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The similarity in the phenotypes of Escherichia coli cells containing a mutation in the radA or sensitive to methyl methanesulfonate (MMS) (sms) gene indicates that the radA and sms mutations are alleles of the same gene. The radA and sms mutants have a similar sensitivity to MMS, medium-dependent sensitivity to UV radiations, and plasmid complementation for UV radiation survival. The radA and sms mutations have the same sequence, except that the radA mutant contains a tyrosine instead of a cysteine residue.

The RadA/Sms protein facilitates DNA repair in Escherichia coli cells damaged by UV radiation, X-rays, and chemical agents. However, the precise mechanism by which RadA/Sms aids DNA repair is unknown. Here we report the production of monoclonal antibodies (MAbs) specific for RadA/Sms for use in biochemical and physiological investigations. Histidine-tagged RadA/Sms (RadA-6xHis) was overproduced in E. coli BL21 cells transformed with the radA/sms coding region in plasmid pRSET A and purified by nickel affinity chromatography. Splenocytes from female BALB/c mice hyperimmunized with the purified protein were fused to SP2/0-Ag14 myeloma cells, and the resultant hybridomas were selected in HAT medium. MAbs were detected in hybridoma culture supernatants by indirect ELISA and Western blot analysis against purified RadA-6xHis. MAbs from four cell lines were further evaluated by Western blotting against peptide maps generated by endoproteinase Glu-C digestion of RadA-6xHis. Each of the four MAbs recognized a unique epitope on the fusion protein. Two of the MAbs (6F5 and 2A2) also detected wild-type (tagless) RadA/Sms produced from the pJS003 plasmid in E. coli K-12 cells. We anticipate that these antibodies will prove useful for the detection, isolation, and functional analysis of RadA/Sms. [ABSTRACT FROM AUTHOR]

The role of DNA repair genes ( and ) in spontaneous mutagenesis was examined in . The spontaneous mutation rate per bacterium per cell division (μ) was determined for the reversion of UAA ( and ), UAG (), and frameshift () mutations, and for the occurrence of forward mutations to valine resistance. Rich growth medium enhanced μ in a wild-type strain but not in a strain. In minimal growth medium, the and strains had the largest μ (1.9–6.2-fold greater than that for isogenic wild-type strains, depending on the mutation assay). The strains carrying or both the and mutations (in combination), i.e., mutations that inhibit error-prone DNA repair, had the lowest μ values (˜10-fold less than the strain). The and mutations also reduced μ (by ˜ 2-fold) in strains. The genetic control of the error prone repair-dependent sector of spontaneous mutagenesis was shown to be qualitatively similar to the genetic control for u.v. radiation mutagenesis. The mutation, which drastically reduced spontaneous mutagenesis, had no effect on genetic recombination. It is proposed that the low level of spontaneous mutagenesis observed in the and the strains is due to errors made during DNA replication, while the enhanced level of spontaneous mutagenesis observed in the wild type, and especially in the and strains, is due to excisable lesions that are produced in the DNA by normal metabolic reactions, and that such unexcised lesions induce mutations via error-prone DNA repair. These results are discussed in terms of their relevance to spontaneous carcinogenesis. [ABSTRACT FROM PUBLISHER]

We evaluated Amblyomma americanum (lone star tick) and Dermacentor variabilis (American dog tick) in northeast Missouri for the presence of Borrelia , Ehrlichia , and Rickettsia bacteria and Heartland virus. We screened 436 individual adult lone star ticks (86% of all ticks collected) and infection rates were 6% for B. lonestari , 19% for E. chaffeensis , 3% for E. ewingii , 36% for R. amblyommatis, and 1% for R. montanensis . In the 189 individual American dog ticks, infection rates were 19% for E. chaffeensis , 15% for E. ewingii , 4% for R. amblyommatis , and 5% for R. montanensis . In addition, we screened 20 pools of adults and 30 pools of nymphs for the Heartland virus which was not detected. Understanding the presence and epidemiology of these causative ( E. chaffeensis and E. ewingii ) and suspected ( B. lonestari, R. amblyommatis, and R. montanensis ) agents in Missouri should increase awareness of potential tick-borne disease in the medical community.

Context: Mobilization of a joint affects local tissue directly but may also have other effects that are mediated through the central nervous system., Objective: To identify differential gene expression in the spinal cords of rats with or without inflammatory joint injury after manual therapy or no treatment., Methods: Rats were randomly assigned to 1 of 4 treatment groups: no injury and no touch (NI/NT), injury and no touch (I/NT), no injury and manual therapy (NI/MT), and injury and manual therapy (I/MT). We induced acute inflammatory joint injury in the rats by injecting carrageenan into an ankle. Rats in the no-injury groups did not receive carrageenan injection. One day after injury, rats received manual therapy to the knee of the injured limb. Rats in the no-touch groups were anesthetized without receiving manual therapy. Spinal cords were harvested 30 minutes after therapy or no touch, and spinal cord gene expression was analyzed by microarray for 3 comparisons: NI/NT vs I/NT, I/MT vs I/NT, and NI/NT vs NI/MT., Results: Three rats were assigned to each group. Of 38,875 expressed sequence tags, 755 were differentially expressed in the NI/NT vs I/NT comparison. For the other comparisons, no expressed sequence tags were differentially expressed. Cluster analysis revealed that the differentially expressed sequence tags were over-represented in several categories, including ion homeostasis (enrichment score, 2.29), transmembrane (enrichment score, 1.55), and disulfide bond (enrichment score, 2.04)., Conclusions: An inflammatory injury to the ankle of rats caused differential expression of genes in the spinal cord. Consistent with other studies, genes involved in ion transport were among those affected. However, manual therapy to the knees of injured limbs or to rats without injury did not alter gene expression in the spinal cord. Thus, evidence for central nervous system mediation of manual therapy was not observed., (© 2014 The American Osteopathic Association.)

Context: Animal models can be used to investigate manual therapy mechanisms, but testing manipulation in animal models is problematic because animals cannot directly report their pain., Objective: To develop a rat model of inflammatory joint injury to test the efficacy of manual therapy in reducing nociception and restoring function., Methods: The authors induced acute inflammatory joint injury in rats by injecting carrageenan into the ankle and then measured voluntary running wheel activity in treated and untreated rats. Treatments included manual therapy applied to the ankle and knee of the injured limb and several analgesic medications (eg, morphine, ketorolac, prednisone)., Results: Intra-articular injection of carrageenan to the ankle produced significant swelling (diameter of the ankle increased by 64% after injection; P=.004) and a robust reduction in voluntary running wheel activity (running distance reduced by 91% compared with controls; P<.001). Injured rats gradually returned to running levels equal to controls over 10 days. Neither manual therapy nor analgesic medications increased running wheel activity relative to untreated rats., Conclusion: Voluntary running wheel activity appears to be an appropriate functional measure to evaluate the impact of an acute inflammatory joint injury. However, efforts to treat the injury did not restore running relative to untreated rats.

Context: With the formal adoption of the seven core competencies, the American Osteopathic Association's Commission on Osteopathic College Accreditation instructed osteopathic medical educators to guide curricular development with these goals in mind. Tools to facilitate and monitor these purposes have been under development separately at each of the nation's colleges of osteopathic medicine., Objective: To demonstrate the utility of a checklist-based curriculum assessment tool, the Matrix for Quality Enhancement, as developed at Kirksville (Mo) College of Osteopathic Medicine-A.T. Still University., Methods: APPLICATION of the Matrix is illustrated using examples selected from our analysis of a set of 16 standardized patient encounters provided as part of a first-year basic science course in medical microbiology. Encounters were developed to improve student understanding of infectious disease entities while also providing a variety of clinical experiences. Feedback on professionalism and humanistic behaviors was also provided. A novel aspect of the Matrix is the inclusion of a component dealing with patient safety., Application: Adding standardized patient encounters to the medical microbiology teaching program at Kirksville College of Osteopathic Medicine was an effective means of integrating educational experiences with the seven core competencies, the requirements of Comprehensive Osteopathic Medical Licensing Examination-USA Level 2-PE (Performance Evaluation), and patient safety issues., Conclusion: The Matrix is a valuable tool for evaluating or developing curricular components that maintain osteopathic integrity while working toward standards for medical education specified by the commission.