The use of laboratory intervention to stem the flow of fresh-frozen plasma

Four methods of laboratory intervention were tested by the hospital blood bank in an effort to modify the use of fresh-frozen plasma (FFP). Over a one-year period, a utilization audit was serially initiated with feedback to physicians, a recurrent educational program was introduced for housestaff delineating guidelines for FFP use, a form was introduced requiring justification for FFP orders, and a policy was established requiring pathologist approval of FFP in patients with normal or no coagulation studies. Overall, in comparing the period following all forms of intervention (February 1986-October 1986) to the baseline period prior to any form of intervention (July 1984-March 1985), FFP use dropped 52% in the face of a 17% increase in red blood cell use. It was concluded that blood bankers can dramatically alter the use of this product using established methods for modifying physician ordering behavior.     

Confirmation that the Type I collagen gene on chromosome 17 is COL1A1 (α1(I)), using a human genomic probe

A cloned 15 kb genomic fragment from the human α1 (I) collagen gene (COL1A1) has been used as a probe on restriction digests of DNA from human-mouse somatic cell hybrids. Positive results on hybrids containing chromosome 17 as their only karyotypically visible human material confirm the assignment of this gene to chromosome 17. Hybrids which contain fragments of chromosome 17 are used to confirm the localization to 17q21-qter.     

The lipid raft microdomain-associated protein reggie-1/flotillin-2 is expressed in human B cells and localized at the plasma membrane and centrosome in PBMCs

Reggie-1/flotillin-2 is a plasma membrane-associated cytoplasmic protein, which defines non-caveolar raft microdomains. Reggie-1/flotillin-2 is enriched in detergent insoluble (TX100) membrane fractions (DIG), co-localizes with activated GPI-linked proteins and the fyn-kinase in neurons and T cells, and thus apparently participates in the assembly of protein complexes essential for signal transduction. In T cells activated by crosslinking the GPI-linked protein Thy-1 or by crosslinking the ganglioside GM1, reggie-1/flotillin-2 co-localizes with the T cell receptor. To determine whether reggie-1/flotillin-2 is also expressed in B cells, primary B cells from human blood and cell lines representing the developmental stages of pro, pre, mature and plasma B cells were analyzed by Western blotting, RT-PCR and immunofluorescence. Here, we show that reggie-1/flotillin-2 is expressed throughout B cell development, as well as in primary B cells, purified by cell sorting. On non-activated mature B cell Raji cell line we found reggie-1/flotillin-2 are exclusively in the detergent (TX100) insoluble membrane fractions that are staining positive for the raft marker GM1. Immunofluorescence microscopy showed that reggie-1/flotillin-2 is localized at the plasma membrane and marks intracellular spots in PBMCs. Confocal co-localization studies showed that reggie-1/flotillin-2 is associated with the plasma membrane, and the centrosomes (microtubule organizing centers) in these PBMCs. Comparison of reggie-1/flotillin-2 cDNA sequences with the genomic sequence database allowed us to determine the exon/intron structures in mouse and human. The gene organizations are highly conserved suggesting an important function of reggie-1/flotillin-2. Since reggie/flotillin proteins co-cluster with the T cell receptor and fyn kinases upon T cell stimulation, our findings of reggie-1/flotillin-2 in B cells suggest a similar role in B cell function.     

Plasmids of Ewingella americana: supplementary epidemiologic markers in an outbreak of pseudobacteremia.

During an outbreak of pseudobacteremia in a children's hospital, Ewingella americana was found in blood cultures from 20 patients. E. americana was inoculated into blood culture bottles at the time of specimen collection due to cross contamination from nonsterile, citrated blood collection tubes used for coagulation studies. Antimicrobial susceptibility testing and plasmid profiling were used to assess the association between patient isolates and isolates from unused blood collection tubes. All E. americana isolates had similar antibiograms (i.e., resistance only to cephalothin) when tested at 37 degrees C. However, when the same isolates were tested for antimicrobial susceptibility at 25 degrees C, a different antibiogram (i.e., resistance to chloramphenicol, ampicillin, and cephalothin) was found. The majority of these isolates also demonstrated a unique four-plasmid profile (130, 56, 4.6, and 3.1 megadaltons), and two of these plasmids (130 and 56 megadaltons) were characterized as temperature-sensitive plasmids. An epidemiologic link between outbreak-associated isolates obtained from different time periods in the outbreak was supported by evidence of a significant trend in the ability of the outbreak-associated isolates to reduce nitrate, together with the presence of the resistance antibiogram at 25 degrees C and the demonstration of the unique four-plasmid profile.     

Pathogenic anti-DNA antibodies in SLE: idiotypic families and genetic origins

We have adopted an idiotypic approach to study the double stranded DNA (dsDNA) binding antibodies of systemic lupus erythematosus (SLE). Three anti-idiotypic reagents, 8.12, 3I, and F4, identify cross reactive idiotypes that are each expressed on anti-dsDNA antibodies in the sera of many patients with SLE. These idiotypic antibodies are implicated in the pathogenesis of SLE as they are present in immune complex deposits in the kidneys of patients with SLE glomerulonephritis. The autoantibody associated idiotypes are also expressed on antibodies that do not bind DNA. We are investigating the origin of the pathogenic anti-dsDNA antibodies of SLE by comparing the autoantibodies, the antibodies to foreign antigens, and the myeloma proteins that express each SLE associated idiotype. In conjunction with serological analysis of these idiotypic systems, molecular genetic studies indicate that both the 8.12 and the 3I autoantibody associated idiotypes may be germline encoded, while the F4 idiotype is generated by somatic mutation. The data further suggest that the antigenic specificity of the pathogenic anti-DNA antibodies of SLE is acquired through somatic mutation of germline immunoglobulin genes. By studying the regulation of genes capable of encoding pathogenic autoantibodies, in both SLE patients and non-autoimmune individuals, we may be able to elucidate the pathogenesis of autoimmune disease and begin to design more effective therapeutic interventions.     

Ewingella americana: recurrent pseudobacteremia from a persistent environmental reservoir.

From September 1981 through April 1984, 20 patients at one hospital were identified with Ewingella americana pseudobacteremia. Case-control studies demonstrated an association between having a positive blood culture for E. americana and having blood for culture obtained simultaneously with blood obtained for coagulation studies (15 of 19 case patients versus 4 of 38 controls; P = 4.5 X 10(-7)). Review of blood-drawing procedures showed that blood for coagulation studies and culture was drawn with the same syringe, and coagulation tubes were filled before blood culture tubes. Some phlebotomists were not using new sterile needles to inoculate blood culture bottles. Collection tubes for coagulation studies were prepared in the hospital, and E. americana was isolated from all 52 unused coagulation tubes tested. Solutions prepared in the hospital may constitute a persistent inanimate environmental reservoir for this uncommon microorganism. Pseudobacteremia can result in unnecessary antimicrobial therapy for some patients, incurring the risks of adverse drug reactions, selection of drug-resistant bacteria, and increased health care costs.     

Risk factors for HIV infection in injection drug users and evidence for onward transmission of HIV to their sexual partners in Chennai, India

OBJECTIVES: Determining HIV prevalence in injection drug users (IDUs) and their regular sex partners in Chennai, India. METHODS: A total of 226 IDUs and their regular sex partners were enrolled during April-July 2003. After informed consent was obtained, a semistructured questionnaire was administered and serum was tested for HIV antibody. RESULTS: The HIV seroprevalence was 30% (68/226) in IDUs and 5% in their regular sex partners (11/226). While in 25% of couples only the male partner was HIV positive, 5% of the couples were concordant for HIV infection and 70% were HIV negative. Fifty-seven percent of the HIV-positive IDUs and 45% of the HIV-infected women thought that they had "no chance" or "very little chance" of getting HIV, reflecting low HIV risk perception. More than 20% IDUs reported borrowing or lending of injection equipment. In univariate analyses "sex" and "condom use" with sex workers had no bearing but "more than twice a day injecting frequency," "history of incarceration," "tattoos," "recruitment from northern part of the city," and ever-injecting drugs in drug-selling places had significant association with HIV infection in IDUs. In an adjusted model, the odds of HIV infection were 2 times higher among IDUs who had ever injected drugs in drug-selling places and 6 times higher in those who were recruited from the northern part of central Chennai. CONCLUSION: Reducing sharing of injection equipment and unsafe tattooing through targeted and environmental interventions, increasing HIV risk perception, and promoting safer sex practices among IDUs and their sex partners are urgent program needs.     

Nosocomial Pseudomonas pickettii colonization associated with a contaminated respiratory therapy solution in a special care nursery.

Pseudomonas pickettii caused respiratory tract colonization in five infants in the special care nursery of a Chicago hospital. All organisms had the same antimicrobial susceptibilities. Endotracheal suctioning with saline from 5-ml unit-dose vials was identified by epidemiologic investigation as a risk factor for colonization. The vials of saline were contaminated with a strain of P. pickettii having the same antimicrobial susceptibility pattern as the isolates from patients. As part of an investigation of the manufacturing plant where the saline solution was produced, P. pickettii was recovered from deionized water used to make the product and from several sites in the processing line. Bypassing of a 180 degrees F (ca. 82 degrees C) water-holding tank appeared to be temporally related to product contamination. The ability of P. pickettii to survive and grow in this solution has been demonstrated in the laboratory. This outbreak demonstrates that, despite pertinent Food and Drug Administration regulations and company programs for identifying such contamination, intrinsically contaminated solutions can occasionally reach the bedside of the patient.     

Effect of histamine and methacholine on nasal airway resistance in atopic and nonatopic subjects: Comparison with bronchial challenge and skin test responses

Serial nasal, intracutaneous, or bronchial challenges were carried out with solutions containing 2- or 3-fold increments in histamine (H) or methacholine (Meth) concentration until nasal airway resistance (NAR) increased by more than 100%, a large intracutaneous reaction was elicited, or FEV₁ decreased by 20% or more. Thirty nonatopic and 48 asymptomatic atopic subjects were studied, the latter group divided into rhinitic patients with and without asthma. Several types of data analysis demonstrated there was no significant difference in the nasal or cutaneous effects of H or Meth between the atopic and nonatopic groups. Comparable results were obtained in a subgroup of 39 subjects (13 normal, 13 atopic, and 13 atopic with asthma) who underwent all six test sequences (i.e., nasal, cutaneous, and bronchial with both drugs). As expected, the asthmatics showed significantly increased bronchial reactivity to both agents. In comparison with Meth, H had a much greater effect on the nasal mucosa and skin than on the bronchi. It is concluded that, contrary to bronchial responses, but in accord with cutaneous reactivity, the nasal responses of nonatopic subjects, atopic persons with allergic rhinitis alone, and subjects with both allergic rhinitis and asthma show no intergroup differences on testing with H or Meth.     

Hydrosalpinges adversely affect markers of endometrial receptivity

While in-vitro fertilization (IVF) was initially developed in women with tubal factor infertility, recent clinical studies have suggested that the presence of hydrosalpinges lowers implantation and pregnancy rates. We postulated that these hydrosalpinges cause impaired endometrial receptivity. A total of 103 women with hydrosalpinges were prospectively evaluated, and compared with 55 infertile and 44 fertile controls. All women had endometrial biopsies during the window of implantation, analysed by conventional histological criteria, and also stained for three integrin markers of endometrial receptivity (alpha1beta1, alpha4beta1 and alpha vbeta3). Women with hydrosalpinges (cases) expressed significantly less of the alpha vbeta3 integrin compared with controls. There was no difference in expression of alpha1beta1 or alpha4beta1 among groups. A significantly greater number of cases had out of phase histology and missing alpha vbeta3 (type I defects) and absent integrin expression despite normal histological maturation (type II) defects, compared with controls. Of 20 women with impaired endometrial receptivity who were also biopsied after hydrosalpinx surgery, 70% demonstrated increased alpha vbeta3 expression. Seventy-seven percent of type I and 57% of type II defects were corrected postoperatively. Using markers of endometrial receptivity, this study demonstrates that inflammatory hydrosalpinges have an adverse effect on endometrial receptivity, which in some cases may be overcome by surgical treatment of the hydrosalpinx.