Biology

Understanding coral responses to environmental change is critical to predicting their health and future distribution. While tropical and subtropical corals generally experience limited environmental changes, temperate corals undergo pronounced seasonal cycles in irradiance, temperature and nutrients. Their ability to cope with environmental changes makes temperate organisms an ideal model to investigate the effects of global change on coral physiology.

The spatial scale of life-history and demographic variation was investigated in the opportunistic polychaete Ophryotrocha labronica La Greca and Bacci. Individuals were collected along the Italian coasts from three thermally different biogeographical regions of the Mediterranean Sea. For each region, populations from four harbours were considered, and for each harbour, two sites were examined. Life-history and demographic traits were investigated after one generation under a common garden experiment, and their variation at the three spatial scales was assessed. All the traits showed high variability with regard to site. A number of life-history and all demographic traits also varied according to the biogeographical region. Conversely, no differences were found between harbours, suggesting that geographical isolation did not contribute to phenotypic variation. Results confirmed the central role of local conditions for the evolution of life history in species colonizing heterogeneous environments, but they also pointed to the importance of large-scale factors in shaping the phenotypic responses of O. labronica, demonstrating the need for a multi-scale approach for obtaining a good measure of natural variation in widespread opportunistic species.

1The relationships between life history, fitness and sexuality, together with their ecological and evolutionary significance, has been analysed comparing the main life-history traits and demography in three closely related species belonging to the genus Ophryotrocha. The species are: the gonochoristic O. labronica, the simultaneous hermaphrodite O. diadema and the protandrous hermaphrodite O. puerilis.2Survivorship and reproductive data were collected weekly and were used to construct life tables and population projection matrices for each species and compare life-history characteristics. Elasticity, life-table response and decomposition analyses were performed to examine the relative contribution of fecundity and survivorship to differences in λ between species.3The gonochoristic and hermaphroditic species differ in all the main life-history parameters and also in demographic characteristics. In particular the value of λ, used commonly to express fitness, is markedly higher in the gonochoristic species while in terms of fitness simultaneous and sequential hermaphroditism are very similar. In the genus Ophryotrocha gonochorism currently represents the most widespread condition, being characteristic of the majority of the known species in the genus.4Given the demographic advantage ensured by gonochorism, it remains be understood why some species have retained simultaneous hermaphroditism and one has evolved a sequential type hermaphroditism; the most probable hypothesis is correlated with the density of the species in natural habitats.The relationships between life history, fitness and sexuality, together with their ecological and evolutionary significance, has been analysed comparing the main life-history traits and demography in three closely related species belonging to the genus Ophryotrocha. The species are: the gonochoristic O. labronica, the simultaneous hermaphrodite O. diadema and the protandrous hermaphrodite O. puerilis.Survivorship and reproductive data were collected weekly and were used to construct life tables and population projection matrices for each species and compare life-history characteristics. Elasticity, life-table response and decomposition analyses were performed to examine the relative contribution of fecundity and survivorship to differences in λ between species.The gonochoristic and hermaphroditic species differ in all the main life-history parameters and also in demographic characteristics. In particular the value of λ, used commonly to express fitness, is markedly higher in the gonochoristic species while in terms of fitness simultaneous and sequential hermaphroditism are very similar. In the genus Ophryotrocha gonochorism currently represents the most widespread condition, being characteristic of the majority of the known species in the genus.Given the demographic advantage ensured by gonochorism, it remains be understood why some species have retained simultaneous hermaphroditism and one has evolved a sequential type hermaphroditism; the most probable hypothesis is correlated with the density of the species in natural habitats.

In the harbour of La Spezia (Italy) six polychaete species belonging to the family Dorvilleidae are present. The population dynamics of Dinophilus gyrociliatus, Ophryotrocha hartmanni, O. japonica, O. labronica, O. puerilis and Schistomeringos rudolphii has been studied from August 2002 to August 2003. The density of each species was considerably different and during the year there was a species succession that alternated in dominating the community. The role of the life-history characteristics of each species and environmental conditions in determining the population dynamics are discussed.

In order to gain insights into links between fitness and physiological function, the phenotypic plasticity of life-history traits and thermal tolerances has been investigated in a laboratory strain of the polychaete Ophryotrocha labronica acclimated to a broad range of temperatures. Reproductive pairs were kept in isolation for two months at one of six temperatures (10, 15, 20, 25, 30 and 35 °C), and survival and a number of life-history traits recorded. Tolerance to heat and cold was assessed in individuals that survived the accli-
mation period by measuring upper and lower thermal limits. Life-history and thermal tolerance traits showed different relationships with temperature: generally non-monotonic for life-history and monotonic for thermal limits. The thermal history of the strain used in the study seemed to be partly responsible for
this pattern, affecting the thermal responses of most life-history traits, but not those of the upper and lower thermal limits. Survival correlated positively with tolerance to cold but not with heat tolerance. This result supports the idea that tolerance to heat is under stronger selection in O. labronica, resulting in a
lower level of plasticity for this trait. Some life-history features positively co-varied with tolerance to heat
(e.g. body size), whilst in other cases reproductive traits showed apparent trade-offs with thermal limits.
In all cases correlations amongst traits were apparently driven by their relationship with temperature.
Even if we found no evolutionary trade-offs among the thermal responses of life-history traits and thermal limits, the strategies evolved by organisms to cope with thermal variability result from the mutual interaction of these traits, which plays a key role in defining organisms' responses to temperature changes and
fluctuations.

Recent efforts of researchers to elucidate the molecular mechanisms of biological systems have been revolutionized greatly with the use of high throughput and cost-effective techniques such as next generation sequencing (NGS). Application of NGS to microbial genomics is not just limited to predict the prevalence of microorganisms in food samples but also to elucidate the molecular basis of how microorganisms respond to different food-associated conditions, which in turn offers tremendous opportunities to predict and control the growth and survival of desirable or undesirable microorganisms in food. Concurrently, NGS has facilitated the development of new genome-assisted approaches for correlating genotype and phenotype. The aim of this review is to provide a snapshot of the various possibilities that these new technologies are opening up in area of food microbiology, focusing the discussion mainly on lactic acid bacteria and yeasts associated with fermented food. The contribution of NGS to a system level understanding of food microorganisms is also discussed.

The karyotype of the aphid Aphis sambuci has been studied by means of C-banding, fluorochrome staining and AgNO3 staining. Heterochromatic areas are confined exclusively to the X chromosomes. Interestingly, silver staining produces a clear-cut longitudinal differentiation of all chromosomes, which allows reliable identification of homologues.

Electrophoresis following digestion of Myzus persicae genomic DNA with HindIII showed the presence of a prominent band of approximately 200 bp whereas a faint electrophoretic band corresponding to DNA fragments of about 3000 bp was observed after digestion with ApaI. In situ digestion with restriction enzymes, followed by in situ nick translation, showed that ApaI targets are localized at the nucleolus organizer-bearing X telomeric region, whereas HindIII restriction sites are clustered in intercalary C-positive areas on the same X chromosome. Fluorescent in situ hybridization (FISH) carried out by using digoxygenin-labeled HindIII repeats as probe fully confirmed overlapping between the hybridization sites of this probe and the AT-rich intercalary heterochromatic bands on the X chromosome. These findings, together with published data, allow us to conclude that the M. persicae genome possesses three classes of C-positive heterochromatin: (i) a GC-rich argentophilic band located on one telomere of the X chromosome that contains ApaI targets; (ii) AT-rich intercalary bands located on the X chromosome containing clustered HindIII fragments; (iii) AT-rich telomeric bands, located on autosomes, consisting of HaeIII repeats. Molecular analysis has shown that the length of the HindIII repeat consensus sequence is 189 bp with an AT content of 67%. Southern blotting with HindIII monomers revealed a regular ladder of bands composed of multimers of basic length that are characteristic of satellite DNAs. The HindIII repeat displays other features typical of eukaryotic satellite arrays such as overlapping with heterochromatic bands and a high degree of sequence similarity among monomers (84%–94%). A similarity plot showed that sequences were particularly variable in the 50–100 bp region whereas they proved to be highly conservative in the first 50 bp, thus suggesting that this portion of the repeat might be functionally important.

Because of their compact genome, pufferfish (Tetraodontiformes) have been proposed as a model for the study of the vertebrate genome. The genome of pufferfish is peculiar as it has the structural complexity of the genomes of higher vertebrates, but has small introns and lacks large clusters of highly repetitive sequences. Despite such interest, information about the genetics of pufferfish is still scanty. To fill this gap, we have performed a cytogenetic analysis of the pufferfish, Tetraodon fluviatilis, which can be maintained in an aquarium for a long time and, unlike the pufferfish, Fugu rubripes, it is not difficult to obtain. Karyotype analysis shows that T. fluviatilis has 2n = 42 with two metacentric chromosomes, four submetacentrics, two subtelocentrics and 34 acrocentrics. C-banding, followed by DAPI staining, showed that heterochromatin is essentially AT-rich and is located at centromeres. Staining of the same metaphase plates with CMA3 showed the presence of four heterochromatic regions located on two pairs of submetacentric chromosomes. Silver staining and FISH with a 28S rDNA probe showed that these GC-rich regions are nucleolar organizing regions (NORs). Finally, regardless of the technique used, no difference in the chromosome complement was found between males and females.

. This brings to light the question of whether, during assisted reproduction techniques, sperm preparation is 4 To whom correspondence should be addressed able to isolate spermatozoa of 'real' quality or whether Human semen is heterogeneous in quality, not only between spermatozoa with undetectable anomalies are being used. males but also within a single ejaculate.

In parthenogenetic females of a clone of the aphid Megoura viciae (Homoptera, Aphididae), more than 50% of the cells show heteromorphism between homologous NORs which are located on one telomeric region of the two X chromosomes. Using different techniques, such as staining with the CG-specific fluorochrome chromomycin A3, silver staining and in-situ hybridization with an rDNA probe, we have shown that the observed heteromorphism is due to an unequal distribution of ribosomal genes between homologous NOR regions. The total number of rDNA genes per individual aphid remained constant. Moreover, the analysis of cells from single embroys has shown that the observed heteromorphism is not only intraclonal but also intraindividual. These data, together with the finding of X chromosomes connected by chromatin bridges between their NORs, allow us to suggest that mitotic unequal crossing over could be the main cause of NOR heteomorphism in this taxon.

Digestion of Mamestra brassicae DNA with DraI produced a prominent fragment of approximately 200 bp and a ladder of electrophoretic bands with molecular weights which are a multiple of 200 bp. Southern blotting revealed that this ladder is composed of DNA fragments that are multimers of the 200-bp DraI band suggesting that DraI isolated a satellite that has been called Mamestra brassicae satellite DNA 1 (MBSAT1). MBSAT1 is the first satellite DNA isolated in Lepidoptera. In-situ DraI digestion of chromosome spreads, together with fluorescent in-situ hybridization, showed that MBSAT1 sequences are clustered in heterochromatin of the sex chromosomes, Z and W. MBSAT1 was 234 bp long with an AT content of 60.7%. The curvature–propensity plot suggested a curvature in the MBSAT1 structure.

The induction of anti-microbial peptides against Gram positive and negative bacteria in the IZD-MB-0503 cell line from the lepidopteran Mamestra brassicae is demonstrated, while no anti-fungal activity is detected. The identification of a defensin-like molecule active against Gram positive bacteria is described for the first time in Lepidoptera. This molecule shows between 43% and 59% homology with group A defensins from other dipteran and hymenopteran species.

On the basis of a bioinformatic approach, we suggest that in invertebrates many ligands interact with a single, ancestral and generalized receptor driving ligand evolution. In vertebrates, on the other hand, the occurrence of gene/genome duplications induced the shift to a ligand-directed evolution of receptors.

The procorticotrophin-releasing hormone (proCRH) gene from the teleost Ameiurus nebulosus was cloned by direct and inverse PCR-based technologies and characterized. The structure of the proCRH gene shows the presence of four exons and three introns giving a total length of 1416 bp. Sequence similarity with the corresponding proCRH coding sequences in Tilapia mossambica and Homo sapiens is 97.7 and 78%, respectively. Western blot experiments performed with an anti-human CRH (1–41) antibody revealed the presence of an immunoreactive molecule with an approximate MW of 18 kDa, a value comparable to that of the putative catfish proCRH peptide. These data suggest that proCRH could be active in A. nebulosus without any cleavage. ProCRH immunoreactive molecules were found in the central nervous system (CNS) and were immunocytochemically detected in the head kidney and in the pancreatic gland. Western blot and immunocytochemical experiments showed an increase in proCRH expression in the CNS after 15 min but not after 120 min exposure to LPS. In contrast, the increased immunopositivity was detectable in the pancreas only after 120 min of treatment, but in the head kidney throughout the entire period of exposure. Our findings indicate that the CNS responds to the altered conditions for a shorter period of time than the peripheral organs, suggesting a hierarchical and time-regulated stress response. However, an independent response in the peripheral organs cannot be excluded in this scenario.

The cloning and characterisation of a procorticotrophin-releasing hormone (proCRH) and the related CRH fragment in the IZD-MB-0503 cell line from the leptidopteran Mamestra brassicae were performed. PCR amplification of the genomic DNA reveals a fragment of 276 bp, while inverse PCR shows the presence of a gene consisting of 1153 bp. The comparison of the insect genomic proCRH gene with proCRH cDNA obtained by RACE shows the presence of three introns. There was a 61% identity with the corresponding coding sequence in Tilapia mossambica, and a 65.2% identity with the human proCRH coding sequence.

In invertebrates and vertebrates, innate immunity is considered the first line of defense mechanism against non-self material. In vertebrates, cytokines play a critical role in innate immune signalling. To date, however, the existence of genes encoding for invertebrate helical cytokines has been anticipated, but never demonstrated. Here, we report the first structural and functional evidence of a gene encoding for a putative helical cytokine in Drosophila melanogaster. Functional experiments demonstrate that its expression, as well as that of the antimicrobial factors defensin and cecropin A1, is significantly increased after immune stimulation. These observations suggest the involvement of helical cytokines in the innate immune response of invertebrates.