Objectives:
Public health laboratories (PHLs) are essential components of US Public Health Service operations. The health information technology that supports PHLs is central to effective and ...efficient laboratory operations and overall public health response to infectious disease management. This analysis presents key information on how the Nebraska Public Health Laboratory (NPHL) information technology system evolved to meet the demands of the COVID-19 pandemic.
Materials and Methods:
COVID-19 presented numerous, unforeseen information technology system challenges. The most notable challenges requiring changes to NPHL software systems and capability were improving efficiency of the laboratory operation due to high-volume testing, responding daily to demands for timely data for analysis by partner systems, interfacing with multiple testing (equipment) platforms, and supporting community-based specimen collection programs.
Results:
Improvements to the NPHL information technology system enabled NPHL to perform >121 000 SARS-CoV-2 polymerase chain reaction tests from March 2020 through January 2022 at a sustainable rate of 2000 SARS-CoV-2 tests per day, with no increase in laboratory staffing. Electronic reporting of 62 000 rapid antigen tests eliminated paper reporting and extended testing services throughout the state. Collection of COVID-19 symptom data before specimen collection enabled NPHL to make data-driven decisions to perform pool testing and conserve testing kits when supplies were low.
Practice Implications:
NPHL information technology applications proved essential for managing health care provider workload, prioritizing the use of scarce testing supplies, and managing Nebraska’s overall pandemic response. The NPHL experience provides useful examples of a highly capable information technology system and suggests areas for additional attention in the PHL environment, including a focus on end users, collaboration with various partners, and investment in information technology.
Summary This study investigated the diagnostic accuracy of whole slide imaging (WSI) in breast needle biopsy diagnosis in comparison with standard light microscopy (LM). The study examined the ...effects of image capture magnification and computer monitor quality on diagnostic concordance of WSI and LM. Four pathologists rendered diagnoses using WSI to examine 85 breast biopsies (92 parts; 786 slides) consisting of benign and malignant cases. Each WSI case was evaluated using images captured at either ×20 or ×40 magnifications and viewed using a Digital Imaging and Communication in Medicine (DICOM) grade, color-calibrated monitor or a standard, desktop liquid-crystal display (LCD) monitor. For each combination, the WSI result was compared with the original, LM diagnosis. The overall concordance rate observed between WSI and LM was 97.1% (95% confidence intervals CI: 94.3%-98.5%). After a washout period, all cases were reviewed a second time by each pathologist after using LM, and the second LM diagnosis was compared with the WSI diagnosis rendered by the same pathologist. Intraobserver concordance between WSI and LM was 95.4% (95% CI: 92.2%-97.4%). The second LM diagnoses were also compared with the original LM diagnoses, and the observed interobserver LM concordance rate was 97.3% (95% CI: 93.1%-99.0%). The study data demonstrated that breast needle biopsy diagnoses rendered by WSI were equivalent to diagnoses rendered by LM. No diagnostic differences were detected between the underlying viewing system parameters of monitor quality and image capture resolution. The results of this study demonstrated that WSI can be effectively used in subspecialty diagnostic cases where a minimum amount of tissue is available.
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DNA-dependent RNA primase is essential for de novo primer synthesis during DNA replication in all living organisms. Bacterial DnaG primase is an attractive target for inhibition ...because it is essential, low in copy number and structurally distinct from eukaryotic and archaeal primases. DnaG primase is sensitive to known inhibitors including suramin and doxorubicin. Recently, tilorone was discovered by high throughput screening to be an inhibitor of Bacillus anthracis primase DnaG but it failed to reduce the growth of B. anthracis in vitro. In this study we determined that tilorone also inhibited DnaG primase from Staphylococcus aureus. C2-Symmetric fluorenone-based compounds, similar to tilorone chemical structure were synthesized and tested to identify potential lead compounds that inhibit bacterial growth in B. anthracis, MRSA and Burkholderia thailandensis. These compounds were evaluated by determining the minimum inhibitory concentration (MIC) against several different bacterial species which demonstrated 17.5 and 16μg/ml MIC profiles. Importantly, some of the fluorenone-based compounds with a long carbon chain showed a relatively low MIC against B. anthracis, S. aureus, MRSA, Francisella tularensis, and B. thailandensis, suggesting it may be a promising lead compound.
Vaccination is the most effective intervention to prevent influenza and control the spread of the virus. Alternatives are needed to the traditional egg-based vaccine strategy for a more rapid ...response to new outbreaks. Two different hemagglutinin (HA) fragments (rHA1
and rHA1
) derived from influenza A virus subtype H1N1 were expressed in Escherichia coli and characterized by immunoblot, gel filtration, hemagglutination, and competitive binding assays. rHA1
included neutralizing epitopes and the trimerization domain, whereas rHA1
included only the head of HA with the neutralizing epitopes. Mice were immunized with rHA1
or rHA1
, and sera were tested for the presence of neutralizing antibodies. Mice were then challenged with H1N1 and infection severity was monitored. rHA1
trimerized, whereas rHA1
was unable to form oligomers. Both rHA1
and rHA1
elicited the production of neutralizing antibodies, but only oligomerized rHA1
protected against live virus challenges in mice. This study demonstrated that bacterially expressed HA was capable of folding properly and eliciting the production of neutralizing antibodies, and that HA oligomerization contributed to protection against viral challenge. Therefore, prokaryotic-derived vaccine platforms can provide antigenic and structural requirements for viral protection, as well as allow for the rapid and cost-effective incorporation of multiple antigens for broader protection.
Human albumin is thought to hydrolyze esters because multiple equivalents of product are formed for each equivalent of albumin. Esterase activity with p-nitrophenyl acetate has been attributed to ...turnover at tyrosine 411. However, p-nitrophenyl acetate creates multiple, stable, acetylated adducts, a property contrary to turnover. Our goal was to identify residues that become acetylated by p-nitrophenyl acetate and determine the relationship between stable adduct formation and turnover. Fatty acid-free human albumin was treated with 0.5 mM p-nitrophenyl acetate for 5 min to 2 weeks, or with 10 mM p-nitrophenyl acetate for 48 h to 2 weeks. Aliquots were digested with pepsin, trypsin, or GluC and analyzed by mass spectrometry to identify labeled residues. Only Tyr-411 was acetylated within the first 5 min of reaction with 0.5 mM p-nitrophenyl acetate. After 0.5-6 h there was partial acetylation of 16-17 residues including Asp-1, Lys-4, Lys-12, Tyr-411, Lys-413, and Lys-414. Treatment with 10 mM p-nitrophenyl acetate resulted in acetylation of 59 lysines, 10 serines, 8 threonines, 4 tyrosines, and Asp-1. When Tyr-411 was blocked with diisopropylfluorophosphate or chlorpyrifos oxon, albumin had normal esterase activity with β-naphthyl acetate as visualized on a nondenaturing gel. However, after 82 residues had been acetylated, esterase activity was almost completely inhibited. The half-life for deacetylation of Tyr-411 at pH 8.0, 22 °C was 61 ± 4 h. Acetylated lysines formed adducts that were even more stable. In conclusion, the pseudo-esterase activity of albumin is the result of irreversible acetylation of 82 residues and is not the result of turnover.
Ebola virus (EBOV), species Zaire ebolavirus, may persist in the semen of male survivors of Ebola virus disease (EVD). We conducted a study of male survivors of the 2014-2016 EVD outbreak in Liberia ...and evaluated their immune responses to EBOV. We report here findings from the serologic testing of blood for EBOV-specific antibodies, molecular testing for EBOV in blood and semen, and serologic testing of peripheral blood mononuclear cells (PBMCs) in a subset of study participants.
We tested for EBOV RNA in blood by quantitative reverse transcription polymerase chain reaction (qRT-PCR), and for anti-EBOV-specific immunoglobulin M (IgM) and immunoglobulin G (IgG) antibodies by enzyme-linked immunosorbent assay (ELISA) for 126 study participants. We performed PBMC analysis on a subgroup of 26 IgG-negative participants.
All 126 participants tested negative for EBOV RNA in blood by qRT-PCR. The blood of 26 participants tested negative for EBOV-specific IgG antibodies by ELISA. PBMCs were collected from 23/26 EBOV IgG-negative participants. Of these, 1/23 participants had PBMCs that produced anti-EBOV-specific IgG antibodies upon stimulation with EBOV-specific glycoprotein (GP) and nucleoprotein (NP) antigens.
The blood of EVD survivors, collected when they did not have symptoms meeting the case definition for acute or relapsed EVD, is unlikely to pose a risk for EBOV transmission. We identified 1 IgM/IgG negative participant who had PBMCs that produced anti-EBOV-specific antibodies upon stimulation. Immunogenicity following acute EBOV infection may exist along a spectrum, and absence of antibody response should not be exclusionary in determining an individual's status as a survivor of EVD.
Our goal was to determine whether chlorpyrifos oxon, dichlorvos, diisopropylfluorophosphate (DFP), and sarin covalently bind to human albumin. Human albumin or plasma was treated with ...organophosphorus (OP) agent at alkaline pH, digested with pepsin at pH 2.3, and analyzed by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. Two singly charged peaks
m/
z 1718 and 1831, corresponding to the unlabeled peptide fragments containing the active site Tyr411 residue, were detected in all samples. The sequences of the two peptides were VRYTKKVPQVSTPTL and LVRYTKKVPQVSTPTL. The peptide–OP adducts of these peptides were also found. They had masses of 1854 and 1967 for chlorpyrifos oxon, 1825 and 1938 for dichlorvos, 1881 and 1994 for DFP, and 1838 and 1938 for sarin; these masses fit a mechanism whereby OP bound covalently to Tyr411. The binding of DFP to Tyr411 of human albumin was confirmed by electrospray tandem mass spectrometry and analysis of product ions. None of the OP–albumin adducts lost an alkoxy group, leading to the conclusion that aging did not occur. Our results show that OP pesticides and nerve agents bind covalently to human albumin at Tyr411. The presence of Tyr411 on an exposed surface of albumin suggests that an antibody response could be generated against OP–albumin adducts.
Shiga-toxin producing Escherichia coli (STEC) O26:H11 is the second most common cause of severe diarrhea and hemolytic uremic syndrome worldwide. The implementation of whole genome sequencing (WGS) ...enhances the detection and in-depth characterization of these non-O157 STEC strains. The aim of this study was to compare WGS to phenotypic serotyping and pulse field gel electrophoresis (PFGE) for characterization of STECO26 strains following a zoonotic outbreak from cattle to humans.
This study evaluated seven E. coli strains; two strains isolated from two children with gastrointestinal symptoms and five strains from five calves suspected as the source of infection. Six of these isolates were serotyped phenotypically and by WGS as E. coli O26:H11 while one bovine isolate could be serotyped only by WGS as E. coli O182:H25. Stx1 was detected in two human- and two bovine-isolates using PCR and WGS. Using WGS, all four STECO26 isolates belong to sequence type (ST) 21 while the two stx1 negative E. coli O26 were ST29. All four STECO26 isolates were indistinguishable by PFGE. However, the data generated by WGS linked the two human STECO26 isolates to only one bovine STECO26 strain by having identical high-quality single nucleotide polymorphisms (hqSNPs) and identical virulence factor profiles while the remaining bovine STECO26 isolate differed by 7 hqSNPs and lacked virulence factor toxB.
These data demonstrated that WGS provided significant information beyond traditional epidemiological tools allowing for comprehensive characterization of the STEC. Using this approach, WGS was able to identify the specific source of infection in this study.
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