A mutation in the fibroblast growth factor 14 gene is associated with autosomal dominant cerebellar ataxia [corrected

Hereditary spinocerebellar ataxias (SCAs) are a clinically and genetically heterogeneous group of neurodegenerative disorders for which >/=14 different genetic loci have been identified. In some SCA types, expanded tri- or pentanucleotide repeats have been identified, and the length of these expansions correlates with the age at onset and with the severity of the clinical phenotype. In several other SCA types, no genetic defect has yet been identified. We describe a large, three-generation family with early-onset tremor, dyskinesia, and slowly progressive cerebellar ataxia, not associated with any of the known SCA loci, and a mutation in the fibroblast growth factor 14 (FGF14) gene on chromosome 13q34. Our observations are in accordance with the occurrence of ataxia and paroxysmal dyskinesia in Fgf14-knockout mice. As indicated by protein modeling, the amino acid change from phenylalanine to serine at position 145 is predicted to reduce the stability of the protein. The present FGF14 mutation represents a novel gene defect involved in the neurodegeneration of cerebellum and basal ganglia.     

A ribosomal RNA gene intergenic spacer based PCR and DGGE fingerprinting method for the analysis of specific rhizobial communities in soil

A direct molecular method for assessing the diversity of specific populations of rhizobia in soil, based on nested PCR amplification of 16S-23S ribosomal RNA gene (rDNA) intergenic spacer (IGS) sequences, was developed. Initial generic amplification of bacterial rDNA IGS sequences from soil DNA was followed by specific amplification of (1) sequences affiliated with Rhizobium leguminosarum "sensu lato" and (2) R. tropici. Using analysis of the amplified sequences in clone libraries obtained on the basis of soil DNA, this two-sided method was shown to be very specific for rhizobial subpopulations in soil. It was then further validated as a direct fingerprinting tool of the target rhizobia based on denaturing gradient gel electrophoresis (DGGE). The PCR-DGGE approach was applied to soils from fields in Brazil cultivated with common bean (Phaseolus vulgaris) under conventional or no-tillage practices. The community fingerprints obtained allowed the direct analysis of the respective rhizobial community structures in soil samples from the two contrasting agricultural practices. Data obtained with both primer sets revealed clustering of the community structures of the target rhizobial types along treatment. Moreover, the DGGE profiles obtained with the R. tropici primer set indicated that the abundance and diversity of these organisms were favoured under NT practices. These results suggest that the R. leguminosarum-as well as R. tropici-targeted IGS-based nested PCR and DGGE are useful tools for monitoring the effect of agricultural practices on these and related rhizobial subpopulations in soils.     

Tuberculosis Disease during Pregnancy and Treatment Outcomes in HIV-Infected and Uninfected Women at a Referral Hospital in Cape Town

BackgroundTuberculosis during pregnancy and treatment outcomes are poorly defined in high prevalence tuberculosis and HIV settings.MethodsA prospective cohort study of pregnant and postpartum women identified to be routinely on antituberculosis treatment was conducted at Tygerberg Hospital, Cape Town, South Africa, from January 2011 through December 2011. Maternal tuberculosis disease spectrum and tuberculosis-exposed newborns were characterized by maternal HIV status. Maternal tuberculosis treatment outcomes were documented and a multivariable regression model identified predictors of unfavourable tuberculosis treatment outcomes. Infant outcomes were also described.ResultsSeventy-four women with tuberculosis, 53 (72%) HIV-infected, were consecutively enrolled; 35 (47%) were diagnosed at delivery or postpartum and 22 (30%) of women reported previous antituberculosis treatment. HIV-infected women were 5.67 times more likely to have extrapulmonary tuberculosis (95% CI 1.18–27.25, p = 0.03). All 5 maternal deaths were amongst HIV-infected women. Birth outcomes were available for 75 newborns (2 sets of twins, missing data for 1 stillbirth). Of the 75 newborns, 49 (65%) were premature and 44 (59%) were low birth weight (LBW; <2500 grams). All 6 infants who died and the 4 stillbirths were born to HIV-infected women. Unfavourable tuberculosis treatment outcomes were documented in 33/74 (45%) women. Unfavourable maternal tuberculosis outcome was associated with delivery of LBW infants (OR 3.83; 95% CI 1.40–10.53, p = 0.009).ConclusionsA large number of pregnant women with tuberculosis presented at a provincial referral hospital. All maternal and infant deaths occurred in HIV-infected women and their newborns. Maternal tuberculosis treatment outcomes were poor.     

Development of a Murine Model with Optimal Routes for Bacterial Infection and Treatment, as Determined with Bioluminescent Imaging in C57BL/6 Mice

Mice intragastrically infected with Listeria monocytogenes EGDe and Staphylococcus aureus Xen 36 showed no visible signs of infection over 48 h. However, high numbers (6.2 × 10⁵ cfu/mg feces) of S. aureus Xen 36 were detected 4 h, and 3.3 × 10⁵ cfu/mg feces of L. monocytogenes EGDe 8 h, after administration. Mice intraperitoneally infected with S. aureus Xen 36 (1 × 10⁷ cfu) developed infection immediately after administration and for at least the following 48 h. Injection with higher cell numbers of S. aureus Xen 36 (2 × 10⁸ cfu) resulted in more intense bioluminescence (infection) of the peritoneal cavity. Injection of S. aureus Xen 36 in the tail and penile veins resulted in localized tissue infection for the first 120 h. Injection of S. aureus Xen 36 into the thigh produced a faint bioluminescent signal for 15 min. Nisin F injected into the peritoneal cavity at the same area of infection led to an immediate statistically significant decrease in infection (from 2 × 10⁶ p/s/cm²/sr to 3 × 10⁵ p/s/cm²/sr) within 2 h. Similar results were recorded when nisin F was injected subcutaneously. Intraperitoneal administration is an optimal administration route for bacterial infection and treatment with antimicrobial peptides.     

The nature and perception of fluctuations in human musical rhythms

Although human musical performances represent one of the most valuable achievements of mankind, the best musicians perform imperfectly. Musical rhythms are not entirely accurate and thus inevitably deviate from the ideal beat pattern. Nevertheless, computer generated perfect beat patterns are frequently devalued by listeners due to a perceived lack of human touch. Professional audio editing software therefore offers a humanizing feature which artificially generates rhythmic fluctuations. However, the built-in humanizing units are essentially random number generators producing only simple uncorrelated fluctuations. Here, for the first time, we establish long-range fluctuations as an inevitable natural companion of both simple and complex human rhythmic performances. Moreover, we demonstrate that listeners strongly prefer long-range correlated fluctuations in musical rhythms. Thus, the favorable fluctuation type for humanizing interbeat intervals coincides with the one generically inherent in human musical performances.     

Identification of a key determinant of hepatitis C virus cell culture adaptation in domain II of NS3 helicase

Efficient replication of hepatitis C virus (HCV) replicons in cell culture is associated with specific sequences not generally observed in vivo. These cell culture adaptive mutations dramatically increase the frequency with which replication is established in vitro. However, replicons derived from HCV isolates that have been shown to replicate in chimpanzees do not replicate in cell culture even when these adaptive mutations are introduced. To better understand this apparent paradox, we performed a gain-of-function screen to identify sequences that could confer cell culture replication competence to replicons derived from chimpanzee infectious HCV isolates. We found that residue 470 in domain II of the NS3 helicase is a critical determinant in cell culture adaptation. Substitutions in residue 470 when combined with the NS5A-S232I adaptive mutation are both necessary and sufficient to confer cell culture replication to otherwise inactive replicons, including those derived from genotype 1b HCV-BK and genotype 1a HCV-H77 isolates. The specific substitution at residue 470 required for replication is context-dependent, with R470M and P470L being optimal for the activity of HCV-BK and HCV-H77 replicons, respectively. Together these data indicate that mutations in the NS3 helicase domain II act in concert with previously identified adaptive mutations and predict that introduction of compatible residues at these positions can confer cell culture replication activity to diverse HCV isolates.     

Self-Transmissible Mercury Resistance Plasmids with Gene-Mobilizing Capacity in Soil Bacterial Populations: Influence of Wheat Roots and Mercury Addition

A set of mercury resistance plasmids was obtained from wheat rhizosphere soil amended or not amended with mercuric chloride via exogenous plasmid isolation by using Pseudomonas fluorescens R2f, Pseudomonas putida UWC1, and Enterobacter cloacae BE1 as recipient strains. The isolation frequencies were highest from soil amended with high levels of mercury, and the isolation frequencies from unamended soil were low. With P. putida UWC1 as the recipient, the isolation frequency was significantly enhanced in wheat rhizosphere compared to bulk soil. Twenty transconjugants were analyzed per recipient strain. All of the transconjugants contained plasmids which were between 40 and 50 kb long. Eight selected plasmids were distributed among five groups, as shown by restriction digestion coupled with a similarity matrix analysis. However, all of the plasmids formed a tight group, as judged by hybridization with two whole-plasmid probes and comparisons with other plasmids in dot blot hybridization analyses. The results of replicon typing and broad-host-range incompatibility (Inc) group-specific PCR suggested that the plasmid isolates were not related to any previously described Inc group. Although resistance to copper, resistance to streptomycin, and/or resistance to chloramphenicol was found in several plasmids, catabolic sequences were generally not identified. One plasmid, pEC10, transferred into a variety of bacteria belonging to the β and γ subdivisions of the class Proteobacteria and mobilized as well as retromobilized the IncQ plasmid pSUP104. A PCR method for detection of pEC10-like replicons was used, in conjunction with other methods, to monitor pEC10-homologous sequences in mercury-polluted and unpolluted soils. The presence of mercury enhanced the prevalence of pEC10-like replicons in soil and rhizosphere bacterial populations.The potential use of genetically modified bacteria in agriculture has raised questions pertaining to the spread of introduced recombinant DNA through soil bacterial communities. Gene transfer in soil via conjugation has received much attention, and the focus of most studies has been the transfer and fate of introduced plasmids (6, 22, 27–29, 39). Under favorable conditions, in specific soil microhabitats, or under selection conditions, both self-transmissible and mobilizable plasmids present in introduced hosts can be transferred to introduced recipients, as well as to a variety of indigenous bacteria (15, 20, 27, 28, 33). In particular, rhizospheres of crop plants, such as wheat and sugar beet, provide conditions conducive to conjugal plasmid transfer between bacterial inhabitants (15, 36). When genetically modified bacteria are developed as inoculants for the rhizosphere, insertion of heterologous DNA into non-self-transmissible plasmids or the chromosome might restrict conjugal transfer of this DNA to members of the indigenous bacterial community. However, mobilizing or retromobilizing (33) plasmids present in indigenous soil bacteria could potentially still effect the transfer of the less mobile heterologous DNA via chromosome or plasmid mobilization, which may involve cointegration (9, 19, 31). Such plasmids might thus be responsible for the escape of heterologous DNA from genetically modified bacteria introduced into soil.There is a paucity of knowledge concerning the incidence of plasmids with mobilizing capacity in soils and rhizospheres, as well as concerning the effects of soil factors, such as stresses resulting from pollution or from natural causes (e.g., rhizosphere acidity), on plasmid prevalence and transfer (e.g., reference 38). Whereas it has been suggested that chemical stress often does not enhance plasmid incidence in selected soil bacterial populations (40), pollution in river water or mines (in particular mercury pollution) has been found to exert a selective (enhancing) effect (4, 13).Plasmids of environmental bacteria have classically been obtained by endogenous isolation procedures (20). Endogenous isolation implies that putative plasmid hosts with the phenotype of interest are isolated from soil, after which plasmids are extracted from pure cultures of these strains. On the other hand, pioneering studies performed with river stone epilithon (9) and later extended to soil and sediment (32) have shown that plasmids can be obtained directly from indigenous bacterial communities in new hosts by exogenous isolation. In this approach, plasmids are captured in selectable recipient strains by using mating between these strains and the total bacterial community obtained from an environmental sample. Following incubation, the mating mixture is plated with selection for the recipient and an additional marker gene presumedly located on a plasmid present in the indigenous bacteria (6). The advantage of the exogenous isolation procedure is that no culturing step is required in the mating, which thus allows isolation of plasmids from nonculturable hosts. Furthermore, plasmids are directly selected for their transfer capacity, in addition to the presence of a specific selectable marker.In this study, exogenous plasmid isolation was employed to obtain transferable plasmids from soil bacteria by using mercury resistance as the selectable marker. The objective of this work was to gain insight into the potential present in soil bacterial populations to (retro)mobilize genes out of introduced bacteria into members of the soil bacterial community. Since the incidence of plasmids in soil bacteria is likely influenced by soil ecological factors and selection pressure, the presence of wheat roots and selection by mercury (25) were studied as experimental variables.