Occurrence and diversity of tetracycline resistance genes in lagoons and groundwater underlying two swine production facilities

In this study, we used PCR typing methods to assess the presence of tetracycline resistance determinants conferring ribosomal protection in waste lagoons and in groundwater underlying two swine farms. All eight classes of genes encoding this mechanism of resistance [tet(O), tet(Q), tet(W), tet(M), tetB(P), tet(S), tet(T), and otrA] were found in total DNA extracted from water of two lagoons. These determinants were found to be seeping into the underlying groundwater and could be detected as far as 250 m downstream from the lagoons. The identities and origin of these genes in groundwater were confirmed by PCR-denaturing gradient gel electrophoresis and sequence analyses. Tetracycline-resistant bacterial isolates from groundwater harbored the tet(M) gene, which was not predominant in the environmental samples and was identical to tet(M) from the lagoons. The presence of this gene in some typical soil inhabitants suggests that the vector of antibiotic resistance gene dissemination is not limited to strains of gastrointestinal origin carrying the gene but can be mobilized into the indigenous soil microbiota. This study demonstrated that tet genes occur in the environment as a direct result of agriculture and suggested that groundwater may be a potential source of antibiotic resistance in the food chain.     

Regulation of CB1 cannabinoid receptor internalization by a promiscuous phosphorylation-dependent mechanism

Agonists stimulate cannabinoid 1 receptor (CB₁R) internalization. Previous work suggests that the extreme carboxy-terminus of the receptor regulates this internalization – likely through the phosphorylation of serines and threonines clustered within this region. While truncation of the carboxy-terminus (V460Z CB₁) and consequent removal of these putative phosphorylation sites prevents endocytosis in AtT20 cells, the residues necessary for CB₁R internalization remain elusive. To determine the structural requirements for internalization, we evaluated endocytosis of carboxy-terminal mutant CB₁Rs stably expressed in HEK293 cells. In contrast to AtT20 cells, V460Z CB₁R expressed in HEK293 cells internalized to the same extent and with similar kinetics as the wild-type receptor. However, mutation of serine and/or threonine residues within the extreme carboxy-terminal attenuated internalization when these receptors were expressed in HEK293 cells. These results establish that the extreme carboxy-terminal phosphorylation sites are not required for internalization of truncated receptors, but are required for internalization of full-length receptors in HEK293 cells. Analysis of β-arrestin-2 recruitment to mutant CB₁R suggests that putative carboxy-terminal phosphorylation sites mediate β-arrestin-2 translocation. This study indicates that the local cellular environment affects the structural determinants of CB₁R internalization. Additionally, phosphorylation likely regulates the internalization of (full-length) CB₁Rs.     

Solution properties of sodium alginate

Sodium alginate fractions derived from three different sources—Laminaria hyperboria (75% guluronate), Fucus vesicularus (95% mannuronate), and Azotobacter vinelandii (85% mannuronate)—were investigated in aqueous solution over a wide range of ionic strength and pH using the techniques of light scattering, viscometry, and osmometry. Light-scattering data extrapolated to infinite ionic strength yielded b₀ = 4.7 ± 0.3 and 3.0 ± 0.2 nm for the unperturbed effective bond lengths of the guluronate- and mannuronate-rich samples, respectively. These values are in the same ratio as predicted by conformational analysis, although lower by a factor of 0.7, probably due, in part at least, to the fact that measurements cannot be made on pure homopolymers. A comparison of the light-scattering and the viscosity data indicated that Φ in the Flory-Fox equation is lower than for more flexible polymers and increases with molecular weight, probably due to decreasing hydrodynamic permeability. Mark-Houwink exponents obtained from data extrapolated to infinite ionic strength were found to be considerably greater than 0.5, and we attribute this entirely to a variation in Φ. Comparison of the results obtained for the two mannuronate-rich samples indicated that the value of Φ and its variation with molecular weight can, in the case of alginates, differ markedly for chains, which, although having chemical differences, have similar chain statistics.     

Cryptic species in the cosmopolitan Bugula neritina complex (Bryozoa,Cheilostomata)

Previous analyses of the mitochondrial gene cytochrome c oxidase subunit 1 (COI) and γ‐proteobacterial endosymbiont diversity have suggested that the marine bryozoan Bugula neritina is a complex of three cryptic species, namely Types S, D and N. Types D and N were previously reported to have restricted distributions along California (western USA) and Delaware and Connecticut (eastern USA), respectively, whereas Type S is considered widespread in tropical, subtropical and temperate regions due to anthropogenic transport. Here, Bayesian species delimitation analysis of a data set composed of two mitochondrial (COI and large ribosomal RNA subunit [16S]) and two nuclear genes (dynein light chain roadblock type‐2 protein [DYN] and voltage‐dependent anion‐selective channel protein [VDAC]) demonstrated that Types S, D and N correspond to three biological species. This finding was significantly supported, in spite of the combinations of priors applied for ancestral population size and root age. Furthermore, COI sequences were used to assess the introduction patterns of the cosmopolitan Type S species. Two COI haplotypes of Type S (S1a and S1d) were found occurring at a global scale. Mantel tests showed correlation between these haplotypes and local sea surface temperature tolerance. Accordingly, the distributions of Type S haplotypes may reflect intraspecific temperature tolerance variation, in addition to the role of introduction vectors. Finally, we show that the Type N may also have been introduced widely, as this species was found for the first time in Central California and north‐eastern Australia.