Healthcare personnel intestinal colonization with multidrug-resistant organisms.

OBJECTIVES: This study aims to assess the association between patient contact and intestinal carriage of multidrug-resistant organisms (MDRO) by sampling healthcare personnel (HCP) and staff without patient contact. METHODS: For this observational study, we recruited 400 HCP who worked in our 200-bed research hospital and 400 individuals without patient contact between November 2013 and February 2015. Participants submitted two self-collected perirectal swabs and a questionnaire. Swabs were processed for multidrug-resistant Gram-negative bacteria and vancomycin-resistant enterococci (VRE). Questionnaires explored occupational and personal risk factors for MDRO carriage. RESULTS: Among 800 participants, 94.4% (755/800) submitted at least one swab, and 91.4% (731/800) also submitted questionnaires. Extended spectrum ß-lactamase-producing organisms were recovered from 3.4% (26/755) of participants, and only one carbapenemase-producing organism was recovered. No VRE were detected. The potential exposure of 68.9% (250/363) of HCP who reported caring for MDRO-colonized patients did not result in a rate of MDRO carriage among HCP (4.0%; 15/379) significantly higher than that of staff without patient contact (3.2%; 12/376; p 0.55). CONCLUSIONS: This is the largest US study of HCP intestinal MDRO carriage. The low colonization rate is probably reflective of local community background rates, suggesting that HCP intestinal colonization plays a minor role in nosocomial spread of MDROs in a non-outbreak setting. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT01952158.

Horizontal Transfer of Carbapenemase-Encoding Plasmids and Comparison with Hospital Epidemiology Data.

Carbapenemase-producing organisms have spread worldwide, and infections with these bacteria cause significant morbidity. Horizontal transfer of plasmids carrying genes that encode carbapenemases plays an important role in the spread of multidrug-resistant Gram-negative bacteria. Here we investigate parameters regulating conjugation using an Escherichia coli laboratory strain that lacks plasmids or restriction enzyme modification systems as a recipient and also using patient isolates as donors and recipients. Because conjugation is tightly regulated, we performed a systematic analysis of the transfer of Klebsiella pneumoniae carbapenemase (blaKPC)-encoding plasmids into multiple strains under different environmental conditions to investigate critical variables. We used four blaKPC-carrying plasmids isolated from patient strains obtained from two hospitals: pKpQIL and pKPC-47e from the National Institutes of Health, and pKPC_UVA01 and pKPC_UVA02 from the University of Virginia. Plasmid transfer frequency differed substantially between different donor and recipient pairs, and the frequency was influenced by plasmid content, temperature, and substrate, in addition to donor and recipient strain. pKPC-47e was attenuated in conjugation efficiency across all conditions tested. Despite its presence in multiple clinical species, pKPC_UVA01 had lower conjugation efficiencies than pKpQIL into recipient strains. The conjugation frequency of these plasmids into K. pneumoniae and E. coli patient isolates ranged widely without a clear correlation with clinical epidemiological data. Our results highlight the importance of each variable examined in these controlled experiments. The in vitro models did not reliably predict plasmid mobilization observed in a patient population, indicating that further studies are needed to understand the most important variables affecting horizontal transfer in vivo.

ECT and the youth: catatonia in context.

Electroconvulsive therapy (ECT) has been in psychiatric practice for well over half a century, but it continues to incite controversy. However, it is regarded amongst psychiatrists as a safe and effective treatment and at times even a lifesaver. It offers a fairly swift but a time-limited response, opening up opportunities for initiation of more longer lasting treatments. The use of ECT in the youth is limited, and as such good studies are few and far between. The recent Practice Parameters by the American Academy of Child and Adolescent Psychiatry, specifically addressing ECT in adolescents, is indeed a welcome addition. Electrocovulsive therapy is as effective in the youth as it is in the adults, and the indications and contraindication are the same. The administration of ECT follows the same general principles in all age groups. One particular indication is of the use in catatonia, a motor syndrome that could occur with affective disorders, schizophrenia or medical conditions, in which it is considered to be extremely effective. The association between catatonia and autism and spectrum disorders has been noted, and in this situation, ECT is considered by some to be effective. Ethical considerations and that of capacity and informed consent are of paramount importance as are the human rights. Working in partnership with the parents/carers all the way is a must. The lack of information leaflets on ECT especially designed for young patients and their parents has to be rectified soon. Registers based on geographical health regions for those below the age of 18 will assist tremendously in epidemiological studies as well as pave the way toward more evidence-based studies that are essential.

DNA ligase IV deficiency in mice leads to defective neurogenesis and embryonic lethality via the p53 pathway.

DNA ligase IV (LIG4) is a nonhomologous end-joining (NHEJ) protein used for V(D)J recombination and DNA repair. In mice, Lig4 deficiency causes embryonic lethality, massive neuronal apoptosis, arrested lymphogenesis, and various cellular defects. Herein, we assess potential roles in this phenotype for INK4a/ARF and p53, two proteins implicated in apoptosis and senescence. INK4a/ARF deficiency rescued proliferation/senescence defects of Lig4-deficient fibroblasts but not other phenotypic aspects. In contrast, p53 deficiency rescued embryonic lethality, neuronal apoptosis, and fibroblast proliferation/senescence defects but not lymphocyte development or radiosensitivity. Young Lig4/p53 double null mice routinely died from pro-B lymphomas. Thus, in the context of Lig4 deficiency, embryonic lethality and neuronal apoptosis likely result from a p53-dependent response to unrepaired DNA damage, and neuronal apoptosis and lymphocyte developmental defects can be mechanistically dissociated.

The nonhomologous end-joining pathway of DNA repair is required for genomic stability and the suppression of translocations.

We have used spectral karyotyping to assess potential roles of three different components of the nonhomologous DNA end-joining pathway in the maintenance of genomic stability in mouse embryonic fibroblasts (MEFs). MEFs homozygous for mutations that inactivate either DNA ligase IV (Lig4) or Ku70 display dramatic genomic instability, even in the absence of exogenous DNA damaging agents. These aberrant events range from chromosomal fragmentation to nonreciprocal translocations that can involve several chromosomes. DNA-dependent protein kinase catalytic subunit deficiency also promotes genome instability. Deficiency for the p53 cell cycle checkpoint protein has little effect on spontaneous levels of chromosomal instability in Lig4-deficient fibroblasts. However, in the context of ionizing radiation treatment, p53 deficiency allowed visualization of massive acute chromosomal destruction in Lig4-deficient MEFs, which in surviving cells manifested as frequent nonreciprocal translocations. We conclude that nonhomologous DNA end-joining plays a crucial role as a caretaker of the mammalian genome, and that an alternative repair pathway exists that often leads to nonreciprocal translocations.

Interplay of p53 and DNA-repair protein XRCC4 in tumorigenesis, genomic stability and development.

XRCC4 is a non-homologous end-joining protein employed in DNA double strand break repair and in V(D)J recombination. In mice, XRCC4-deficiency causes a pleiotropic phenotype, which includes embryonic lethality and massive neuronal apoptosis. When DNA damage is not repaired, activation of the cell cycle checkpoint protein p53 can lead to apoptosis. Here we show that p53-deficiency rescues several aspects of the XRCC4-deficient phenotype, including embryonic lethality, neuronal apoptosis, and impaired cellular proliferation. However, there was no significant rescue of impaired V(D)J recombination or lymphocyte development. Although p53-deficiency allowed postnatal survival of XRCC4-deficient mice, they routinely succumbed to pro-B-cell lymphomas which had chromosomal translocations linking amplified c-myc oncogene and IgH locus sequences. Moreover, even XRCC4-deficient embryonic fibroblasts exhibited marked genomic instability including chromosomal translocations. Our findings support a crucial role for the non-homologous end-joining pathway as a caretaker of the mammalian genome, a role required both for normal development and for suppression of tumours.

Fulminant babesiosis treated with clindamycin, quinine, and whole-blood exchange transfusion.

BACKGROUND: Babesiosis is an increasingly recognized parasitic infection with manifestations that range from a subclinical or mild flu-like illness to life-threatening disease. Risk factors that may be associated with a more severe clinical course include immunosuppression, splenectomy, and advanced age. The most effective chemotherapeutic regimen, clindamycin plus quinine, is sometimes ineffective in cases of severe disease. CASE REPORT: A previously healthy, 58-year-old man was infected by Babesia microti, presumably through a tick bite. He developed fulminant disease characterized by severe hemolytic anemia, disseminated intravascular coagulation, acute renal failure, and respiratory failure. There was no history of splenectomy or immunodeficiency. He was given oral clindamycin (300 mg/4x/day) 2 days before admission. Oral quinine (650 mg/3x/day) was added upon hospitalization. There was no clinical improvement despite antibiotic therapy with clindamycin and quinine. On the second hospital day, a whole-blood exchange transfusion was performed to simultaneously lower the parasite load and replace the patient's plasma. With an automated blood cell separator, 87 percent of the patient's total blood volume was exchanged. As replacement fluid, 6.7 L of packed RBCs reconstituted with FFP (average Hct, 33%) was used. The patient's Hct increased from 26.9 percent before the exchange to 28.3 percent after the exchange. The percentage of parasitized RBCs decreased from 13.8 percent just before exchange to 4.2 percent immediately after exchange. There was rapid clinical improvement after the whole-blood exchange transfusion. The patient's subsequent clinical course was marked by a disappearance of the parasitemia and continued slow, general improvement. Therapy with clindamycin was continued for 14 days after the exchange transfusion and quinine for 17 days. CONCLUSION: In cases of severe babesiosis, prompt institution of whole-blood exchange transfusion, in combination with appropriate antimicrobial therapy, can be life-saving.

Late embryonic lethality and impaired V(D)J recombination in mice lacking DNA ligase IV.

The DNA-end-joining reactions used for repair of double-strand breaks in DNA and for V(D)J recombination, the process by which immunoglobulin and T-cell antigen-receptor genes are assembled from multiple gene segments, use common factors. These factors include components of DNA-dependent protein kinase (DNA-PK), namely DNA-PKcs and the Ku heterodimer, Ku70-Ku80, and XRCC4. The precise function of XRCC4 is unknown, but it interacts with DNA ligase IV. Ligase IV is one of the three known mammalian DNA ligases; however, the in vivo functions of these ligases have not been determined unequivocally. Here we show that inactivation of the ligase IV gene in mice leads to late embryonic lethality. Lymphopoiesis in these mice is blocked and V(D)J joining does not occur. Ligase IV-deficient embryonic fibroblasts also show marked sensitivity to ionizing radiation, growth defects and premature senescence. All of these phenotypic characteristics, except embryonic lethality, resemble those associated with Ku70 and Ku80 deficiencies, indicating that they may result from an impaired end-joining process that involves both Ku subunits and ligase IV. However, Ku-deficient mice are viable, so ligase IV must also be required for processes and/or in cell types in which Ku is dispensable.

A critical role for DNA end-joining proteins in both lymphogenesis and neurogenesis.

XRCC4 was identified via a complementation cloning method that employed an ionizing radiation (IR)-sensitive hamster cell line. By gene-targeted mutation, we show that XRCC4 deficiency in primary murine cells causes growth defects, premature senescence, IR sensitivity, and inability to support V(D)J recombination. In mice, XRCC4 deficiency causes late embryonic lethality accompanied by defective lymphogenesis and defective neurogenesis manifested by extensive apoptotic death of newly generated postmitotic neuronal cells. We find similar neuronal developmental defects in embryos that lack DNA ligase IV, an XRCC4-associated protein. Our findings demonstrate that differentiating lymphocytes and neurons strictly require the XRCC4 and DNA ligase IV end-joining proteins and point to the general stage of neuronal development in which these proteins are necessary.