Structure of plant–Hymenoptera networks in two coastal shrub sites in Mexico

The study of interaction networks between plants and pollinators allows us to explore interaction patterns at the community level, detect changes in visit frequency and evaluate the nestedness of the networks. The latter allows rare plant species to be visited by more abundant species of pollinators, potentially allowing community diversity to be maintained, and this approach makes it possible to discern the rewiring (changes in connections) of species when their preferred resource is not available. In this study, the topology, species identity and rewiring were compared between two contrasting sites, one within a conservation area and the other subjected to continuous disturbance. The networks of both sites were significantly nested and shared a high number of common species of both plants and pollinators. However, the sites differed notably in the number of exclusive interactions, suggesting a high percentage of interaction rewiring. The introduced bee species, Apis mellifera, was the most frequent species at both sites and also the most connected in terms of the number of its interactions. This is explained by its generalist foraging characteristics that allow it to form part of the networks’ core group. In general, our results underscore the importance of knowing the identity of the participating species when studying networks, and how connections change between them, as well as the potential effect of habitat destruction and the role of invasive species in the rearrangement of the interactions; all factors that can exert an influence on the functioning of plant–pollinator networks.     

Prevalence and phylogenetic relationship of two β-carbonic anhydrases in affiliates of Enterobacteriaceae

Enterobacteriaceae, one of the major families of microorganisms that inhabit the soil and gut, internally regulate constant fluctuations in soil and gut pH by buffering these changes through the presence of carbonic anhydrase (CA). In our study, we prove the prevalence of β-CA, derived from the can gene, in members of Enterobacteriaceae by using a combination of experimental and bioinformatics approaches. Enzyme purification and western blot analysis revealed the presence of β-CA in Enterobacter sp. RS1. Genetic studies confirmed the presence of β-CA in both Enterobacter sp. RS1 and Citrobacter freundii SW3. Our analysis of the divergence of cynT and can genes among harboring members indicated that the can gene was more prominent in Enterobacteriaceae than cynT. Sequence analysis of the can gene revealed a >25 % similarity among all sequences and a >50 % similarity among sequences from the Enterobacteriaceae family. The β-CA from C. freundii SW3 and Enterobacter sp. RS1, isolated from soil and used in this study, possessed a high similarity with the can gene. The close association among Enterobacteriaceae genera usually found in the soil and gut and the sequence similarity of β-CA in the different genera of Enterobacteriaceae suggest the importance of the can gene in oscillating environmental conditions.     

First detection of OKP-A β-lactamase in two Serratia marcescens isolates in China

Two strains of Enterobacteriaceae producing prodigiosin were isolated from meat in the Sichuan province of China in 2010. The strains were identified by Vitek system, 16S rDNA, rpoB, pfs and luxS genes. Minimum inhibitory concentrations were determined using the broth microdilution method. The two strains were screened for the presence of β-lactamase genes (blaTEM, blaSHV, blaOKP, and blaCTX-M genes). Based on PCR amplification and 16S rDNA sequencing the analysed strains were identified as Serratia marcescens. In addition, morphological and biochemical identification showed that the two stains were definitely S. marcesens. Antimicrobial susceptibility test showed that both strains were resistant to ampicillin and first-generation cephalosporins while being susceptible to cefotaxime, ceftiofur, ceftriaxone, imipenem and aztreonam. It was found that blaOKP had been identified first from the two S. marcescens strains, ch1 and ch2. The isolates were closely related as shown by pulsed-field gel electrophoresis (PFGE). The narrow-spectrum OKP-A β-lactamase gene blaOKP-A-13 was found to be chromosomally located in S. marcescens. The isolates produced a β-lactamase with a pI of approximately 8.2, which corresponds to the OKPA family. Findings indicate that OKP enzymes are not Klebsiella pneumoniae-specific chromosomal ?-lactamases, and the first isolation of S. marcescens producing OKP-A ?-lactamase suggests that the blaOKP gene may be disseminated between different species.     

Evidence for the occurrence of two forms of β-lipotropin in rat pituitary

A peptide isolated from porcine gut according to its glucagon-like activity in liver (bioactive enteroglucagon) has been characterized immunologically, biologically and chemically: its potency relative to pancreatic glucagon in interacting with an antiglucagon antibody, hepatic glucagon-binding sites and hepatic adenylate cyclase was ～100%, 20% and 10%, respectively. In contrast, it is ～20-times more potent than glucagon in oxyntic glands, justifying the term ‘oxyntomodulin’. Chemically, it consists in the 29 amino acid-peptide glucagon elongated at its C-terminal end by the octapeptide Lys—Arg—Asn—Lys—Asn—Asn—Ile &;—Ala; accordingly, it is called ‘glucagon-37’     

Localization of adhesive proteins in two newly subdivided zones in electron-lucent matrix of human platelet α-granules

Summary Platelet -granules have been reported to consist of two zones, nucleoid and electron-lucent matrix, with different densities under electron microscopy. When washed human platelets were prepared by a rapid freeze-substitution method using liquid helium, we found that the electron-lucent matrix could be further subclassified into two zones having different densities: the intermediate and the light zones. The light zone was located at the periphery opposite the most dense nucleoid and contained several tubular structures with diameters of about 20 nm. The intermediate zone often laid between the nucleoid and light zone. By careful inspection, intermediate and light zones could even be identified in the platelets embedded in Lowicryl K4M, which where then used to localize several adhesive proteins in these two zone by immunocytochemical studies using the respective polyclonal antibodies. Fibrinogen, thrombospondin, and fibronectin were detected only in the intermediate zone. In contrast, von Willebrant factor (vWF) was localized only in the light zone, suggesting an association between vWF and the tubular structures in the light zone. In the nucleoid, none of these adhesive proteins were detected. Glycoprotein IIb/IIIa, a receptor for these adhesive proteins on the platelet surface, was detected not only on the outer surface of the cell membranes but also on the inner surface of the -granule membrane. These data indicate that two zones with different densities in electron-lucent matrix and functions exist in the platelet -granules.     

Molecular characterization of two types of 22 kilodalton α-zein genes in a gene cluster in maize

Summary Five genes of the -zein subfamily four (SF4) are located in a 56 kb genomic region of the maize inbred line W22. Their nucleotide and deduced amino acid sequences have been determined. The sequences define two types of -zein SF4 genes — type 1 (T1) and type 2 (T2). The single T1 -zein SF4 gene codes for an -zein protein with a M_r of about 22 000. This is the first -zein SF4 gene sequenced that contains no early in-frame stop codons in its coding sequence. The four T2 -zein SF4 genes in this cluster contain one or two early in-frame stop codons. In addition, our T1 and T2 genes differ markedly in the base sequences of their distal 5 non-translated flanking regions. The nucleotide and the deduced amino acid sequences of these two types of -zein SF4 genes are similar ( > 90 %) to one another and to all known -zein SF4 genes and cDNAs. Of the known W22 -zein SF4 genes, only one in six does not contain an early in-frame stop codon. If the number of -zein SF4 genes is 15–20, then we estimate that only about 4 of the W22 -zein SF4 genes are without in-frame early stop codons.     

Observation of two quenchers of chlorophyll fluorescence in chloroplasts at −196°C

Fluorescence induction at ?196°C has been monitored in chloroplasts rapidly frozen after poising at different redox potentials at room temperature. It was found that, as at room temperature, the initial level of fluorescence observed upon shutter opening (F_o), relative to the final level observed after 10 seconds of illumination (F_m) increased as the redox potential of the chloroplasts was lowered. Redox titration revealed the presence of two quenching components with E_m,7.8 at ?70 mV and ?275 mV accounting for approx. 75% and 25% of the variable fluorescence (F_v). Parallel observation of fluorescence yield at room temperature similarly gave two components, with E_m,7.8 at ?95 mV and ?290 mV, also accounting for approx. 75% and 25%. Simultaneous measurement of fluorescence emission at ?196°C at 695 nm and 735 nm indicated that both emissions are quenched by the same redox components.     

Changes in expression of two endogenous β-galactoside-binding isolectins in the dermis of chick embryonic skin during development in ovo and in vitro

In order to elucidate the roles of metal-independent animal lectins, we systematically investigated changes in expression of 2 kinds of -galactoside-binding isolectins (MW 14 and 16 kDa) in the dermis of chick embryonic tarsometatarsal skin during the course of development. These lectins were immunohistochemically located at different stages of development both in ovo and in vitro by light and electron microscopy. Light-microscopic observation showed that while positive staining for the 14-kDa lectin was weak at days 8 and 10 it became intense after day 13. In contrast, staining for the 16-kDa lectin was intense at days 8, 10, and 13, but it became weak after day 17 when keratinization of the epidermis was completed. Immuno-electron-microscopic observation revealed that both the 14 and 16-kDa lectins were located on the basement membrane, in the extracellular matrix, and in both the cytoplasm and the nucleus of dermal fibroblasts. Distribution of the 2 isolectins was also examined in cultured skin explants in vitro. The results were almost the same as those obtained in ovo when the skin explant was keratinized in the presence of hydrocortisone. However, in the skin explant where keratinization was prevented and mucous metaplasia was induced by the addition of vitamin A, the distribution of the 14-kDa lectin in the epidermis was significantly affected. These results indicate that (1) the expression of the 2 isolectins is differently regulated in both the dermis and epidermis, (2) the 16-kDa lectin is involved in the early stage of the formation of the dermis and the basement membrane and is replaced by the 14-kDa lectin as keratinization of the epidermis occurs, and (3) the expression of the 2 isolectins in the dermis is not significantly affected by the induction of mucous metaplasia, in contrast to their drastic changes in the epidermis.     

Are olfactory cues involved in nest recognition in two social species of estrildid finches?

Reliably recognizing their own nest provides parents with a necessary skill to invest time and resources efficiently in raising their offspring and thereby maximising their own reproductive success. Studies investigating nest recognition in adult birds have focused mainly on visual cues of the nest or the nest site and acoustic cues of the nestlings. To determine whether adult songbirds also use olfaction for nest recognition, we investigated the use of olfactory nest cues for two estrildid finch species, zebra finches (Taeniopygia guttata) and Bengalese finches (Lonchura striata var. domestica) during the nestling and fledgling phase of their offspring. We found similar behavioural responses to nest odours in both songbird species. Females preferred the odour of their own nest over a control and avoided the foreign conspecific nest scent over a control during the nestling phase of their offspring, but when given the own odour and the foreign conspecific odour simultaneously we did not find a preference for the own nest odour. Males of both species did not show any preferences at all. The behavioural reaction to any nest odour decreased after fledging of the offspring. Our results show that only females show a behavioural response to olfactory nest cues, indicating that the use of olfactory cues for nest recognition seems to be sex-specific and dependent on the developmental stage of the offspring. Although estrildid finches are known to use visual and acoustic cues for nest recognition, the similar behavioural pattern of both species indicates that at least females gain additional information by olfactory nest cues during the nestling phase of their offspring. Thus olfactory cues might be important in general, even in situations in which visual and acoustic cues are known to be sufficient.     

A novel mutation in exon 17 of the β-subunit of rod phosphodiesterase in two RP sisters of a consanguineous family

We report the molecular analysis of the subunit of the rod phosphodiesterase (PDEB) gene in a consanguineous autosomal recessive retinitis pigmentosa family that shows homozygosity for polymorphisms in the genomic region comprising this gene, and positive linkage between a PDEB marker and the diesease. The two affected sisters are homozygous for a T to G transversion in codon 699 of the PDEB gene, leading to the substitution of a leucine by an arginine residue. This change, enclosed in the catalytic domain of the PDEB, could result in a modification of the protein structure preventing the physiological hydrolysis of cGMP.     

Differential RNA accumulation of two β-tubulin genes in arbuscular mycorrhizal fungi

RNA was isolated from spores of different arbuscular mycorrhizal (AM) fungi and used for RT-PCR with degenerate primers for beta-tubulin genes. PCR products were cloned and the sequence of several clones was analysed for each fragment. Comparison of sequences identified two loci for beta-tubulin genes with different GC content and codon usage. Btub1 sequences were most similar to beta-tubulin genes from the Oomycota, while Btub2 sequences showed highest similarity to sequences from the Zygomycota. RT-PCR experiments were carried out to monitor RNA accumulation patterns of Btub1 and Btub2 in asymbiotic germinating spores and in symbiotic extraradical hyphae of three different AM fungi. This indicated that Btub1 is constitutively expressed in Gigaspora rosea, but down-regulated during symbiosis in Glomus mosseae and Glomus intraradices. In contrast, Btub2 showed constitutive expression in the two Glomus species, but down-regulation in G. rosea. Further analysis of different fungi indicated that Btub2 primers could be used to specifically monitor RNA accumulation of AM fungi in environmental samples.     

Construction of two recombination yeast two-hybrid vectors by in vitro recombination

Recombination-based restrictionless, ligation-independent cloning has been proven to be advantageous over restriction digestion and ligation cloning. To utilize the recombination cloning and previously constructed two-hybrid cDNA libraries, a new Gateway yeast two-hybrid bait vector, pEZY202, and a new prey vector, pEZY45, were constructed. The two-hybrid vectors were generated by in vitro recombination using a protocol that can be easily adapted for the conversion of other existing vectors. The new vectors were used to assay the interaction between the WW domain of PQBP1 (PQBPww) and the WW domain binding protein WBP11. Both PQBPww and WBP11 were cloned into a Gateway donor vector by in vitro recombination. They were then subcloned into pEZY45 and pEZY202, respectively, by in vitro recombination. The binding between PQBPww and WBP11 was reported in a two-hybrid experiment using the new vectors. The results of testing the new vectors in combination with the original vectors indicated that the new bait vector could be used to screen cDNA libraries that are constructed using the original prey vectors.     

Analysis of chaotic oscillations induced in two coupled Wilson–Cowan models

Although it is known that two coupled Wilson–Cowan models with reciprocal connections induce aperiodic oscillations, little attention has been paid to the dynamical mechanism for such oscillations so far. In this study, we aim to elucidate the fundamental mechanism to induce the aperiodic oscillations in the coupled model. First, aperiodic oscillations observed are investigated for the case when the connections are unidirectional and when the input signal is a periodic oscillation. By the phase portrait analysis, we determine that the aperiodic oscillations are caused by periodically forced state transitions between a stable equilibrium and a stable limit cycle attractors around the saddle-node and saddle separatrix loop bifurcation points. It is revealed that the dynamical mechanism where the state crosses over the saddle-node and saddle separatrix loop bifurcations significantly contributes to the occurrence of chaotic oscillations forced by a periodic input. In addition, this mechanism can also give rise to chaotic oscillations in reciprocally connected Wilson–Cowan models. These results suggest that the dynamic attractor transition underlies chaotic behaviors in two coupled Wilson–Cowan oscillators.     

Detection and partial characterization of two antigenically distinct β-amylases in developing kernels of wheat

A -amylase (EC 3.2.1.2) was identified in the outer pericarp (P) of developing seeds of wheat (Triticum aestivum L.) and compared with the well known -amylase which is synthesized during seed development in the starchy endosperm (E). The enzyme P already exists in the tissues before anthesis and vanishes at the time when E starts to accumulate. The isoelectric-focusing patterns of P and E are very similar. The relative molecular weight (M_r) of P is slightly higher than that of E (66 and 64.5 kDa, respectively). Both P and E exhibit common epitopes in addition to epitopes specific for each of them. The two enzymes were identified in small amounts in the green tissues of the developing seeds (inner pericarp and testa). No antigenic difference was detected between P and the -amylases of roots and leaves.Abbreviations P pericarp -amylase - E endosperm -amylase - IS1 anti--amylase immune serum - IS2 anti- and anti- amylase immune serum - IS3 anti- amylase immune serum - IEF isoelectric focusing - IgG immunoglobulin G The authors thank Dr. P. Ziegler (Universität Bayreuth, FRG) for stimulating discussion and for useful suggestions during the writing of the text. The authors thank Miss C. Mayer for her skillful technical assistance.     

Nucleotide sequence of two 3′-termini of rRNA in the sea urchin Lytechinus variegatus

The 3′-terminus of 26 S rRNA from the sea urchin Lytechinus variegatus has been determined by oligonucleotide fingerprinting, S1 nuclease mapping and terminal nucleotide analysis. There are two species of 26 S rRNA of approximately equal abundance, one 19 nucleotides longer than the other.     

Colocalization of pinopsin with two types of G-protein α-subunits in the chicken pineal gland

Pinopsin is a photoreceptive molecule present in the outer segments of chicken pinealocytes. In this paper, the localization of alpha-subunits of G-proteins, rod transducin (Gt1) and Gq/11, was examined by immunoelectron microscopy to investigate whether these G-proteins colocalize with pinopsin in the outer segments. Ultrathin sections of the chicken pineal gland were double-immunolabeled with antibodies to pinopsin and either Gt1alpha or Gq/11alpha. As shown previously, the outer segments around the follicular lumen exhibited divergent morphology with ciliary, bulbous, or lamellate shapes, and most of them displayed pinopsin immunoreactivity. The majority (>90%) of pinopsin-immunopositive outer segments were labeled by anti-Gt1alpha and/or anti-Gq/11alpha antibodies. Application of double-immunolabeling to serial sections demonstrated that a large number of the pinopsin-immunopositive outer segments contained both Gt1alpha and Gq/11alpha immunoreactivities. These results suggest that Gt1alpha and Gq/11alpha are functionally coupled with light-activated pinopsin within a single outer segment.