Sexing Sirenians: Validation of Visual and Molecular Sex Determination in Both Wild Dugongs (Dugong dugon) and Florida Manatees (Trichechus manatus latirostris)

Sexing wild marine mammals that show little to no sexual dimorphism is challenging. For sirenians that are difficult to catch or approach closely, molecular sexing from tissue biopsies offers an alternative method to visual discrimination. This paper reports the results of a field study to validate the use of two sexing methods: (1) visual discrimination of sex vs (2) molecular sexing based on a multiplex PCR assay which amplifies the male-specific SRY gene and differentiates ZFX and ZFY gametologues. Skin samples from 628 dugongs (Dugong dugon) and 100 Florida manatees (Trichechus manatus latirostris) were analysed and assigned as male or female based on molecular sex. These individuals were also assigned a sex based on either direct observation of the genitalia and/or the association of the individual with a calf. Individuals of both species showed 93 to 96% congruence between visual and molecular sexing. For the remaining 4 to 7%, the discrepancies could be explained by human error. To mitigate this error rate, we recommend using both of these robust techniques, with routine inclusion of sex primers into microsatellite panels employed for identity, along with trained field observers and stringent sample handling.

Sexing up the city in the international beauty contest: the performative nature of spatial planning and the fictive spectacle of place branding

This paper responds to recent calls for more academic research and critical discussion on the relationship between spatial planning and city branding. Through the lens of Liverpool, the article analyses how key planning projects have delivered major transformations in the city's built environment and cultural landscape. More specifically, in concentrating on the performative nature of spatial planning it reveals the physical, symbolic and discursive re-imaging of Liverpool into a 'world class city'. Another aspect of the paper presents important socioeconomic datasets and offers a critical reading of the re-branding in showing how it presents an inaccurate representation of Liverpool. The evidence provided indicates that a more accurate label for Liverpool is a polarised and divided city, thereby questioning the fictive spectacle of city branding. Finally, the paper ends with some critical commentary on the role of spatial planning as an accessory to the sophistry of city branding.

Biometrics and sexing criteria of the yellow-faced honeyeater lichenostomus chrysops

Morphometric data on 99 adult and 13 juvenile Yellow-faced Honeyeaters Lichenostomus chrysops that were independently sexed using molecular techniques were analysed to investigate size dimorphism between the sexes. Our results support previous studies that have demonstrated Yellow-faced Honeyeaters are sexually dimorphic in size, with males being the larger sex. Discriminant analyses of morphometric data were used to develop a simple method for sexing adult Yellow-faced Honeyeaters in the hand. As five observers collected the measurements our sexing criteria are conservative and should have wide application for field ornithologists working on the species.

Methods for aging and sexing the Black-eared Miner, Manorina melanotis, and Yellow-throated Miner, M. flavigula

Morphometric data on 92 Black-eared Miners and 47 Yellow-throated Miners that had been independently sexed using molecular techniques were analysed to investigate size dimorphism between the sexes. We found that both species are sexually dimorphic in size, with males being the larger sex. Discriminant analyses of morphometric data were used to develop a simple method for sexing both species in the hand. Additionally, alula shape was consistent with other methods that we applied for ageing individuals. Sex-specific size differences between Black-eared and Yellow-throated Miners detected here add further support to the contention that they represent different taxa. The application of these sexing and ageing techniques for both species of mallee miner will improve ongoing field management of the endangered Black-eared Miner.

A morphological model for sexing nestling peregrine falcons (Falco Peregrinus Macropus) verified through genetic analysis

Adult peregrine falcons (Falco peregrinus macropus) have monotypic plumage and display strong reversed sexual dimorphism, with females significantly larger than males. Reversed sexual dimorphism is measurable among nestlings in the latter stages of their development and can therefore be used to differentiate between sexes. In the early stages of development, however, nestlings cannot be sexed with any degree of certainty because morphological differentiation between the sexes is not well developed. During this study we developed a model for sexing younger nestlings based on genetic analysis and morphometric data collected as part of a long-term banding study of this species. A discriminant function model based on morphological characteristics was developed for determining the sex of nestlings (n = 150) in the field and was shown to be 96.0% accurate. This predictive model was further tested against an independent morphometric dataset taken from a second group of nestlings (n = 131). The model correctly allocated sex to 96.2% of this second group of nestlings. Sex can reliably be determined (98.6% accurate) for nestlings that have a wing length of at least 9 cm using this model. Application of this model, therefore, allows the banding of younger nestlings and, as such, significantly increases the period of time over which banding can occur. Another important implication of this model is that by banding nestlings earlier, they are less likely to jump from the nest, therefore reducing the risk of injury to both the brood and the bander.

Sexing little penguins eudyptula minor using bill measurements

In Little Penguins Eudyptula minor there are no reliable plumage or body size differences that can be used visually to distinguish the sex of individuals. However, sexual dimorphism of morphometric measures has been noted, with males always being a little larger than females. In this study, differences between E. minor sexes at eight colonies in south-eastern Australia were determined statistically via discriminant function analysis (DFA) and through the utilization of DNA-based techniques developed for non-ratite birds. The DFA correctly determined gender in 91.1% of cases and molecular methods were 100% accurate. Our DFA success rate of classification is similar to that previously published for Little Penguins in Victoria. In this study statistically significant differences in mean bill depths and lengths were found between Little Penguin colonies at St Kilda, Phillip Island and Gabo Island, compared to colonies at Kangaroo Island, Granite Island, Middle Island and London Bridge. As birds in eastern populations (St Kilda, Phillip Island, Gabo Island) exhibit statistically significantly smaller beaks (bill depth and bill length), separate discriminant functions were investigated for each phenotypically distinct geo-spatial cohort. Interestingly, cluster analysis for bill length identified three groups: western (Kangaroo Island and Granite Island), eastern (St Kilda, Phillip Island and Middle Island) and the London Bridge Little Penguin colony, which constituted a separate group. We conclude that while there is a slight increase in DF power for colonies west of Cape Otway and for some specific colonies, colony-specific DFA is not required to identify the sex of Little Penguins in south-eastern Australia.

Sexing accuracy and indicators of maturation status in captive Murray cod Maccullochella peelii peelii using non-invasive ultrasonic imagery

Macroscopic- and histological-based assessments of gonad condition were compared with ultrasound images to determine the feasibility of this technology as a non-invasive diagnostic tool for identifying sex and assessing maturation status of Murray cod. Four age-classes (1⁺, 2⁺, 3⁺ and 6⁺ years), were sub-sampled at monthly intervals throughout their annual reproductive cycle and scanned with a 5 MHz linear transducer. An interpretation of sex was made from the resulting images and maximum cross-sectional gonad diameter and area were recorded. Fish were subsequently dissected to confirm gender, and the weights and maturation status of gonads determined and then compared with their respective image profile. Ovaries of females were usually a distinctive feature in ultrasound images, being particularly obvious in older and/or more developed fish. In contrast, the identification of male testis was more problematic. Nonetheless, identifying sex from ultrasound images was consistently achieved by recording the presence/absence of a female ovary (96% total sexing accuracy). Maximum cross-sectional ovary diameter and area were highly correlated with gonad weight (r² = 0.90 and 0.89, respectively) suggesting that indices of maturation status, comparable to the gonadosomatic index (GSI), can be obtained non-destructively from ultrasound scans of females. A less distinct relationship occurred between these dimensions and weight of testes (r² = 0.41). Significant increases (P < 0.05) in mean gonad index (GI, calculated from gonad diameter) occurred for most gonad development stages. However, differences in mean GI between maturation stages were confounded by phenotypic variability, indicating that GI may be limited to population level studies. Nevertheless, ultrasound images of ovaries at each development stage were visually distinctive and enabled qualitative evaluations of maturity, thereby complementing quantitative GI assessments. Repeated serial-monitoring of the same population using ultrasound appears to have great potential for tracking maturation-induced changes in broodfish.

An evaluation of three field techniques for sexing live Gould's petrels (Pterodroma leucoptera) (Procellariidae)

Many petrels show no obvious sex-linked dimorphism in plumage or size and consequently many researchers fail to sex the living individuals they study. Several methods of sex discrimination that do not rely on plumage- or obvious size-dimorphism can be used to sex live petrels. The effectiveness of three such techniques was evaluated: body condition at the time of laying, cloacal inspection, and discriminant function analysis (DFA) of external morphometrics. Gould’s Petrel (Pterodroma leucoptera leucoptera) was used as the subject species. Sexing of breeding adults on the basis of body condition at laying proved to be highly accurate (100% of birds sexed correctly) but required detailed knowledge of the breeding biology. Following training, cloacal inspection proved to be an accurate (96%) method of determining the sex of breeding adults, but not of chicks. Unlike molecular sexing, the latter two methods of sex discrimination provide immediate knowledge of the sex of individuals in the field. DFA of external morphometrics predicted the sex of adults with an accuracy of 73% and the sex of near-fledged chicks with an accuracy of 66%. However, the probability of correct assignment of sex was low in most cases and, therefore, this is the least useful of the three techniques assessed here.

New and improved molecular sexing methods for museum bird specimens

We present two new avian molecular sexing techniques for nonpasserine and passerine birds (Neognathae), which are more suitable for use with museum specimens than earlier methods. The technique for nonpasserines is based on a new primer (M5) which, in combination with the existing P8 primer, targets a smaller amplicon in the CHD1 sex-linked gene than previously. Primers targeting ATP5A1, an avian sex-linked gene not previously used for sex identification, were developed for passerines. Comprehensive testing across species demonstrated that both primer pairs sex a range of different species within their respective taxonomic groups. Rigorous evaluation of each method within species showed that these permitted sexing of specimens dating from the 1850s. For corn bunting museum specimens, the ATP5A1 method sexed 98% of 63 samples (1857–1966). The M5/P8 CHD1 method was similarly successful, sexing 90% of 384 moorhen specimens from six different museum collections (1855–2001). In contrast, the original P2/P8 CHD1 sexing method only identified the sex of less than half of 111 museum moorhen samples. In addition to dried skin samples, these methods may be useful for other types of material that yield degraded or damaged DNA, and are hence potential new sexing tools for avian conservation genetics, population management and wildlife forensics.

Sexing of murine and bovine embryos by developmental high-titer arrest induced by H-Y antisera

Murine and bovine embryos at the late morula stage were cultured in medium containing high-titer rat H-Y antisera. After 12 h of incubation, embryos blocked at the late morulae stage were classified as males and those at the blastocyst stage were classified as females. Sexing of murine embryos by PCR and cytogenetics revealed that 83% of the embryos classified as males and 82% of those classified as females had their sex correctly predicted (P < 0.05). Bovine embryos were transferred to recipient females. Pregnancy rates were 71.4% (10/14) for embryos classified as males and 68.8% (11/16) for embryos classified as females. The sex was correctly predicted for 80% (8/10) of the embryos classified as males and for 81.8% (9/11) of those classified as females (overall accuracy, 80.9%, P < 0.05). Therefore, the induction of developmental arrest by high-titer male-specific antisera was an efficient strategy for non-invasive embryo sexing. The procedure was straightforward and has considerable commercial potential for sexing bovine embryos. (c) 2004 Elsevier B.V. All rights reserved.

Development of a real-time PCR method for rapid sexing of human preimplantation embryos

Genes on the X chromosome are known to be responsible for more than 200 hereditary diseases. After IVF, the simple selection of embryo sex before uterine transfer can prevent the occurrence of affected offspring among couples at risk for these genetic disorders. The aim of this investigation was to develop a rapid method of preimplantation genetic diagnosis (PGD) using real-time polymerase chain reaction (PCR) for the sexing of human embryos, and to compare it to the fluorescence in-situ hybridization technique, considered to be the gold standard. After biopsies were obtained from 40 surplus non-viable embryos for transfer, a total of 98 blastomeres were analysed. It was possible to analyse 24 embryos (60%) by both techniques, generating a total of 70 blastomeres (35 per technique), white 28 blastomeres from 16 embryos (40%) were analysed only by real-time PCR. A rapid and safe method was developed in the present study for the sexual diagnosis of a single human cell (blastomere and buccal cell) using the emerging technology of real-time PCR. (C) 2009, Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.