Reproductive biology of Leptodactylus fuscus (Anura, Leptodactylidae) in the subtropical climate, Rio Grande do Sul, Brazil

The aim of this study was to characterize, for the central region of the State of Rio Grande do Sul, Brazil, the reproductive biology of Leptodactylus fuscus (Schneider, 1799), based on the analysis of gonadal development of males and females, reproductive effort, size-fecundity relationships, and occurrence of sexual dimorphism in body size. Mature individuals were found from October 1996 to February 1997 and from October 1997 to December 1997. The highest input of juveniles in the population was recorded in March 1997. There was a positive and significant correlation between the number of mature individuals and the mean monthly temperature. The population did not present sexual dimorphism in size. Males presented significant correlation only between snout-vent length and testes length. All females had oocytes at four different maturation stages and there were no significant correlations regarding size-fecundity variables. The correlation between ovarian size factor and females snout-vent length was not significant either. The main difference between this population and those that inhabit tropical climate was that temperature was responsible for stimulating the reproduction activity, instead of rainfall.

Spatial and temporal distribution of Pseudis minuta (Anura, Hylidae, Hylinae) and environmental variables related to its reproductive activity in Reserva Biológica do Lami, southern Brazil

We studied the reproductive biology of a population of Pseudis minuta Günther, 1858 from Reserva Biológica do Lami (30º 15' S; 51º 05' W), Porto Alegre, southern Brazil. We assessed the spatial and temporal distribution of individuals (males, females, juveniles) and explored potential relationships with environmental variables. Field activities encompassed bimonthly surveys in three semi-permanent ponds, each one during approximately two days and two nights, from August 2004 to July 2005. We recorded differences in the sites used by males, females and juveniles, with males occupying deeper and more distant places from the border. The temporal distributions of individuals, calling sites and amplectant pairs indicated that the reproductive activity of P. minuta is related to some of the studied abiotic factors. Calling males presented statistical differences in relation to non-calling males for all daily abiotic variables analyzed (air temperature, water temperature, relative humidity and rainfall), as well as to monthly temperature and rainfall. The number of active males, females and juveniles was influenced by at least one of the daily or monthly environmental variables analyzed. We conclude that the reproduction in this species is seasonal and may be partially determined by abiotic factors.

Reproductive modes and fecundity of an assemblage of anuran amphibians in the Atlantic rainforest, Brazil

Reproductive modes and size-fecundity relationships are described for anurans from Picinguaba, a region of Atlantic rainforest on the northern coast of the state of São Paulo, Brazil. We observed 13 reproductive modes, confirming a high diversity of modes in the Atlantic rainforest. This diversity of reproductive modes reflects the successful use of diversified and humid microhabitats by anurans in this biome. We measured the snout-vent length of 715 specimens of 40 species of anurans. The size-fecundity relationship of 12 species was analyzed. Female snout-vent lengths explained between 57% and 81% of clutch size variation. Anurans with aquatic modes laid more eggs than those with terrestrial or arboreal modes. Larger eggs were deposited by species with specialized reproductive modes.

Reproductive traits of Gymnogeophagus labiatus (Teleostei, Cichlidae), in an upper stretch of Sinos river, Caraá, Brazil

This work describes the reproduction of Gymnogeophagus labiatus (Hensel, 1870) from an upper stretch of Sinos river, southern Brazil, based on the analysis of 174 males and 132 females captured in monthly samples taken from January to December 2007. Results showed that reproductive activity occur in spring and summer although ripe males were found along the year. The standard length of the smallest ripe male was 104.74 mm (Lt) and the smallest ripe female was 55.00 mm (Lt). There was a significant difference in total sex ratio, with 1.32 males to each female (χ2 = 5.76). Males were much more abundant in March (1.75 males: 1 female) and December (5 males: 1 female). Females were more abundant in the 62├77 mm interval (1 male: 2.36 female) while males were more abundant in the 77├92 mm size interval (2.57 males: 1 female). The largest length intervals were composed of only males. Mean absolute fecundity was 113.4 (± 31.24 sd) and mean relative fecundity was 0.0125 (± 0.0026 sd) oocytes/mg. In ripe ovaries, small-diameter oocytes were observed at high frequencies while larger ones occurred at lower frequencies. This pattern is common in fishes with asynchronous oocyte development. Characteristics of G. labiatus, such as low fecundity, asynchrony in oocyte development, multiple spawning, and its well-known parental care behavior, are consistent with an equilibrium strategy, as proposed for other cichlids.

Compared nymphal development of Tingis americana (Hemiptera, Tingidae) in two Handroanthus species (Bignoniaceae) and reproductive parameters in seedlings of Handroanthus heptaphyllus

Nymphs and adults of Tingis americana Drake, 1922 were found feeding on leaves of Handroanthus heptaphyllus (Vell.) Mattos and Handroanthus chrysotrichus (Mart. ex A. DC.) Mattos in the Botanic Garden, Porto Alegre, state of Rio Grande do Sul, Brazil. This is the first record of T. americana on these host plants and in the southern Brazil. We aimed to compare the nymphal development on both hosts and to analyze the reproductive parameters on H. heptaphyllus (25 ± 1ºC; 60 ± 10% RH; 16 h photophase). The mean nymphal period (days) was shorter in individuals reared on H. heptaphyllus (12.69 ± 0.076) than on H. chrysotrichus (19.11 ± 0.208) (P < 0.0001), however, nymph viability was similar. On H. heptaphyllus, the mean embryonic period lasted 12.32 ± 0.274 days and the egg viability was 92%. The mean total and daily fecundity were 310.0 ± 19.40 eggs/female and 7.46 ± 0.302 eggs/female/day, respectively. Paired males and females showed similar longevity (P = 0.0691), while unpaired females lived longer than unpaired males (P = 0.0460).

Reproductive aspects of Dendropsophus sanborni (Anura, Hylidae) in northeastern Argentina

Reproductive aspects of Dendropsophus sanborni (Anura, Hylidae) in northeastern Argentina. The aim of this investigation was to characterize reproductive aspects of Dendropsophus sanborni (Schmidt, 1944) (Anura, Hylidae) in the northwest of Corrientes province, in northeastern Argentina. The reproductive cycles of males and females, the existence of sexual dimorphism and the male’s vocalization period were analyzed. Samplings were made from September 2011 to August 2012. Dendropsophus sanborni showed a prolonged reproductive pattern because reproductive activity was observed during most part of the year. Calling males were recorded all months except in July. The spermatogenic cycle can be characterized as potentially continuous. In males, snout-vent length was significantly correlated with mean testicular volume. The species showed sexual dimorphism in body mass, with females larger than males. Post-ovulatory females were found from March to June and most females had oocytes at different stages of maturity, with abundant post-vitellogenic oocytes from August to December.

Reproductive strategy of a non-annual rivulid in a perennial wetland

ABSTRACT Atlantirivulus riograndensis (Costa & Lanés, 2009) is a fish registered to the basin of Patos lagoon and the adjacent coastal plains in southern states of Rio Grande do Sul and Santa Catarina, Brazil, found in shallow water courses with that have large quantities of aquatic vegetation and forest edges. The objective of this study was to investigate the reproductive strategy and tactics of this species including the sex ratio, the length at first maturity, spawning type, fecundity and the possible associations among reproduction and abiotic factors. Sampling of specimens occurred in perennial wetlands within the Banhado dos Pachecos wildlife refuge, in the city of Viamão, state of Rio Grande do Sul, Brazil, which is a conservation unit that belongs to an area of environmental protection in the Pampa Biome. The capture of 30 A. riograndensis specimens per month occurred from January to December of 2012. A total of 188 females and 172 males were captured and the total sex ratio was 1:1 in the sampled population. Sexual maturity of the species occurs after 13.59 and 11.92 mm (SL) for females and males, respectively. Both a multiple spawning and a long reproductive period (since August to March) were confirmed by the presence of post-ovulatory follicles that were observed through histological analysis and the values of the gonadosomatic index in females considered spawning capable. The average absolute fecundity of the species is of 19.33 (± 6.18) vitellogenic oocytes in mature ovaries. No significant relationship was found between mean GSI and the abiotic data. Reproductive tactics presented by A. riograndensis indicate a species with an opportunistic reproductive strategy, following the pattern of other species of the Rivulidae family.

Collaboration strategies and technological innovation: a contractual perspective of the relationship between firms and techonological centers

In this paper, a theoretical framework for analyzing the selection of governance structures for implementing collaboration agreements between firms and Technological Centers is presented and empirically discussed. This framework includes Transaction Costs and Property Rights’ theoretical assumptions, though complemented with several proposals coming from the Transactional Value Theory. This last theory is used for adding some dynamism in the governance structure selection. As empirical evidence of this theoretical explanation, we analyse four real experiences of collaboration between firms and one Technological Center. These experiences are aimed to represent the typology of relationships which Technological Centers usually face. Among others, a key interesting result is obtained: R&D collaboration activities do not need to always be organized through hierarchical solutions. In those cases where future expected benefits and/or reputation issues could play an important role, the traditional more static theories could not fully explain the selected governance structure for managing the R&D relationship. As a consequence, these results justify further research about the adequacy of the theoretical framework presented in this paper in other contexts, for example, R&D collaborations between firms and/or between Universities or Public Research Centers and firms.

Firm strategies in R&D: cooperation and participation in R&D programs

This paper presents empirical evidence on firms’ decisions to cooperate in R&D and the extent to which government sponsored R&D programs increase cooperation. Using a sample of firms from the Spanish innovation survey we jointly estimate the determinants of firm participation in R&D programs and of choice of cooperation partners. We find that (i) firms participating in national and European programs have different profiles, suggesting program complementarity; (ii) private-private and publicprivate cooperation are associated on average with firms with different characteristics, and (iii) national R&D programs seem to have a positive effect on all types of cooperation, but especially on public-private partnerships.

Reproductive isolation between different forms of Lutzomyia longipalpis (Lutz & Neiva), (Diptera: Psychodidae), the vector of Leishmania donovani chagasi Cunha & Chagas and its significance to kala-azar distribution in South America

The males of the sandfly Lutzomyia longipalpis occur in two forms, one which bears a single pair of pale spots on tergite 4 and another in which an additional pair of spots characterizes tergite 3. In crosses between laboratory reared stocks of the two forms originating from allopatric and sympatric sites in Brazil nearly all males of one form fail to inseminate females of the other. In addition, insemination failure between some allopatric populaytions of Lu. longipalpis with similar tergal spot patterns is recorded, indicating the existence of additional forms in an apparent species complex. The possibility that Lu. longipalpis sensu latu represents more than a single taxon is discussed and the relevance of these findings to future epidemiological studies on kala-azar is considered.

Development of metabolic labeling strategies to study changes in protein expression

Résumé : Les progrès techniques de la spectrométrie de masse (MS) ont contribué au récent développement de la protéomique. Cette technique peut actuellement détecter, identifier et quantifier des milliers de protéines. Toutefois, elle n'est pas encore assez puissante pour fournir une analyse complète des modifications du protéome corrélées à des phénomènes biologiques. Notre objectif était le développement d'une nouvelle stratégie pour la détection spécifique et la quantification des variations du protéome, basée sur la mesure de la synthèse des protéines plutôt que sur celle de la quantité de protéines totale. Pour cela, nous volions associer le marquage pulsé des protéines par des isotopes stables avec une méthode d'acquisition MS basée sur le balayage des ions précurseurs (precursor ion scan, ou PIS), afin de détecter spécifiquement les protéines ayant intégré les isotopes et d'estimer leur abondance par rapport aux protéines non marquées. Une telle approche peut identifier les protéines avec les plus hauts taux de synthèse dans une période de temps donnée, y compris les protéines dont l'expression augmente spécifiquement suite à un événement précis. Nous avons tout d'abord testé différents acides aminés marqués en combinaison avec des méthodes PIS spécifiques. Ces essais ont permis la détection spécifique des protéines marquées. Cependant, en raison des limitations instrumentales du spectromètre de masse utilisé pour les méthodes PIS, la sensibilité de cette approche s'est révélée être inférieure à une analyse non ciblée réalisée sur un instrument plus récent (Chapitre 2.1). Toutefois, pour l'analyse différentielle de deux milieux de culture conditionnés par des cellules cancéreuses humaines, nous avons utilisé le marquage métabolique pour distinguer les protéines d'origine cellulaire des protéines non marquées du sérum présentes dans les milieux de culture (Chapitre 2.2). Parallèlement, nous avons développé une nouvelle méthode de quantification nommée IBIS, qui utilise des paires d'isotopes stables d'acides aminés capables de produire des ions spécifiques qui peuvent être utilisés pour la quantification relative. La méthode IBIS a été appliquée à l'analyse de deux lignées cellulaires cancéreuses complètement marquées, mais de manière différenciée, par des paires d'acides aminés (Chapitre 2.3). Ensuite, conformément à l'objectif initial de cette thèse, nous avons utilisé une variante pulsée de l'IBIS pour détecter des modifications du protéome dans des cellules HeLa infectée par le virus humain Herpes Simplex-1 (Chapitre 2.4). Ce virus réprime la synthèse des protéines des cellules hôtes afin d'exploiter leur mécanisme de traduction pour la production massive de virions. Comme prévu, de hauts taux de synthèse ont été mesurés pour les protéines virales détectées, attestant de leur haut niveau d'expression. Nous avons de plus identifié un certain nombre de protéines humaines dont le rapport de synthèse et de dégradation (S/D) a été modifié par l'infection virale, ce qui peut donner des indications sur les stratégies utilisées par les virus pour détourner la machinerie cellulaire. En conclusion, nous avons montré dans ce travail que le marquage métabolique peut être employé de façon non conventionnelle pour étudier des dimensions peu explorées en protéomique. Summary : In recent years major technical advancements greatly supported the development of mass spectrometry (MS)-based proteomics. Currently, this technique can efficiently detect, identify and quantify thousands of proteins. However, it is not yet sufficiently powerful to provide a comprehensive analysis of the proteome changes correlated with biological phenomena. The aim of our project was the development of ~a new strategy for the specific detection and quantification of proteomé variations based on measurements of protein synthesis rather than total protein amounts. The rationale for this approach was that changes in protein synthesis more closely reflect dynamic cellular responses than changes in total protein concentrations. Our starting idea was to couple "pulsed" stable-isotope labeling of proteins with a specific MS acquisition method based on precursor ion scan (PIS), to specifically detect proteins that incorporated the label and to simultaneously estimate their abundance, relative to the unlabeled protein isoform. Such approach could highlight proteins with the highest synthesis rate in a given time frame, including proteins specifically up-regulated by a given biological stimulus. As a first step, we tested different isotope-labeled amino acids in combination with dedicated PIS methods and showed that this leads to specific detection of labeled proteins. Sensitivity, however, turned out to be lower than an untargeted analysis run on a more recent instrument, due to MS hardware limitations (Chapter 2.1). We next used metabolic labeling to distinguish the proteins of cellular origin from a high background of unlabeled (serum) proteins, for the differential analysis of two serum-containing culture media conditioned by labeled human cancer cells (Chapter 2.2). As a parallel project we developed a new quantification method (named ISIS), which uses pairs of stable-isotope labeled amino acids able to produce specific reporter ions, which can be used for relative quantification. The ISIS method was applied to the analysis of two fully, yet differentially labeled cancer cell lines, as described in Chapter 2.3. Next, in line with the original purpose of this thesis, we used a "pulsed" variant of ISIS to detect proteome changes in HeLa cells after the infection with human Herpes Simplex Virus-1 (Chapter 2.4). This virus is known to repress the synthesis of host cell proteins to exploit the translation machinery for the massive production of virions. As expected, high synthesis rates were measured for the detected viral proteins, confirming their up-regulation. Moreover, we identified a number of human proteins whose synthesis/degradation ratio (S/D) was affected by the viral infection and which could provide clues on the strategies used by the virus to hijack the cellular machinery. Overall, in this work, we showed that metabolic labeling can be employed in alternative ways to investigate poorly explored dimensions in proteomics.

Changes in reproductive roles are associated with changes in gene expression in fire ant queens.

Abstract In species with social hierarchies, the death of dominant individuals typically upheaves the social hierarchy and provides an opportunity for subordinate individuals to become reproductives. Such a phenomenon occurs in the monogyne form of the fire ant, Solenopsis invicta, where colonies typically contain a single wingless reproductive queen, thousands of workers and hundreds of winged nonreproductive virgin queens. Upon the death of the mother queen, many virgin queens shed their wings and initiate reproductive development instead of departing on a mating flight. Workers progressively execute almost all of them over the following weeks. To identify the molecular changes that occur in virgin queens as they perceive the loss of their mother queen and begin to compete for reproductive dominance, we collected virgin queens before the loss of their mother queen, 6 h after orphaning and 24 h after orphaning. Their RNA was extracted and hybridized against microarrays to examine the expression levels of approximately 10 000 genes. We identified 297 genes that were consistently differentially expressed after orphaning. These include genes that are putatively involved in the signalling and onset of reproductive development, as well as genes underlying major physiological changes in the young queens.