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Objective: During the current pandemic, COVID-19 has been detected in patients using real-time reverse transcriptase-polymerase chain reaction (RT-PCR) that confirms the presence of SARS-CoV-2 RNA. The demand for increased testing, particularly for asymptomatic individuals required alternative approaches to single-patient RT-PCR testing, such as pooling., Methods: This study explored the impact of dilution on the detectability of SARS-CoV-2 in asymptomatic patients using RT-PCR and demonstrated that pooling can be effective in low prevalence populations., Results: The RT-PCR results for the 3:1, 5:1, and 7:1 aliquot samples showed little differences in CT values, confirming detection capability at these dilutions., Conclusion: Based on the results of the present study, a pooled approach with up to 5:1 sample aliquots and using the current RT-PCR methodology likely will detect SARS CoV2 RNA among asymptomatic patients., (© American Society for Clinical Pathology 2020. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Rapid delivery of proper antibiotic therapies to infectious disease patients is essential for improving patient outcomes, decreasing hospital lengths-of-stay, and combating the antibiotic resistance epidemic. Antibiotic stewardship programs are designed to address these issues by coordinating hospital efforts to rapidly deliver the most effective antibiotics for each patient, which requires bacterial identification and antimicrobial susceptibility testing (AST). Despite the clinical need for fast susceptibility testing over a wide range of antibiotics, conventional phenotypic AST requires overnight incubations, and new rapid phenotypic AST platforms restrict the number of antibiotics tested for each patient. Here, we introduce a novel approach to AST based on signal amplification of bacterial surfaces that enables phenotypic AST within 5 hours for non-fastidious bacteria. By binding bacterial surfaces, this novel method allows more accurate measurements of bacterial replication in instances where organisms filament or swell in response to antibiotic exposure. Further, as an endpoint assay performed on standard microplates, this method should enable parallel testing of more antibiotics than is currently possible with available automated systems. This technology has the potential to revolutionize clinical practice by providing rapid and accurate phenotypic AST data for virtually all available antibiotics in a single test.

Background: In 2010, the Tennessee Department of Health, in collaboration with the Centers for Disease Control and Prevention (CDC), expanded influenza surveillance in Tennessee to include other respiratory viruses., Objectives: To determine the prevalence and seasonality of influenza and other respiratory viruses during the influenza seasons of 2010-2012., Methods: Nasal and nasopharangeal swabs/washings from persons with influenza-like illness were collected across Tennessee. Influenza and other respiratory viruses were identified using a molecular-based respiratory virus panel. Influenza A positives were subtyped using real-time PCR according to the CDC protocol. Data were analyzed to describe frequency and seasonality of circulating strains., Results: Of the 933 positive specimens, 60·3% were identified as influenza viruses, 19·8% rhinovirus/enterovirus, 8·6% respiratory syncytial virus (RSV), 5·8% metapneumovirus, 3·0% adenovirus, and 2·5% parainfluenza viruses. In the 2010-2011 season, influenza B was prominent during weeks 48-3, while influenza A(H1N1) was most frequently identified during weeks 4-10. Influenza A(H3N2) was present at lower levels during weeks 48-17. However, in the 2011-2012 season, overall numbers of influenza cases were reduced and influenza A (H3N2) was the most abundant influenza strain. The expanded surveillance for other respiratory viruses noted an increase in identified specimens from the first to the second season for adenovirus, metapneumovirus, RSV, and rhinovirus/enterovirus., Conclusions: This study provides data of the influenza strains in circulation in Tennessee. It also establishes a baseline and time of year to expect other respiratory viruses that will aid in detecting outbreaks of non-influenza respiratory viruses in Tennessee., (© 2013 John Wiley & Sons Ltd.)

Long term sequential study of immune responses in the same individuals is difficult from the time commitment required and the problem of maintaining enough subjects to provide for comparative analysis. We closely studied one hundred women with silicone mammary devices through cross sectional analysis up to 26 years post implantation and a similar sample of women to 6 years post explantation. The T cell index, calculated from tritiated thymidine incorporation during lymphoblast transformation, rose to a post implant peak at 10.5-12.0 years, falling progressively over the next 14.0-15.5 years to values indicative of probable immune quiescence. Post explantation, the index rose over the first 3 years and then sharply declined to within the range for unexposed controls. The shape of these time curves contains considerable information referent cell dynamics for both stimulatory and inhibitory factors and for demonstrating net group effects, appropriate to analysis in the cross sectional perspective. When a subset of four women was studied frequently and sequentially up to 8 years, an internal oscillatory pattern emerged, focusing attention on both the stimulatory and the inhibitory aspects of long term clonal expansion. IL-2 has stimulatory and inhibitory properties at different levels of production and is considered a prime candidate as the essential cytokine. The equations have details, however, which require exploration beyond any such provisional conclusion. The analytic process was aided by normalization of oscillatory data to eliminate subject variability and by Pareto optimization to assess the trend shown by normalization. Pareto analysis revealed two minimally coordinated oscillations, one over time and the other along net clonal expansion or decline of the siloxane specific T lymphocyte clone. The segments of the time related oscillation greatly exceeded the reaction times of cytokines currently known to be active in T cell regulation. Although the ultimate controlling factor(s) may be cytokine or chemokine combinations, the data are compatible with some more basic regulatory factor(s) of cell integrity, including limits on the number of cell divisions which can be sustained in long term immunopathic lesions, among other processes.

Silicones have an adverse effect on human health well beyond that suggested by the recent superficial public controversy. The evidence for immune responses to injected/implanted silicones is extensive, detailed, often very specific, and not at all new. Comprehending the immunopathogenicity, realized and potential, of silicone has grown as our general understanding of the immune system has developed. Several major issues in furthering this comprehension pertain to the nature of the essential epitope, special risk of silicones to women, and definition of the chronic disease complex so evident clinically, one defying classification within currently traditional disease categories and states. The commentary presented here emphasizes the immunopathic evidence, explores the question of the essential epitope, estimates the minimal threshold of silicone load for immune reactivity, presents a profile of autoantibodies for siliconosis, and calls attention to specific silicone-based female contraceptive modalities. The silicone content of personal care products, not always revealed by retail package labeling, is explored as a potential sensitizing factor in the environment.

Thirty-nine patients with fibromyalgia syndrome (FMS) according to American College of Rheumatology criteria were studied for cell-mediated sensitivity to environmental chemicals. Lymphocytes were tested by standard [(3)H]thymidine incorporation in vitro for T cell memory to 11 chemical substances. Concanavalin A (Con A) was used to demonstrate T cell proliferation. Controls were 25 contemporaneous healthy adults and 252 other concurrent standard controls without any aspect of FMS. Significantly higher (P < 0.01) stimulation indexes (SI) were found in FMS for aluminum, lead, and platinum; borderline higher (0.05 > P > 0.02) SI were found for cadmium and silicon. FMS patients showed sporadic responses to the specific substances tested, with no high-frequency result (>50%) and no obvious pattern. Mitogenic responses to Con A indicated some suppression of T cell functionality in FMS. Possible links between mitogenicity and immunogenic T cell proliferation, certain electrochemical specifics of granuloma formation, maintenance of connective tissue, and the fundamental nature of FMS are considered., (Copyright 2000 Academic Press.)

We present two atypical cases of protein studies that were evaluated by immunofixation and immunosubtraction using capillary zone electrophoresis and high-resolution agarose gel electrophoresis. The first study showed an abnormal peak in the beta region by capillary zone electrophoresis that was located in the gamma region of the high-resolution agarose gel electrophoresis. Further investigation showed that this monoclonal protein was displaced due to binding with beta-lipoproteins. In the second case, a large peak was detected in the alpha-2 region and was shown by capillary zone electrophoresis to be a non-proteinaceous material that mimicked a paraprotein.

A large surgical wound is required for implantation of silicone mammary devices. Formation of capsules around silicone devices follows wound healing processes except that the healing is conformed and significantly delayed by the physical presence of the implant. Multilayered capsules are thicker and lymphocytic and plasmalymphocytic vasculitis, markers for delayed hypersensitivity, also correlate with thicker capsules. Polyurethane-coated devices induce very thick capsules that remain so for over 20 years. By contrast, gel and saline content devices show maximum thickness at 6. 5 years. Active T(H) lymphocyte memory does not differ by implant type for individuals with devices in place and that for gel content devices peaks at 10.5 years. There was a significant decrease in T cell indexes only after the removal of saline content devices. Comparison of the rate of formation of the periprosthetic capsule with the healing time of large wounds of similar size indicates that silicone devices interfere with the healing process, requiring substantially more time. This extended period has the potential for enhancing autoimmune conversion as a consequence of persistent delayed hypersensitivity., (Copyright 1999 Academic Press.)

Objective: Evaluate capillary zone electrophoresis for analyzing beta proteins for concentrations of transferrin and complement., Design: Thirty normal sera were used to establish expected ranges for these proteins by re-gating on the specific peaks from the original histograms. A total of 61 sera with elevated beta proteins were evaluated by this method. An additional 50 sera with an acute phase reaction were evaluated., Setting: All tests were done at Baptist Regional Laboratories on a Beckman CZE/1000 2000 instrument., Patients or Other Participants: Sera were taken from excess sera used for clinical testing. Thirty normal serum donors were tested. The 61 sera with elevated beta proteins were evaluated on a prospective basis following clinical testing. The 50 sera with acute phase patterns were evaluated retrospectively following clinical testing., Intervention: None, Results: Among the 61 sera with increased beta protein, 34 (56%) had elevated transferrin but 57 (93%) had elevated complement levels. In the 50 sera from the acute phase reactant group, 24 sera had elevated complement., Outcome: The study showed that capillary zone electrophoresis could effectively be used to evaluate specific levels of transferrin and complement without further testing., Conclusions: These findings show that with this method, transferrin and complement can be quantitatively measured without further analysis and that the majority of beta protein increases are due to elevated complement levels in our population.

A variant of hepatitis E virus (HEV), designated HEV US-1, was identified in a hepatitis patient in the United States (US); the patient had no history of travel to areas where HEV is endemic. Nucleotide sequences were obtained from the 5' end of open reading frame (ORF) 1 (1418 nt), the 3' end of ORF1 (1359 nt), the entire ORF2 and ORF3 regions, and the 3'-untranslated region (2127 nt). The HEV US-1 strain is significantly divergent from other human HEV isolates with nucleotide identities ranging from 76.8 to 77.5%. Phylogenetic analyses indicate that HEV US-1 and a recently discovered HEV variant from swine may represent separate isolates of a new strain of HEV, significantly divergent from the Mexican and Burmese strains. Synthetic peptides derived from the carboxyl amino acids of ORF2 and ORF3 were shown to be useful for detecting exposure to HEV. In addition, IgM class antibodies directed against HEV US-1 synthetic peptides were detected in the US patient infected with HEV US-1, but were absent using synthetic peptides from the Burmese or Mexican strains of HEV. A preferential reactivity to HEV US-1 specific peptides has lead to the identification of a second isolate of this virus also from a patient with acute hepatitis from the US. The discovery of these HEV variants may be important in understanding the worldwide distribution of HEV infection.

Recent evidence confirms the fundamental involvement of the human immune system in the reaction to implantation of silicone-based medical devices. An as yet-to-be particularized epitope of many complex substances sharing siloxane structures is presented through the MHC-II apparatus with development and retention of T cell memory. This memory can be tested for in practical terms using one or more forms of silica, which links the immuno-histopathology and autoimmune attributes of "silicosis" with those of "siliconosis." The lesions of siliconosis are typical of those for persistent antigens and delayed, cell mediated hypersensitivity. The basic descriptive pathology of the reaction to silicone has been known since soon after introduction of silicones in medical procedures, with the exception of some details related to the more recent discoveries on the role of cytokines in the immunopathic process. The clinical consequences of siliconosis are common and can be severe in some individuals implanted with silicone devices.

Mononuclear cells were isolated by Ficoll-Hypaque density gradient centrifugation from randomly selected silicone breast implant recipients for testing. Restricted antibody to HLA-DR (28-33 kD) depleted the concanavalin A mitogenic response which was expected but failed to inhibit the proliferative response to silicon dioxide. Further testing with monoclonal antibodies to HLA-DP, -DQ, and a second -DR with specificity for the NS1 region of the MHC class II genome, all markedly inhibited proliferation of T cells despite otherwise adequate stimulation by concanavalin A or silicon dioxide. Monoclonal antibodies directed against B7-1 also inhibited proliferation of T cells following stimulation with concanavalin A or silicon dioxide. These results confirm the T-cell response to silicon dioxide is monocyte-dependent and not a superantigen as has been speculated.

A patient who had recently traveled from India to Tennessee was found to have acute hepatitis E. Improved recognition and serologic testing have increased the diagnosis of hepatitis E in the United States. For those at risk, hepatitis E should be considered in the differential diagnosis of acute hepatitis.

Since the isolation of human immunodeficiency virus (HIV), numerous laboratory tests have been developed to confirm the immunologic response to the virus. In 1991, the state of Tennessee mandated that clinical laboratories performing Western blot analyses for HIV report all new cases of confirmed HIV infection. This report is a summary of HIV confirmed cases evaluated by this facility from 1985 until July 1, 1995. During each year, the laboratory screens approximately 20,000 persons for HIV and all positive screens are confirmed by an FDA-approved Western blot. During the period described, the laboratory has confirmed HIV serologic positivity in 4,096 individuals, principally from Shelby County and surrounding Tennessee counties. Although these data are not complete, they clearly indicate that we have a continuing problem with this infectious disease. Based on the current rate of positive cases seen thus far in 1995, this facility may confirm nearly 900 new cases of HIV infection in this year alone.

The current study evaluated immune response to silicon dioxide in children born to women with silicone breast implants. In part one of the study, the T lymphocytes of 21 of 24 such children were significantly stimulated by silicon dioxide (silica). Part two consisted of eleven children, four born preimplantation and seven born postimplantation. None of the preimplant offspring showed T cell responses to silica; five of the seven postimplant children were positive for T cell memory for silica. Part three was a blinded study based on statistically significant differences in T cell stimulation with silicon dioxide between postimplant children and controls. These findings indicate a common immune reaction, that of T cell memory, occurs in mothers and their children born after exposure to silicone mammary implants placed prior to pregnancy. Since not all such children were breast fed the result favors transplacental passage of immunogens such as silicone oligomers or through maternofetal cellular traffic.

Difficulties in showing the biologic activity of silicones in vitro have contributed to the controversy over effects of silicone mammary implants in vivo. We adapted a standard lymphocyte stimulation test to detect evidence of cellular immunity in patients with silicone gel implants. Initially, lymphocytes were harvested from 70 implant patients, 76 normal controls without implants or symptoms, and 18 patients with classic rheumatic disorders and without a history of silicone implants. The harvested lymphocytes were stimulated with PWM, PHA, Con A, and pharmaceutical grade colloidal silicon dioxide (silica). Implant patients showed increased SI compared to controls and those with rheumatic disorders. The mean SI of implant patients was 195.0 +/- 19.3, 18-fold that of normal controls (< 0.0001). Patients with rheumatic disease showed the same SI as controls (P = 0.3577). A follow-up study included 220 normal controls without implants, 942 silicone gel implant patients with demonstrable rheumatic symptoms, and 34 implant patients without symptoms at the time of the study. The average SI for the 220 normal controls was 10.0 +/- 0.41. Among the symptomatic implant women, 860 (91.3%) had SI significantly above those of the normal controls. Of these, 171 (18.2%) had SI between 25 and 50, 316 (33.5%) had SI between 50 and 100, and 373 (39.6%) had SI over 100. The data presented confirms that silicone implant patients respond immunologically to the silicon dioxide contained in mammary prostheses.

Objective: To compare the performance of the MONO-LEX system to that of the MONO-TEST in detecting infectious mononucleosis (IM) heterophil antibodies., Setting: Baptist Regional Laboratories, Memphis, Tennessee., Practice Description: Not applicable. PRODUCT COMPARISON: The MONO-LEX system, manufactured by Trinity Laboratories, Raleigh, North Carolina, distributed by Gull Laboratories, Salt Lake City, Utah, and the MONO-TEST kit, manufactured by Wampole Laboratories, Cranbury, New Jersey, are agglutination assays that show visible agglutination when reagents are mixed with serum or plasma containing IM heterophil antibodies., Main Outcome Measurements: Accuracy in detecting heterophil antibodies in the study specimens, ease of interpreting test results, and cost-effectiveness., Results: Of the 139 specimens tested, 132 produced the same results using both products and seven produced discrepant results. The possibility that the MONO-TEST produced a "prozone effect" was eliminated. Titration of sera that tested positive for heterophil antibody by both methods revealed titers as high as 1:152 by the MONO-LEX compared with 1:128 by the MONO-TEST. A total of 31 specimens that tested positive for IgM antibodies to Epstein-Barr virus nuclear antigen by enzyme immunoassay were assayed using the two methods; MONO-LEX detected heterophil antibodies in eight sera while the MONO-TEST detected antibody in only four samples., Conclusion: The MONO-LEX surpassed the MONO-TEST in sensitivity, cost-effectiveness, and ease of interpretation.

A total of 469 males over the age of 50 years were tested by the prostate specific antigen (PSA) with follow-up evaluations on those that exceeded 4.0 ng/ml. Thirty six (7.7 percent) of the participants had abnormal results, 23 (63.9 percent) had abnormal digital rectal examinations (DRE) and had biopsies done. Ten of the 23 biopsied participants had positive findings of prostate malignancy and 16 participants had benign prostatic hyperplasia. Three participants with abnormal PSA had negative DRE and no evidence of malignancy by ultrasound. Based on age-specific reference ranges, only one positive biopsy would have been missed while five of the 16 males who had negative biopsies would not have had to undergo this procedure. This would have resulted in a significant savings of medical costs as well as undue worry by the patient.

A new qualitative membrane immunoassay for use in detecting IgG antibodies to cytomegalovirus (CMV) was compared with a standard microplate enzyme immunoassay (EIA). A total of 179 patients were tested by the EIA for IgG- and IgM-specific antibodies to CMV and then compared with the new membrane immunoassay. Among the 179 sera, five specimens were repeatably invalid. Among the 174 evaluable sera, 128 (73.6%) were positive for IgG antibody to CMV by the EIA method. Of these 128 sera, the Transtat-CMV was positive in 124 (96.9%). The remaining 46 sera were negative for IgG antibodies by both methods. The Transtat-CMV membrane immunoassay appears to be a reliable and easily used method not requiring additional equipment or extended incubation times. As a qualitative-only assay, it cannot be used for diagnosis.

With the isolation of the human immunodeficiency virus (HIV), numerous assays have been developed to detect antibodies to the retrovirus. The manufacturers' recommend that all sera exhibiting a positive enzyme immunoassay be confirmed by the more specific Western blot test (WBT). Recently, a new WBT strip was released for detection of antibodies to HIV. Four hundred patients' sera and 22 CDC proficiency samples were each tested with these strips and with a WBT strip prepared within the laboratory using the CDC techniques. The strips were then evaluated for banding pattern with the reactions recorded on a scale of 0 to 4+ for each band consistent with HIV response. Criteria for positive results were based on the ASTPHLD/CDC criteria. Primary emphasis was on antibody banding to gp41 (envelope protein) and p24 (core protein). The new strips showed appropriate changes in level of antibody, especially for declining antibody to p24. When the banding pattern for antibody to gp41 was compared to the standard strips, the new strips showed a sensitivity of 99.6% and specificity of 99.4%. The presence or absence of antibody to p24 on the new strips versus the standard strips showed a sensitivity of 93.7% and specificity of 99.5%.

An enzyme immunoassay that detects IgM and IgG antibodies to the nuclear antigen of Epstein-Barr virus (EBV) is evaluated. The EIA was compared with a rapid monoscreen (Immunoscan-IM) and an indirect fluorescent antibody procedure to determine the correlation of the new test with the current methodology. Among 73 patients tested for heterophilic antibody, 36 were positive; 25 of these had IgM antibodies to nuclear antigen. The sensitivity of the EIA compared to the mono screen was 69.4% and specificity was 97.3%. In comparison with fluorescence assays for IgG to the nuclear antigen, 94 patients were tested with 84 showing positivity by the fluorescent method. Among these, 80 were positive by EIA yielding a sensitivity and specificity of 95.2% and 90.0%, respectively. The EIA does not appear sensitive enough for replacement of the monoscreen but was a sensitive, cost-effective alternative to the indirect fluorescent antibody test.

Pseudomonas paucimobilis, previously known as group IIK biotype 1, was found to have a significantly different enzyme profile from that of Flavobacterium multivorum, formerly group IIK biotype 2. P. paucimobilis produces several esterases and phosphatases which may contribute to its virulence.

The Chemstrip-L test was evaluated in 249 body fluids for its ability to detect leukocytes. Three of 42 cerebrospinal fluids were positive for leukocyte esterases, of which all three were cases of culture-confirmed bacterial meningitis. After the addition of predetermined numbers of neutrophils, the lowest level of detection of neutrophils in cerebrospinal fluids was 40 per microliter. Seventy-five of 200 (38%) peritoneal fluids had positive leukocyte esterase tests. Sixty-six of 72 (89%) peritoneal fluids with 100 or greater neutrophils per microliter had positive leukocyte esterase tests. Protein content, if increased, may interfere with the results of the L-test.

This investigation found that individuals with microagglutination antibody titers of 1:32 or greater to Legionella pneumophila produced bactericidal activity against the bacterium. Those individuals with microagglutination antibody titers of 1:16 or less demonstrated no bactericidal activity.

This investigation was done to determine whether dipicolinic acid was present in the Legionnaires' disease bacterium. A colorimetric assay for dipicolinic acid was done and the results for the bacterium were compared with those obtained for Escherichia coli and Bacillus subtilis. Dipicolinic acid was not detected in the Legionnaires' disease bacterium. This demonstrated that the bacterium was unable to produce typical bacterial endospores under the experimental conditions examined.

We evaluated 230 body fluid samples, of which 131 were peritoneal effluents and 99 were other body fluids. Of these, 63 dialysates were culture positive, and 54 (85.7%) of these 63 were leukocyte esterase positive. Of 99 other body fluids, 8 were both culture positive and leukocyte esterase positive.

Kluyvera, a proposed genus formerly known as Enteric group 8, was found to have similar enzyme profiles among the present 3 groups. Kluyvera species group 3 showed the most heterogeneous enzyme reactions.

In the present investigation, only one of 244 (0.4%) urines with abnormal microscopic findings would have been missed because of normal biochemical results of pH, protein, glucose, ketones, bilirubin, urobilinogen, hemoglobin, and leukocyte esterase. These biochemical tests detected 106 of 107 (99.1%) urines with significant microscopic findings. Sixty-three (25.8%) of 244 urines tested could have been eliminated from microscopic examination based on the negative results of the biochemical screen.

This investigation determined the serum agglutination activity and serum bactericidal response after rabbit immunization with Pseudomonas paucimobilis. Agglutination activity of antisera showed a twofold increase in titer from before immunization to 4 weeks post-immunization and peaked at 8 weeks post-immunization with a titer of 1:512. 2-Mercaptoethanol reduction of immunoglobulin M decreased agglutination titers. No major antigens were found to be common from crude antigen preparations of P. paucimobilis, Pseudomonas aeruginosa and Pseudomonas cepacia when tested with antisera to P. paucimobilis. Serum bactericidal activity was found in post-immunization antisera at 8 and 12 weeks against P. paucimobilis, with no activity present before immunization or at 4 weeks post-immunization. Antisera against P. paucimobilis showed no bactericidal activity against P. aeruginosa or P. cepacia.

The present study evaluated the usefulness of the 1-min leukocyte esterase-nitrite tests in a tertiary-care hospital as a screening procedure to detect significant bacteriuria and correlated the findings with culture results. A total of 531 urine samples were reviewed, of which 484 were evaluated. Of the evaluated samples, 113 positive cultures (23.4%) were found, of which 93 (82.3%) were detected by leukocyte esterase-nitrite tests. In addition, 365 of 371 (98.4%) urine samples with negative bacterial cultures were negative in leukocyte esterase and nitrite tests.

The chromogenic Limulus amoebocyte lysate test effectively detected 66 (100%) culture-proven gram-negative peritonitis cases among 185 continuous ambulatory peritoneal dialysis patients with clinical evidence of infectious peritonitis.