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Objective The present study analyzed the influence of co-transferring embryos with high and low cloning efficiencies produced via somatic cell nuclear transfer (SCNT) on pregnancy outcomes in dogs. Methods Cloned dogs were produced by SCNT using donor cells derived from a Tibetan Mastiff (TM) and Toy Poodle (TP). The in vivo developmental capacity of cloned embryos was evaluated. The pregnancy and parturition rates were determined following single transfer of 284 fused oocytes into 21 surrogates and co-transfer of 47 fused oocytes into four surrogates. Results When cloned embryos produced using a single type of donor cell were transferred into surrogates, the pregnancy and live birth rates were significantly higher following transfer of embryos produced using TP donor cells than following transfer of embryos produced using TM donor cells. Next, pregnancy and live birth rates were compared following single and co-transfer of these cloned embryos. The pregnancy and live birth rates were similar upon co-transfer of embryos and single transfer of embryos produced using TP donor cells but were significantly lower upon single transfer of embryos produced using TM donor cells. Furthermore, the parturition rate for TM dogs and the percentage of these dogs that remained alive until weaning was significantly higher upon co-transfer than upon single transfer of embryos. However, there was no difference between the two embryo transfer methods for TP dogs. The mean birth weight of cloned TM dogs was significantly higher upon single transfer than upon co-transfer of embryos. However, the body weight of TM dogs did not significantly differ between the two embryo transfer methods after day 5. Conclusion For cloned embryos with a lower developmental competence, the parturition rate and percentage of dogs that remain alive until weaning are increased when they are co-transferred with cloned embryos with a greater developmental competence.

As the spread of antimicrobial resistance (AMR) genes becomes an increasing global threat, improved understanding of mobile genetic elements which contribute to the spread of antimicrobial resistance genes, becomes more critical. We created transconjugants from the mating of three chromosomally isogenic Klebsiella pneumoniae carbapenemase (bla KPC) positive Citrobacter freundii isolates with a laboratory strain of Escherichia coli and evaluated the movement of small cryptic plasmids (SCPs), p3223 and p1916, when larger bla KPC -plasmids were transferred. In all of the 143 transconjugants, multiple plasmids, both large and small, transferred with each mating. When two bla KPC- plasmids were present in the host, frequently (87%; 98/113) both would be transferred during mating. p3223 is found in a wide range of bacterial hosts that harbor AMR genes; p1916 has been identified in only a limited number of publicly available sequences to date. From our evaluation, there is still much to learn about SCPs, and the high rate of co-transfer of multiple plasmids from real-world carbapenemase-producing Enterobacteriales. • There is frequent in vitro co-transfer of small plasmids with bla KPC -plasmids from hospital acquired C. freundii isolates. • During in vitro conjugation, multiple bla KPC -plasmids within a C. freundii host get transferred to an E. coli recipient. • There can be a plethora of plasmid diversity within bacterial isolates that have isogenic chromosomes within an institution. • p3223 has a host range that includes many species of Gammaproteobacteria but little is known about its function. [ABSTRACT FROM AUTHOR]

The present study was conducted to evaluate the possibility of using in vitro-produced parthenogenetic ( PA) embryos for co-transfer with morulae that had been collected in vivo and cryopreserved. The proportion of PA blastocysts (20.5%) was higher than that of their in vitro fertilization ( IVF) counterparts (16.6%). Although there were no differences in morphology or diameter between the two groups, the number of cells in early PA blastocysts after in vitro culture for 6 days was lower than for IVF blastocysts (25.7 and 30.4 cells, respectively), and the number in recovered PA blastocysts was also smaller than that in recovered IVF blastocysts (37.4 and 50.2 cells, respectively). When 10 morulae warmed after vitrification were co-transferred with 10 PA blastocysts (total 20 embryos) to the uterus of five recipients, the rates of pregnancy and farrowing did not differ, but the average period until spontaneous abortion tended to be longer relative to the control (when 20 morulae were transferred). These data suggest that in vitro-produced PA embryos offer the possibility of assisted pregnancy for cryopreserved embryos; further experiments will be needed to confirm the beneficial effect of this approach on piglet production. [ABSTRACT FROM AUTHOR]

Background: Genomic uptake of DNA by prokaryotes often encompasses more than a single gene. In many cases, several horizontally transferred genes may be acquired together. Accordingly, we expect that horizontally transferred genes cluster spatially in the genome more often than expected if transfers were independent. Further, genes that depend on each other functionally may be unlikely to have beneficial fitness effects when taken up individually by a foreign genome. Hence, we also expect the co-acquisition of functionally related genes, resulting in the clustering of horizontally transferred genes in functional networks. Results: Analysing spatial and metabolic clustering of recent horizontal (or lateral) gene transfers among 21 γ-proteobacteria, we confirm both predictions. When comparing two datasets of predicted transfers that differ in their expected false-positive rate, we find that the more stringent dataset shows a stronger enrichment of clustered pairs. Conclusions: The enrichment of interdependent metabolic genes among predicted transfers supports a biologically significant role of horizontally transferred genes in metabolic adaptation. Our results further suggest that spatial and metabolic clustering may be used as a benchmark for methods that predict recent horizontal gene transfers. Reviewers: This article was reviewed by Peter Gogarten in collaboration with Luiz Thiberio Rangel, and by Yuri Wolf. [ABSTRACT FROM AUTHOR]

Lambdoid (or Lambda-like) phages are a group of related temperate phages that can infect Escherichia coli and other gut bacteria. A key characteristic of these phages is their mosaic genome structure, which served as the basis for the 'modular genome hypothesis'. Accordingly, lambdoid phages evolve by transferring genomic regions, each of which constitutes a functional unit. Nevertheless, it is unknown which genes are preferentially transferred together and what drives such co-transfer events. Here we aim to characterize genome modularity by studying co-transfer of genes among 95 distantly related lambdoid (pro-)phages. Based on gene content, we observed that the genomes cluster into 12 groups, which are characterized by a highly similar gene content within the groups and highly divergent gene content across groups. Highly similar proteins can occur in genomes of different groups, indicating that they have been transferred. About 26 % of homologous protein clusters in the four known operons (i.e. the early left, early right, immunity and late operon) engage in gene transfer, which affects all operons to a similar extent. We identified pairs of genes that are frequently co-transferred and observed that these pairs tend to be near one another on the genome. We find that frequently co-transferred genes are involved in related functions and highlight interesting examples involving structural proteins, the cI repressor and Cro regulator, proteins interacting with DNA, and membrane-interacting proteins. We conclude that epistatic effects, where the functioning of one protein depends on the presence of another, play an important role in the evolution of the modular structure of these genomes.

Objective: The present study analyzed the influence of co-transferring embryos with high and low cloning efficiencies produced via somatic cell nuclear transfer (SCNT) on pregnancy outcomes in dogs., Methods: Cloned dogs were produced by SCNT using donor cells derived from a Tibetan Mastiff (TM) and Toy Poodle (TP). The in vivo developmental capacity of cloned embryos was evaluated. The pregnancy and parturition rates were determined following single transfer of 284 fused oocytes into 21 surrogates and co-transfer of 47 fused oocytes into four surrogates., Results: When cloned embryos produced using a single type of donor cell were transferred into surrogates, the pregnancy and live birth rates were significantly higher following transfer of embryos produced using TP donor cells than following transfer of embryos produced using TM donor cells. Next, pregnancy and live birth rates were compared following single and co-transfer of these cloned embryos. The pregnancy and live birth rates were similar upon co-transfer of embryos and single transfer of embryos produced using TP donor cells but were significantly lower upon single transfer of embryos produced using TM donor cells. Furthermore, the parturition rate for TM dogs and the percentage of these dogs that remained alive until weaning was significantly higher upon co-transfer than upon single transfer of embryos. However, there was no difference between the two embryo transfer methods for TP dogs. The mean birth weight of cloned TM dogs was significantly higher upon single transfer than upon co-transfer of embryos. However, the body weight of TM dogs did not significantly differ between the two embryo transfer methods after day 5., Conclusion: For cloned embryos with a lower developmental competence, the parturition rate and percentage of dogs that remain alive until weaning are increased when they are co-transferred with cloned embryos with a greater developmental competence.

Abstract: The aim of this study was to determine the complete sequence of the multi-resistance plasmid pV7037 to gain insight into the structure and organization of this plasmid. Of the four XbaI clones of pV7037, one clone of 17,577bp has already been sequenced and shown to carry a multi-resistance gene cluster. The remaining three clones of approximately 12.5, 6.5 and 4.5kb were sequenced, the entire plasmid sequence correctly assembled and investigated for reading frames. In addition, two reading frames one coding for an ABC transporter and the other coding for an rRNA methylase were cloned and expressed in a S. aureus host to see whether they confer antimicrobial resistance properties. Plasmid pV7037 proved to be 40,971bp in size. Besides the previously determined resistance gene cluster, it carried a functionally active tet(L) gene for tetracycline resistance, a complete cadDX operon for cadmium resistance and also a variant of the β-lactamase transposon Tn552. Two single bp deletions, which resulted in frame shifts, functionally deleted the genes for the BlaZ β-lactamase and the signal transducer protein BlaR1 in this Tn552 variant of pV7037. Plasmid pV7037 seems to be composed of various parts previously known from plasmids and transposons of staphylococci and other Gram-positive bacteria. However, there are also parts of the plasmid which do not show any homology to so far known sequences deposited in the databases. The novel ABC transporter and rRNA methylase genes identified on pV7037 do not seem to play a role in antimicrobial resistance. The co-location of numerous antimicrobial resistance genes bears the risk of co-transfer and co-selection of resistance genes, but also persistence of resistance genes even if no direct selective pressure by the use of the respective antimicrobial agents is applied. [Copyright &y& Elsevier]

In this study, the additive effects of a late embryogenesis abundant ( LEA) gene and a basic leucine zipper ( bZIP) gene on salt and osmotic stress in Tamarix plants were analyzed. The constructs containing one or both of the LEA and bZIP genes were transformed into tobacco. Northern blot analysis showed the genes were overexpressed under the control of the CaMV 35S promoter in both dual and single gene-transgenic tobacco lines. The effects of salt and osmotic stress in transgenic tobacco plant were investigated. Following exposure to NaCl, mannitol, and PEG6000 stress, dual gene-transgenic lines showed higher seed generation and growth rates than single gene-transgenic lines and the wild-type. In response to NaCl stress, the dual gene-transgenic lines showed lower malondialdehyde and higher leaf chlorophyll content than single gene-transgenic lines and the wild-type. These results suggested that the co-expression of LEA and bZIP resulted in an additive enhancement of stress tolerance in dual gene-transgenic tobacco. [ABSTRACT FROM AUTHOR]

Abstract: It has long been known that several embryos are needed to establish and maintain pregnancy during early gestation in pigs. In this study, we assessed whether co-transfer of parthenogenotes with a single embryo was sufficient to maintain development of the embryo. Embryos were recovered at morula and early blastocyst stages from gilts that had been artificially inseminated. Parthenogenotes were produced from oocytes matured in vitro, activated by electrical stimulation, and then cultured for 110h. Those that had developed to morula- or blastocyst-like stages at 110h post-activation were transferred to recipient pigs either with or without morula or blastocyst stage embryos. Parthenogenotes that were transferred to recipients in the absence of embryos were viable up to 30 days post-activation and formed limb-buds; at 40 days, however, all were dead or degenerate. Among pigs that received both parthenogenotes and a single embryo, seven of nine (77.8%) delivered a normal piglet at full-term. This study therefore demonstrates that parthenogenotes can be used in place of embryos to establish pregnancy and promote development of a single co-transferred embryo. This method may be applied to rescue high value porcine embryos that are difficult to produce, such as transgenic cloned embryos, or for embryos frozen as a genetic resource. [Copyright &y& Elsevier]

Abstract: Resistance to various classes of antimicrobial agents has been encountered in many bacteria of medical and veterinary relevance. Particular attention has been paid to zoonotic bacteria such as Salmonella. Over the years, various studies have reported the presence of genes and mutations conferring resistance to antimicrobial agents in Salmonella isolates. This review is intended to provide an update on what is currently known about the genetic basis of antimicrobial resistance in Salmonella. [Copyright &y& Elsevier]

A new plasmid series has been created for Agrobacterium-mediated plant transformation. The pBECKS2000 series of binary vectors exploits the Cre/ loxP site-specific recombinase system to facilitate the construction of complex T-DNA vectors. The new plasmids enable the rapid generation of T-DNA vectors in which multiple genes are linked, without relying on the availability of purpose-built cassette systems or demanding complex, and therefore inefficient, ligation reactions. The vectors incorporate facilities for the removal of transformation markers from transgenic plants, while still permitting simple in vitro manipulations of the T-DNA vectors. A `shuttle' or intermediate plasmid approach has been employed. This permits independent ligation strategies to be used for two gene sets. The intermediate plasmid sequence is incorporated into the binary vector through a plasmid co-integration reaction which is mediated by the Cre/ loxP site-specific recombinase system. This reaction is carried out within Agrobacterium cells. Recombinant clones, carrying the co-integrative binary plasmid form, are selected directly using the antibiotic resistance marker carried on the intermediate plasmid. This strategy facilitates production of co-integrative T-DNA binary vector forms which are appropriate for either (1) transfer to and integration within the plant genome of target and marker genes as a single T-DNA unit; (2) transfer and integration of target and marker genes as a single T-DNA unit but with a Cre/ loxP facility for site-specific excision of marker genes from the plant genome; or (3) co-transfer of target and marker genes as two independent T-DNAs within a single-strain Agrobacterium system, providing the potential for segregational loss of marker genes. [ABSTRACT FROM AUTHOR]

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Carbonic anhydrase activity was found and characterized in isolated homogeneous preparations of microvilli and basal infoldings of the rat renal tubular cells. This membrane-bound activity was about 3% of the total carbonic anhydrase activity of the renal cortex. Compared to the renal cytoplasmic soluble form, the membrane-bound enzymes had the same K for CO and were equally resistant to inhibition by chloride. However, they were less inhibited by different sulfonamides. The implication of the existence of membrane-bound enzymes in the renal tubular cells is discussed. [ABSTRACT FROM AUTHOR]