches had closest matches to microbes previously isolated from tammar wallaby pouches.ConclusionsThis study represents the first contamination-controlled investigation into the marsupial pouch microbiota, and sets a rigorous framework for future pouch microbiota studies. Our results indicate that SHNW pouches contain communities of microorganisms that are substantially altered by the host reproductive cycle. We recommend further investigation into the roles that pouch microorganisms may play in marsupial reproductive health and joey survival.

Marsupial brood care in Cretaceous tanaidaceans

Parental care in animal evolution has long fascinated biologists, but tracing this complex of behavioural repertoires is challenging, as these transitory states often leave no corporeal traces as fossils. Among modern invertebrates, the tanaidaceans (Malacostraca: Peracarida), a lineage of marsupial crustaceans, show an interesting variety of brooding strategies. Here we report on fossil tanaidaceans from the Cretaceous of Spain and France that provide conclusive evidence for marsupial care of brood-offspring. Two exceptionally preserved female specimens of Alavatanais carabe and A. margulisae from Late Albian Peñacerrada I amber (Spain) possess four pairs of rudimentary oostegites, indicating formation of a marsupium. From Recent data, given the taxonomic distribution of a marsupium of four pairs of oostegites, we hypothesize that this may be plesiomorphic for the Tanaidomorpha. We also report on a peculiar tanaidacean specimen referable to the fossil family Alavatanaidae, Daenerytanais maieuticus gen. et sp. nov., from Early Cenomanian La Buzinie amber (France), preserved with its marsupial pouch and content. Our discoveries provide early evidence of the peracarid reproductive strategy, as seen in modern Tanaidacea, and argue that this form of parental care may have played a role in the diversification of the lineage during this period.

Antimicrobial Protection of Marsupial Pouch Young.

Marsupials diverged from eutherian mammals about 148 million years ago and represent a unique lineage of mammals with distinctive morphological and reproductive characteristics. Marsupials have significantly shorter gestation periods than eutherians. Pregnancy typically ranges from 15 to 35 days, with young being born at a very early developmental stage and lacking differentiated lymphoid tissues and mature effector cells. Recent microbiome studies of the marsupial pouch revealed that marsupial young can face intense microbial challenges after birth, as the pouch contains a broad range of Gram-positive and Gram-negative bacteria. Antimicrobials are believed to play a significant role in the immune protection of marsupial newborns during their pouch life. The skin of the post-reproductive pouch secretes antimicrobial lysozyme and dermcidin, which may contribute to the decreased density of certain bacteria in the pouch. A range of antimicrobial agents, such as immunoglobulins, lysozyme, transferrin, and cathelicidins, have been identified in marsupial milk. Antimicrobial assays have revealed that marsupial cathelicidins have broad-spectrum activity against a variety of bacteria and fungi, including several multi-drug resistant strains. In this article, we will review the action mechanisms of these antimicrobial compounds and discuss how they protect marsupial newborns from potentially pathogenic bacteria inside the pouch. We will also discuss the potential of marsupial antimicrobial compounds as a source of novel antibiotics.

Genesis of Tropical Storm Debby (2006) within an African Easterly Wave: Roles of the Bottom-Up and Midlevel Pouch Processes

Abstract The “bottom up” generation of low-level vortices (LVs) and midlevel vortices (MVs) during the genesis of Tropical Storm Debby (2006) and the roles of a midlevel “marsupial pouch” associated with an African easterly wave (AEW) are examined using an 84-h simulation with the finest grid size of 1.33 km. Results show that several MVs are generated in leading convective bands and then advected rearward into stratiform regions by front-to-rear ascending flows. Because of different Lagrangian storm-scale circulations, MVs and LVs are displaced along different paths during the early genesis stages. MVs propagate cyclonically inward within the AEW pouch while experiencing slow intensification and merging under the influence of converging flows. The MVs’ merging into a mesovortex is accelerated as they come closer to each other in the core region. In contrast, the low-level Lagrangian circulation is opened as a wave trough prior to tropical depression (TD) stage, so the LVs tend to “escape” from the pouch region. Only after the low-level flows become closed do some LVs congregate and contribute directly to Debby’s genesis. The TD stage is reached when the midlevel mesovortex and an LV are collocated with a convective zone having intense low-level convergence. Results also show the roles of upper-level warming in hydrostatically maintaining the midlevel pouch and producing mesoscale surface pressure falls. It is found that the vertically tilted AEW with a cold dome below is transformed to a deep warm-core TD vortex by subsiding motion. A conceptual model describing the key elements in the genesis of Debby is also provided.

Ovoviviparous reproduction in Australian specimens of the intertidal isopod Cirolana harfordi

Cirolana harfordi is an intertidal isopod found in the SW Pacific (Eastern Australia: New South Wales and Victoria), NW Pacific (Japan and Russia) and the Pacific coast of North America. C. harfordi from North America has been reported to reproduce by depositing eggs into a marsupial pouch formed by oostegite plates that grow from the base of the 3rd to 6th pereopods (walking legs), so as to incubate embryos and mancas (young that resemble the adult except the seventh pair of pereopods are absent) external to the ventral cuticle of the mother. In this study, however, C. harfordi individuals from New South Wales, Australia were found to incubate embryos and mancas inside the pereon (thorax) cavity. C. harfordi in Australia is, therefore, displaying ovoviviparous reproduction and not incubation in a marsupial pouch external to the pereon as previously thought. Embryos have a direct development pathway to the internal manca stage that is held within the mother’s body cavity until the time of live birth. Additionally, internal mancas show some morphological differences to adults, such as a lack of cusps on the mandibles.

The development of the immune tissues in marsupial pouch young

Current knowledge of the development of the marsupial immune system, particularly in the context of lymphoid tissue development and the appearance of lymphocytes, has been examined and limitations identified. While primary lymphoid tissues like the thymus have been extensively studied, secondary lymphoid tissues such as the spleen and lymph nodes have been examined to a lesser extent, partly due to the difficulty of macroscopically identifying these structures, particularly in very small neonates. In addition, little research has been conducted on the mucosal-associated lymphoid tissues; tissues that directly trap antigens and play an important role in the maturity of adaptive immune responses. Research on the development of the marsupial immune tissues to date serves as a solid foundation for further research, particularly on the mechanisms behind the development of the immune system of marsupials. With the recent sequencing and annotation of whole marsupial genomes, the current wealth of sequence data will be essential in the development of marsupial specific reagents, including antibodies, that are required to widen our specific knowledge of the complex marsupial immune system and its development.

Rudimentary Marsupial Pouch in Man.

Rudimentary Marsupial Pouch in Man.

The importance of low-deformation vorticity in tropical cyclone formation

Abstract. Studies of tropical cyclone (TC) formation from tropical waves have shown that TC formation requires a wave-relative quasi-closed circulation: the "marsupial pouch" concept. This results in a layerwise nearly contained region of atmosphere in which the modification of moisture, temperature and vorticity profiles by convective and boundary layer processes occurs undisturbed. The pouch concept is further developed in this paper. TCs develop near the centre of the pouch where the flow is in near solid body rotation. A reference-frame independent parameter is introduced that effectively measures the level of solid-body rotation in the lower troposphere. The parameter is the product of a normalized Okubo-Weiss parameter and absolute vorticity (OWZ). Using 20 yr of ERA-interim reanalysis data and the IBTrACS global TC database, it is shown 95% of TCs including, but not limited to, those forming in tropical waves are associated with enhanced levels of OWZ on both the 850 and 500 hPa pressure levels at the time of TC declaration, while 90% show enhanced OWZ for at least 24 h prior to declaration. This result prompts the question of whether the pouch concept extends beyond wave-type formation to all TC formations world-wide. Combining the OWZ with a low vertical shear requirement and lower troposphere relative humidity thresholds, an imminent genesis parameter is defined. The parameter includes only relatively large-scale fluid properties that are resolved by coarse grid model data (>150 km), which means it can be used as a TC detector for climate model applications. It is also useful as a cyclogenesis diagnostic in higher resolution models such as real-time global forecast models.

An Architectonic Study of the Neocortex of the Short-Tailed Opossum (Monodelphis domestica)

Short-tailed opossums (Monodelphis domestica) belong to the branch of marsupial mammals that diverged from eutherian mammals approximately 180 million years ago. They are small in size, lack a marsupial pouch, and may have retained more morphological characteristics of early marsupial neocortex than most other marsupials. In the present study, we used several different histochemical and immunochemical procedures to reveal the architectonic characteristics of cortical areas in short-tailed opossums. Subdivisions of cortex were identified in brain sections cut in the coronal, sagittal, horizontal or tangential planes and processed for a calcium-binding protein, parvalbumin (PV), neurofilament protein epitopes recognized by SMI-32, the vesicle glutamate transporter 2 (VGluT2), myelin, cytochrome oxidase (CO), and Nissl substance. These different procedures revealed similar boundaries among areas, suggesting that functionally relevant borders were detected. The results allowed a fuller description and more precise demarcation of previously identified sensory areas, and the delineation of additional subdivisions of cortex. Area 17 (V1) was especially prominent, with a densely populated layer 4, high myelination levels, and dark staining of PV and VGluT2 immunopositive terminations. These architectonic features were present, albeit less pronounced, in somatosensory and auditory cortex. The major findings support the conclusion that short-tailed opossums have fewer cortical areas and their neocortex is less distinctly laminated than most other mammals.

Molecular analysis of tammar ( Macropus eugenii) mammary epithelial cells stimulated with lipopolysaccharide and lipoteichoic acid

The immunological function of the metatherian mammary gland plays a crucial part in neonatal survival of the marsupial young. Marsupial pouch young do not develop adult like immune responses until just prior to leaving the pouch. The immune components of the maternal milk secretions are important during this vulnerable early post-partum period. In addition, infection of the mammary gland has not been recognized in metatherians, despite the ready availability of pathogens in the pouch. Regardless of which, little is known about the immunobiology of the mammary gland and the immune responses of mammary epithelial cells in metatherians. In this study, a molecular approach was utilized to examine the response of tammar ( Macropus eugenii) mammary epithelial cells to Escherichia coli derived lipopolysaccharide (LPS) and Staphylococcus aureus derived lipoteichoic acid (LTA). Using custom-made cDNA microarrays, candidate genes were identified in the transciptome, which were involved in antigen presentation, inflammation, cell growth and proliferation, cellular damage and apoptosis. Quantification of mRNA expression of several of these candidate genes, along with seven other genes (TLR4, CD14, TNF-α, cathelicidin, PRDX1, IL-5 and ABCG2) associated with innate immunity in LPS and LTA challenged mammary epithelial cells and leukocytes, was assessed for up to 24 h. Differences in genes associated with cellular damage and pro-inflammatory cytokine production were seen between stimulated mammary epithelial cells and leukocytes. LTA challenge tended to result in lower level induction of pro-inflammatory cytokines, increased PRDX1 mRNA levels, suggesting increased oxidative stress, and increased CD14 expression, but in a non-TLR4-dependent manner. The use of functional genomic tools in the tammar identified differences in the response of tammar mammary epithelial cells (MEC) and leukocytes to challenge with LPS and LTA, and validates the utility of the approach. The results of this study are consistent with a model in which tammar mammary epithelial cells have the capacity to elicit a complex and robust immune response to pathogens.

A proteomic approach to analysis of antimicrobial activity in marsupial pouch secretions

We have documented the antimicrobial activity of pouch secretions of the tammar wallaby, Macropus eugenii, over the period leading up to birth and after birth of the young animal. This activity was greatest against the Gram-negative Escherichia coli and highest at the time of birth. Fractionation of the pouch secretions showed that activity at different times over the reproductive periods was associated with different molecular-weight (MW) components, with compounds in the range up to 50 kDa active immediately prior to and at the time of birth. Proteomic analysis using 1 and 2DE PAGE and LC–MS/MS identified the major components of the pouch secretions at these times, at a range of p I's and MWs. The majority of high-confidence identifications, at a wide range of p I's and MW, were β-lactoglobulin, a known component of marsupial milk. We subsequently conducted a proteomic analysis of mammary gland secretions and digest products from the gut of the young animal, using 2DE PAGE and MALDI MS/MS, to confirm its source and compare it with the observed MW and p I's of β-lactoglobulin. Although we did not directly identify an effector molecule responsible for antimicrobial activity, these results lead us to propose that β-lactoglobulin plays a role in the protection of the young marsupial, a role previously thought to be primarily due to specific secretions from the epithelial surface of the pouch.

Thyroid function and the development of endothermy in a marsupial, the Tasmanian bettong, Bettongia gaimardi (Demarest 1822)

The Tasmanian bettong ( Bettongia gaimardi) is a small rat-kangaroo (marsupial) found only in Tasmania, Australia. The duration of pouch life is 15 weeks. Adults and older young display non-shivering thermogenesis and this paper examines the role of thyroxine in the development of endothermy in pouch young. Free thyroxine (T4) concentrations varied throughout pouch life. The mean (± SE) concentration was 6.2 ± 1.9 pmol L −1 in week 7, increased and peaked at 19.2 ± 4.3 pmol L −1 in week 12, and declined to 5.6 ± 0.4 pmol L −1 by week 20. This was similar to adult levels (3.2 ± 3.8 pmol L −1). These concentrations showed significant differences. From pouch week 12 onwards, T4 injection raised oxygen consumption. Maximum levels of VO 2 after T4 injection occurred at weeks 14–15. Although adult levels were lower, the increase in adult oxygen consumption after T4 injection was about 50%. Peak free T4 levels and metabolic responses to nor-adrenalin occur at week 12 and we hypothesize that thyroid hormone may facilitate the development of adrenergic-receptors in this species. The data presented in the paper further attest to the likely important role of the thyroid gland in the development of endothermy in marsupial pouch young.

Differential expression of the whey acidic protein gene during lactation in the brushtail possum ( Trichosurus vulpecula)

The whey acidic protein (WAP) is a whey protein found in the milk of a number of species. We have isolated and characterised a WAP cDNA clone from the brushtail possum ( Trichosurus vulpecula) and examined its expression in the mammary gland. The amino acid sequences of WAP from the possum and another marsupial, the tammar wallaby, share 69% identity, however, less sequence identity exists between the marsupial and eutherian WAP sequences (30–37%). The possum and tammar WAP genes consist of three four-disulphide core (4-DSC) domains, with a WAP motif at the beginning of each domain. In contrast, the eutherian WAP sequences consist of two 4-DSC domains with the WAP motif only present in the second domain. This WAP motif is also present in a number of protease inhibitors found in a wide range of species. Phylogenetic analysis of marsupial and eutherian WAP sequences suggests that the ancestral WAP gene has three domains and that one of the domains has been deleted from the eutherian gene. The profile of WAP gene expression in the possum mammary gland changed throughout lactation, with WAP mRNA levels reaching a peak between days 106 and 177 of lactation. The level of WAP mRNA in the mammary gland appeared to be correlated with the level of circulating prolactin in the lactating female and was different to that observed for several other whey protein genes. Overlapping expression of the WAP and early lactation protein genes, both of which are putative protease inhibitors, may provide protection of milk immunoglobulins that are required for the prolonged period of passive immune transfer to the marsupial pouch young.

Development of the immune system and immunological protection in marsupial pouch young

At birth the tissues of marsupial immune system are underdeveloped. The young animal is not immunocompetent. Histological and immunohistochemical studies of pouch young epithelial tissues provide a clear picture of tissue development but the timing of onset of immunocompetence awaits definition. The survival of the neonatal marsupial in a microbially rich environment is dependent on maternal strategies, including immunoglobulin transfer via milk and, in some species, prenatally via the yolk sac placenta. It is also likely that pouch secretions play a role. This review summarizes our current knowledge of the pathway of immunological development in marsupials and the protection and threats afforded by the pouch environment.

Immunological protection of the vulnerable marsupial pouch young: two periods of immune transfer during lactation in Trichosurus vulpecula (brushtail possum)

Marsupial young are born with an underdeveloped immune system and are dependent upon passively acquired immune protection provided by the mother’s milk. Colostrum and milk samples were collected from the brushtail possum throughout lactation and the concentration of secretory IgA (sIgA), IgG and transferrin was determined by Western blotting. Two periods of immune transfer were identified. The first, a colostral phase, occurs immediately after birth and involves sIgA, IgG and transferrin. During the early lactation stage, pouch young receive milk of a unique composition as they undergo developmental changes in the pouch that occur in utero for eutherian mammals. At the end of this external gestation, the composition of the milk changes (switch phase) to resemble that of eutherian mammals in the late lactation phase. The second transfer of immunity consists of IgG and transferrin, and occurs during the switch phase prior to maturation of the immune response.

Prevalence of Trypanosoma cruzi infection in opossum (Didelphis albiventris) in Santiago del Estero, Argentina

The opossum of the genus Didelphis is one of the principal wild reservoirs of Trypanosoma cruzi and is widely distributed in the Western Hemisphere. Didelphis albiventris is the most common marsupial in Amamá and Trinidad, two communities in the province of Santiago del Estero, Argentina. The D. albiventris population is replaced every year, and the opossum normally has two reproductive periods, one at the beginning of the spring and another at the beginning of the summer. The two litters are weaned, and they leave the mother's marsupial pouch to join the population, the first (G1) at the beginning of the summer and the second (G2) at the beginning of the fall. Between 1988 and 1991 409 D. albiventris opossums were studied, and xenodiagnoses showed that 35% of them were infected with T. cruzi. Annual cycles of renewed infection were observed, with prevalences that ranged between 22% and 43%. The acquisition of the parasite occurred over the entire year, from the summer through the spring. The prevalence of infection increased with age. The G1 individuals tended to present higher prevalences than the G2 individuals, probably from being exposed to transmission for a longer period of time. In the first two (younger) age categories for the opossums, G2 individuals showed higher prevalences than did the G1 individuals. This indicates a significant increase in transmission intensity during the fall. Opossums should be regarded as a potential source of T. cruzi entry to the domestic transmission cycle.