Clustering of mutations blocking synthesis of xanthan gum by Xanthomonas campestris.

Mutations that block the synthesis of xanthan gum by Xanthomonas campestris B1459S-4L-II were isolated as nonmucoid colonies after treatment with ethyl methanesulfonate. Complete libraries of DNA fragments from wild-type X. campestris were cloned into Escherichia coli by using a broad-host-range cosmid vector and then transferred into each mutant strain by conjugal mating. Cloned fragments that restored xanthan gum synthesis (Xgs+; mucoidy) were compared according to restriction pattern, DNA sequence homology, and complementation of a subset of Xgs- mutations. Groups of clones that contained overlapping homologous DNA were found to complement specific Xgs- mutations. The results suggest clustering of the genetic loci involved in xanthan synthesis. The clustering occurred within three unlinked regions. Two forms of complementation were observed. In most instances, independently isolated cosmid clones that complemented a single mutation were found to be partially homologous. Less frequent was the second form of complementation, in which two cosmid clones that lacked any homologous sequences restored the mucoid phenotype to a single mutant. Finally, xanthan production was measured for wild-type X. campestris carrying multiple plasmid copies of the cloned xanthan genes.     

Role of 1-acyl-2-lyso-phosphatidylcholine acyl transferase in the biosynthesis of pulmonary phosphatidylcholine.

The majority of pulmonary dipalmitoyl phosphatidylcholine, whose surface tension lowering properties are necessary for maintenance of alveolar structural integrity and lung function, is synthesized via a reacylation mechanism. V_max and K_m values were obtained with the 1-acyl-2-lyso-PC: acyl-CoA acyl transferase system in rabbit lung microsomes and long term cultures of rabbit lung cells derived from type ll pneumonocytes enriched fractions. The data, especially for V_max, can be interpreted to indicate that this reacylation system may be an important factor in the biosynthesis of lung dipalmitoyl phosphatidylcholine.     

ALGAL EXCRETION OF 14C-LABELED COMPOUNDS AND MICROBIAL INTERACTIONS IN CYANIDIUM CALDARIUM MATS1

Acid spring effluents are often covered with mats of the eucaryotic phycocyanin-containing alga. Cyanidium caldarium. The primary bacterial component of such mats is an acidophilic strain of Bacillus coagulans, and the primary fungal component is Dactylaria gallopava. Because of the limited species diversity, C. caldarium mats appeared to be an excellent system for studying algal excretion and various microbial interactions in nature. From 2 to 6% of the NaH¹⁴CO₃ taken up by natural or laboratory populations of the alga was excreted as ¹⁴C-labeled materials. The maximum excretion occurred at temperature, light, and pH values optimum for NaH¹⁴CO₃ uptake. However, when excretion was expressed as a percentage of NaH14 CO₃ uptake, a higher percentage of the radioactivity was excreted at nonoptimal conditions for NaH¹⁴CO₃ uptake. Fungal biomass was directly proportional to algal density, but bacterial numbers varied widely and did not correlate with algal numbers. The bacterial and fungal components could be grown in mixed culture with either growing C. caldarium cultures or in an extract prepared, by healing algal cells.     

EMG and EEG biofeedback training in the treatment of a 10-year-old hyperactive boy with a developmental reading disorder

The serial application of electromyographic (EMG) and sensorimotor (SMR) biofeedback training was attempted with a 10-year-old boy presenting a triad of symptoms: an attention deficit disorder with hyperactivity, developmental reading disorder, and ocular instability. Symptom elimination was achieved, for all three aspects of the triad, following the procedure of first conditioning a decrease in EMG-monitored muscle tension and then conditioning increases in the amplitude of sensorimotor rhythm over the Rolandic cortex. The learned reduction of monitored EMG levels was accompanied by a reduction in the child's motoric activity level to below that which had been achieved by past administration of Ritalin. In addition, the attention deficit disorder with hyperactivity was no longer diagnosable following the EMG biofeedback training. The learned increase in the amplitude of monitored SMR was accompanied by remediation of the developmental reading disorder and the ocular instability. These results remained unchanged, as ascertained by follow-ups conducted over a 24-month period subsequent to the termination of biofeedback training.     

Expression of carbonic anhydrase II mRNA and protein in oligodendrocytes during toxic demyelination in the young adult mouse

The aim of this study was to identify events that might take place in oligodendrocytes early in the process of demyelination, i.e., before the occurrence of massive loss of myelin. It was considered important to focus on demyelination and remyelination in young adults, in whose brains there would be relatively few juvenile glial precursor cells. CAII mRNA and protein were used to monitor changes in oligodendrocytes during cuprizone intoxication in the mice. After four or eight weeks of cuprizone feeding CAII message became less plentiful in oligodendrocyte processes. Two days after removal of cuprizone CAII message had appeared in those cell processes. Four or eight weeks after beginning cuprizone feeding CAII protein had decreased∼25% in forebrain homogenates. The loss of CAII protein was reversible after four weeks on cuprizone, but not after eight weeks. After four weeks of cuprizone feeding the numbers of CAII mRNA-prositive oligodendrocytes had decreased by ∼50%m and after eight weeks, by ∼80%. By 12 weeks, however, the number of oligodendrocytes expressing CAII mRNA had spontaneously returned to normal levels. Before eight weeks of cuprizone feeding, loss of myelinated tracts in the corpus striatum was reversible. Demyelination appreared to become irreversible after nine weeks of intoxication, although expression of CAII mRNA remained reversible. The results suggest that in the brain of the young adult, oligodendrocytes expressing message for CAII can be generated spontaneously shortly before demyelination becomes irreversible, and can survive and continue to express CAII mRNA but not CAII protein. Special issue dedicated to Dr. Marion E. Smith.     

A muscle-specific intron enhancer required for rescue of indirect flight muscle and jump muscle function regulates Drosophila tropomyosin I gene expression.

The control of expression of the Drosophila melanogaster tropomyosin I (TmI) gene has been investigated by P-element transformation and rescue of the flightless and jumpless TmI mutant strain, Ifm(3)3. To localize cis-acting DNA sequences that control TmI gene expression, Ifm(3)3 flies were transformed with P-element plasmids containing various deletions and rearrangements of the TmI gene. The effects of these mutations on TmI gene expression were studied by analyzing both the extent of rescue of the Ifm(3)3 mutant phenotypes and determining TmI RNA levels in the transformed flies by primer extension analysis. The results of our analysis indicate that a region located within intron 1 of the gene is necessary and sufficient for directing muscle-specific TmI expression in the adult fly. This intron region has characteristics of a muscle regulatory enhancer element that can function in conjunction with the heterologous nonmuscle hsp70 promoter to promote rescue of the mutant phenotypes and to direct expression of an hsp70-Escherichia coli lacZ reporter gene in adult muscle. The enhancer can be subdivided further into two domains of activity based on primer extension analysis of TmI mRNA levels and on the rescue of mutant phenotypes. One of the intron domains is required for expression in the indirect flight muscle of the adult. The function of the second domain is unknown, but it could regulate the level of expression or be required for expression in other muscle.     

Teaching a new mouse old tricks: Humanized mice as an infection model for Variola virus

Smallpox, caused by the solely human pathogen Variola virus (VARV), was declared eradicated in 1980. While known VARV stocks are secure, smallpox remains a bioterrorist threat agent. Recent U.S. Food and Drug Administration approval of the first smallpox anti-viral (tecovirimat) therapeutic was a successful step forward in smallpox preparedness; however, orthopoxviruses can become resistant to treatment, suggesting a multi-therapeutic approach is necessary. Animal models are required for testing medical countermeasures (MCMs) and ideally MCMs are tested directly against the pathogen of interest. Since VARV only infects humans, a representative animal model for testing therapeutics directly against VARV remains a challenge. Here we show that three different humanized mice strains are highly susceptible to VARV infection, establishing the first small animal model using VARV. In comparison, the non-humanized, immunosuppressed background mouse was not susceptible to systemic VARV infection. Following an intranasal VARV challenge that mimics the natural route for human smallpox transmission, the virus spread systemically within the humanized mouse before mortality (~ 13 days post infection), similar to the time from exposure to symptom onset for ordinary human smallpox. Our identification of a permissive/representative VARV animal model can facilitate testing of MCMs in a manner consistent with their intended use.     

Flight activity and dispersal of the cabbage seedpod weevil (Coleoptera: Curculionidae) are related to atmospheric conditions

The cabbage seedpod weevil, Ceutorhynchus obstrictus (Marsham) (Coleoptera: Curculionidae), is an invasive pest of canola (Brassica napus L. and Brassica rapa L.) in western Canada. Under current climatic conditions, C. obstrictus is spreading from established populations in southwestern Alberta at ≈ 55 km/yr. We studied the influence of climatic conditions on C. obstrictus flight behavior in 2007 and 2008 and eastward dispersal from the western border of Saskatchewan from 2002 to 2007. Positive linear relationships between increases in mean temperature and flight height and between greater mean maximum temperature and expanded dispersal distances were significant. Increases in relative humidity were associated with reduced flight heights and dispersal distances. We developed models that predict the relationships of temperature and relative humidity with flight height and with dispersal distance. We also discuss implications for C. obstrictus dispersal under current climatic conditions and in the context of predicted climate change.