bluuebunny/crossref_metadata_2025_split huggingface dataset:
1191. Evolution of Hepatitis C Virus Points to Postpartum Recovery of CD8+ T-Cell Selection Pressure
Authors: Smith Stephanie , Xu Zhaohui , Walker Christopher , Honegger Jonathan 
Published: January 2021

Abstract

Background
In acute persisting hepatitis C virus (HCV) infection, brief control of viral replication can occur with onset of a cellular immune response but is eventually lost due to selection of viral variants with escape mutations in CD8 T cell epitopes or development of T cell exhaustion. In chronic infection, the CD8s targeting remaining intact epitopes are exhausted, viral levels are stable, and selection of new escape mutations is rare. Postpartum (PP), some chronically infected women experience a drop in viremia which we hypothesize is due to recovery of the CD8 response and will be associated with selection of de novo escape mutations as seen in acute persisting infection.

Methods
HCV genomic evolution was assessed in 2 women with different patterns of PP viral control through consecutive pregnancies. Subject M016 experienced &gt; 2.5 log10 drops in viral loads after each pregnancy while M062 had stable viral levels (Table 1). Longitudinal viral genomes were assessed by Illumina sequencing of near full length PCR products of viral cDNA. Reads were initially aligned to a genotype reference and then iterative consensus sequences until deep coverage was achieved across the entire PCR amplicon. To identify potential CD8 escape mutations, amino acid substitutions were counted if their frequency increased from &lt; 10% of reads peripartum to &gt; 90% at follow up. We focused on the nonstructural region to avoid substitutions related to antibody selection pressure.
Viral Load Levels

Results
M016 developed 3 nonsynonomous (NS) mutations after her 1st pregnancy and 1 more after her 2nd. Of these, 3 were directed away from the consensus amino acid sequence, and one differed from consensus at baseline and changed tangentially. M062 developed 1 NS mutation within a known class I epitope following her second pregnancy which was directed toward consensus for her genotype (Fig 1). All mutations occurred within predicted class I epitopes.
Nonsynonomous Substitutions in Nonstructural Region

Conclusion
While these pilot data require verification in additional subjects, the observed emergence of non-synonymous mutations directed away from genotype consensus in viral class I epitopes of a woman with postpartum viral control but not in a women with stable viral levels support our hypothesis enhanced CD8 T cell selection pressure contribute to enhanced control of HCV replication after childbirth.

Disclosures
All Authors: No reported disclosures
1192. H-NS-like Proteins in Pseudomonas aeruginosa Coordinately Silence Intragenic and Antisense Transcription
Authors: Lippa Andrew M , Gebhardt Michael J , Dove Simon L 
Published: January 2021

Abstract

Background
The H-NS-like proteins MvaT and MvaU act coordinately as global repressors in Pseudomonas aeruginosa by binding to AT-rich regions of the chromosome, which include horizontally acquired genes and numerous virulence factors. Although cells can tolerate the loss of either protein, identifying their combined regulatory effects has been challenging because the loss of both proteins is lethal due to induction of the prophage Pf4 and subsequent superinfection of the cell. Although in other bacteria, H-NS promotes cellular fitness by inhibiting intragenic transcription from AT-rich target regions, preventing them from sequestering RNA polymerase, a role for MvaT and MvaU in repressing transcription from intragenic promoters has not been demonstrated.

Methods
Here we utilize a parental strain that cannot be infected by Pf4 phage to identify the combined MvaT and MvaU regulon. RNA-seq was utilized to identify genes differentially expressed in cells lacking MvaU or both MvaU and MvaT. ChIP-seq was utilized to identify genes directly regulated by MvaT and MvaU in wild-type cells. Further, ChIP-seq was performed in cells of the parental strain and cells lacking both MvaT and MvaU to map genome-wide σ 70-dependent promoters that were active in the presence or absence of both H-NS-like proteins.

Results
We demonstrate that the loss of both MvaT and MvaU, but not MvaU alone, leads to increased sense, antisense, and intragenic transcription from loci directly controlled by these proteins. We further show that the loss of MvaT and MvaU leads to a striking redistribution of RNA polymerase containing σ 70 to those genomic regions vacated by these proteins.
Loss of MvaT and MvaU causes global changes in gene expression

Loss of MvaT and MvaU results in increased sense and antisense transcription

Loss of both MvaT and MvaU results in genome-wide redistribution of RNA polymerase

Conclusion
Our findings suggest that the ability of H-NS-like proteins to repress intragenic transcription is a universal function of these proteins and describe a second mechanism by which MvaT and MvaU may contribute to the growth of P. aeruginosa.

Disclosures
All Authors: No reported disclosures
1194. Identification and Characterization of Extracellular Inducers of Persistence in Staphylococcus epidermidis and Staphylococcus aureus
Authors: Habtu Danni Y , Chamberlain Neal , Curry Elyse , Hart Ryan 
Published: January 2021

Abstract

Background
This study describes the identification and partial characterization of persistence inducing factors (PIF) from Staphylococcus aureus (S. aureus) and Staphylococcus epidermidis (S. epidermidis). Persistence is an epigenetic process that results in tolerance of bacterial cells to antibiotic treatment, which can result in chronic human infections.

Methods
Others have demonstrated a significant increase in persister numbers during mid-log phase. Inducers of this mid-log increase have yet to be identified in staphylococci. Optical density at 600 nm (OD600) was used instead of time to determine when persister numbers increased during logarithmic growth. Concentrated culture filtrates (CCF) from S. epidermidis RP62A and S. aureus SH1000 were obtained at various OD600’s and following incubation at 16 h. The CCF’s were used to develop a persistence inducing factor (PIF) assay. The PIF assay was used to partially characterize PIF from S. epidermidis RP62A and S. aureus SH1000 for relative molecular weight, temperature and protease sensitivity and inter-species communications.

Results
Optimal OD600’s for the S. epidermidis RP62A and S. aureus SH1000 PIF assays were 2.0 and 0.5, respectively. The highest PIF activity for both species was from CCF following incubation overnight (16 h). S. epidermidis RP62A’s PIF activity was decreased by storage at 4o C (2 weeks or longer) but not following incubation at 20o C (16 h), 37o C (1 h) or 100o C (15 min). S. aureus SH1000’s PIF activity was decreased following storage at 4o C (2 week or longer) and after boiling at 100oC for 5 min but not after incubation at 37o C (1 h). PIF activity from both species was less than 3,000 Mrr. Proteinase-K treatment of S. aureus SH1000 PIF decreased activity but did not decrease PIF activity of S. epidermidis RP62A. PIF from S. epidermidis RP62A did not increase persister numbers when used to treat S. aureus SH1000 cells nor did PIF from S. aureus SH1000 increase persister numbers in S. epidermidis RP62A cells.

Conclusion
Previous attempts to discover PIF’s for staphylococcal species were unsuccessful due to the time-based means used to identify mid-log. Both staphylococcal species appear to produce unique, extracellular, low-molecular-weight inducers of persistence (PIF) when assayed using an OD600-based PIF assay.

Disclosures
All Authors: No reported disclosures
93. A Diagnostic Stewardship Intervention to Improve Utilization of 1,3-β-D-glucan Testing
Authors: Kendall Jonathan A , Colson Jordan , Saeed Lyla , Mizusawa Masako , Yamamoto Takeru 
Published: January 2021

Abstract

Background
1,3-β-D-glucan (BDG) is a cell wall component of fungi such as Aspergillus spp., Candida spp., and Pneumocystis jirovecii. BDG assay is used as a screening test to aid early diagnosis of invasive fungal infections (IFI) that are associated with significant morbidity and mortality in immunocompromised patients. The diagnostic performance varies depending on IFI risks among study populations, thus it is important to appropriately select patients with risk factors for IFI to optimize utilization of the BDG test.
Figure 1.

Figure 2.

Methods
An intervention to improve BDG test utilization was initiated at Truman Medical Center on November 28, 2018. The BDG test order was replaced by a BDG test request. The request was sent to the inbox of an on-call pathology team. Patient information was reviewed and the on-call pathology team called the ordering physician to discuss the case based on the approval algorithm chart. The criteria for BDG test approval were 1) immunocompromised or ICU patient, and 2) on empiric antifungal therapy, or inability to perform specific diagnostic tests such as bronchoscopy. If approved, a BDG test order was immediately processed. Retrospective chart review was conducted for 1 year pre- and post- intervention to obtain demographic, clinical, and laboratory data for 4 patient groups. Group 1 included patients who had BDG tests during pre-intervention period. Group 2 was composed of all patients who had BDG test requests during post-intervention period. Group 2a and 2b were the post-intervention patients with approved and rejected BDG test requests, respectively.
Figure 3.

Results
The number of BDG tests performed per year decreased from 156 pre-intervention to 24 post-intervention. The number of test requests was 65 and 41 of them were rejected which led to $7,380 direct cost savings. There was no significant difference in age or the proportion of immunocompromised and ICU patients between Group 1 and 2. The test positivity rate was significantly higher in Group 2-a compared to Group 1 (45.8 % vs. 25.3%, p=0.038). There was no delay in IFI diagnosis or IFI-related mortality in patients for whom BDG test requests were rejected.

Conclusion
We successfully and safely implemented a diagnostic stewardship intervention for BDG testing and improved test utilization.

Disclosures
All Authors: No reported disclosures
1199. Phylogenomic analysis of Campylobacter jejuni isolated from gastroenteritis cases in Michigan
Authors: Rodrigues Jose A , Blankenship Heather M , Cha Wonhee , Mosci Rebekah , Manning Shannon D 
Published: January 2021

Abstract

Background
C. jejuni is the leading cause of bacterial gastroenteritis worldwide. It has been classified as a serious antibiotic resistant threat, causing 13,000 hospitalizations and 120 deaths annually. Our goal was to describe the diversity of clinical C. jejuni using phylogenomics and classify resistance mechanisms.

Methods
Isolates were collected via sentinel surveillance at four hospitals, and demographic and clinical data were obtained. DNA was extracted and sequenced. Raw reads were processed with Trimmomatic and quality checked with FastQC. De novo genome assembly was performed in Spades. Assembled genomes were filtered for quality and completeness; samples of 1.4-2.1MB were annotated in Prokka followed by pangenome and phylogenetic analyses. Multilocus sequence typing loci and virulence and antibiotic resistance genes were extracted from each genome.

Results
Among the 214 C. jejuni isolates recovered, 86 unique sequence types (STs) were identified; five were novel STs with unique allele combinations. ST353 (8.3%: n=18), ST982 (7.4%: n=16), ST50 (5.1 %: n=11) and ST48 (5.1%: n=11) were the most prevalent STs identified, while the majority (50.1%: n=50) of STs were singletons. The pangenome analysis identified 8781, 615, and 1169 total, core, and shell core genes, respectively, which grouped the isolates into three major clades. Most isolates belonged to clade 1. A neighbor-net analysis detected significant recombination among all 86 STs (pairwise homoplasy index p=&lt; 0.00001) and evidence of horizontal gene transfer across clades. The beta-lactamase gene, blaOXA-605, was the most common resistance gene identified (58.8%: n=125) followed by tet(O) (56.0%: n=121), which mediate resistance to beta-lactams and tetracyclines, respectively. Resistance phenotypes were confirmed using microbroth dilution.
Conclusion: Together, these data demonstrate that the C. jejuni population is highly diverse and carries important resistance determinants. The phylogenomic analyses also provide insight into the evolution of this major foodborne pathogen. Future work will focus on identifying molecular and epidemiological factors associated with specific strain types and resistance and virulence profiles circulating in Michigan.

Disclosures
All Authors: No reported disclosures
1200. Risk of group A streptococcal Transmission Among the Pediatric Population in the Houston Area
Authors: Alamarat zain I , Sommer Lauren , McNeil Jonathon C , Flores Anthony R 
Published: January 2021

Abstract

Background
Disease due to group A Streptococcus (GAS) occurs frequently in children and usually manifests as pharyngitis or superficial skin infections. However, invasive disease (iGAS) such as necrotizing fasciitis or streptococcal toxic shock syndrome is responsible for significant morbidity and mortality. National-level surveillance at the Centers for Disease Control and Prevention (CDC) estimates &gt;10,000 cases and ~1,500 deaths due to iGAS occur annually in the US. Much interest revolves around the ability to detect potential transmission events (PTEs) of GAS disease using surveillance data as such information may change recommendations for chemoprophylaxis of close contacts. Studies by the CDC have shown a secondary attack rate from 66.1 to 102 /100,000, primarily occurring among older adults with co-morbidities. However, previous studies were limited in that the GAS surveillance was limited to iGAS disease.

Methods
Retrospective study using a comprehensive GAS passive surveillance system. GAS isolates and associated metadata were obtained from 2 hospital systems in the Texas Medical Center from 2017-2019. Molecular emm typing of GAS isolates was performed using the CDC protocol. PTEs were defined based on GAS disease isolates originating from the same zip code, occurring within 30 days of each other, and of the same emm type.

Results
A total of 1291 isolates were included in the study – 94 PTEs were identified representing 168 individual GAS isolates of which 74 were defined as index cases. The 4 most common GAS emm types identified among PTEs were emm1 (43/94, 45.7%), emm12 (30/94, 31.9%), emm4 (6/94, 6.4%), and emm6 (5/94, 5.3%). Index cases most frequently resulted in a single PTE (n=74) with an average number of PTEs per index case of 1.3 (range 1 to 3 PTEs). From index cases, 10 GAS isolates were derived from invasive disease (10/74, 13.5%) and 6 from skin and soft tissue infections (SSTI; 6/74, 8.1%). A substantial proportion of PTEs resulted in iGAS (9/94, 9.5%) and SSTI (10/94, 10.6%).

Conclusion
Using comprehensive local surveillance, we were able to identify several potential GAS transmission events. Further analysis – including whole genome sequencing on index and PTE isolates – is needed to better define transmission events.

Disclosures
Jonathon C. McNeil, MD, Nabriva (Research Grant or Support, I serve as the site investigator on a multicenter clinical trial sponsored by Nabriva)
1201. Sarcoidosis Candidate Microbes Identified by Next Generation Sequencing
Authors: Kriesel John D , Eckman Emily , Emerson Lyska , Scholand Marybeth , Hoidal John , Fischer Kael 
Published: January 2021

Abstract

Background
Sarcoidosis is an autoimmune disease characterized by granulomatous lung disease with very prominent mediastinal adenopathy. Acid-fast bacteria, fungi, and viruses have been considered as possible causes of sarcoidosis. We used next-generation or deep sequencing to characterize the microbial content of diseased mediastinal lymph nodes from 10 sarcoidosis patients compared to a set of 10 negative-controls.

Methods
RNA was extracted from fixed paraffinized mediastinal lymph nodes (MLN) from 12 diseased specimens taken from 10 sarcoidosis patients and 2 positive control subjects (TB, MAI), and normal appearing MLN from 10 negative-control subjects (mostly cancer patients). The extracted RNA was sequenced on the Illumina 2500, yielding 125-bp paired-end reads. These reads were aligned to the human genome, human transcriptome, and a nonredundant panmicrobial database. Each experimental sample were compared against the set of 10 negative-controls using the false discovery rate method (q-value). Directed qPCR was performed on all the samples.

Results
100-153 million read-pairs were obtained from the 24 sequenced samples (12 sarcoidosis, 10 negative-control, 2 positive-control). Among these, 0.01-1.32% of the reads were microbial, with a trend towards fewer microbial reads in the sarcoidosis group compared to controls (means 66K vs. 457K, p=0.09). Mycobacterial sequence was significantly enriched (q&lt; 0.05) in the MAI but not the TB sample compared to the negative-controls. Among the 12 sarcoidosis samples, sequence mappings were significantly enriched (q&lt; 0.05) for the following genera: fungal, Magnaporthe (N=4 samples) and Debaromyces (1); bacteria, Odoribacter (1) and Granulicella (1); and viral, Roseolovirus (6) and Mardivirus (6). Further metagenomic analysis eliminated Magnaporthe as a candidate. qPCR confirmed the presence of Odoribacter in 2 specimens and Debaromyces in 1. Roseolovirus (HHV6) could not be detected by qPCR in any of the samples.

Conclusion
We conclude that sequencing is a feasible method for identifying candidate microbes that might trigger sarcoidosis in human subjects. Further research is required to establish or refute the pathogenicity of these organisms in patients with sarcoidosis.

Disclosures
All Authors: No reported disclosures
1202. Subinhibitory Concentrations of Omadacycline Inhibit Staphylococcus aureus Hemolytic Activity in Vitro
Authors: Serio Alisa W , Tanaka S Ken , Wright Kelly , Garrity-Ryan Lynne 
Published: January 2021

Abstract

Background
In animal models of Staphylococcus aureus infection, α-hemolysin has been shown to be a key virulence factor. Treatment of S. aureus with subinhibitory levels of protein synthesis inhibitors can decrease α-hemolysin expression. Omadacycline, a novel aminomethylcycline antibiotic in the tetracycline class of bacterial protein biosynthesis inhibitors, is approved in the United States for treatment of community-acquired bacterial pneumonia (CABP) and acute bacterial skin and skin structure infections (ABSSSI) in adults. This study was performed to determine the durability of inhibition and effect of subinhibitory concentrations of omadacycline on S. aureus hemolytic activity.

Methods
All experiments used the methicillin-sensitive S. aureus strain Wood 46 (ATCC 10832), a laboratory strain known to secrete high levels of α-hemolysin. Minimum inhibitory concentrations (MICs) of omadacycline and comparator antibiotics (tetracycline, cephalothin, clindamycin, vancomycin, linezolid) were determined. Growth of S. aureus with all antibiotics was determined and the percentage of hemolysis assayed. “Washout” experiments were performed with omadacycline only.

Results
S. aureus cultures treated with 1/2 or 1/4 the MIC of omadacycline for 4 hours showed hemolysis units/108 CFU of 47% and 59% of vehicle-treated cultures, respectively (Fig. 1A, 1B). In washout experiments, treatment with as little as 1/4 the MIC of omadacycline for 1 hour decreased the hemolysis units/108 CFU by 60% for 4 hours following removal of the drug (Table 1).
Figure 1

Table 1

Conclusion
Omadacycline inhibited S. aureus hemolytic activity in vitro at subinhibitory concentrations and inhibition was maintained for ≥ 4 hours after removal of extracellular drug (Fig. 2). The suppression of virulence factors throughout the approved omadacycline dosing interval, in addition to the in vitro potency of omadacycline, may contribute to the efficacy of omadacycline for ABSSSI and CABP due to virulent S. aureus. This finding may apply to other organisms and other virulence factors that require new protein synthesis to establish disease.
Figure 2

Disclosures
Alisa W. Serio, PhD, Paratek Pharmaceuticals, Inc. (Employee, Shareholder) S. Ken Tanaka, PhD, Paratek Pharmaceuticals, Inc. (Employee, Shareholder) Kelly Wright, PharmD, Paratek Pharmaceuticals, Inc. (Employee, Shareholder) Lynne Garrity-Ryan, PhD, Paratek Pharmaceuticals, Inc. (Employee, Shareholder)
1203. Systemic, Mucosal Immune Activation And Psycho-sexual Health in HIV-Infected And Uninfected Women: Evaluation of Biomarkers And Environmental Stimuli
Authors: Nijhawan Parni , Nijhawan Parni , Carraro Anna , Marocco raffaella , Vita Serena , Kertusha Blerta , Mascia Claudia , Lichtner Miriam , Mastroianni Claudio , Zingaropoli Maria Antonella , Masci Georgio , Ianiri Palmieri , Vito Mercurio , Fondaco Laura , Del Borgo Cosmo , Mirella D’Achille 
Published: January 2021

Abstract

Background
HIV infection in women in disproportionate ratios as compared to men has been a grave concern over the years. It is in proportion to reproductive and hormonal differences making women more vulnerable. It elicits an Immune response which can be monitored by analysing various factors such as mucosal immunity, sexual behaviour, biomarkers in the plasma, serum and vaginal lavage and vaginal infections.

Aim
Evaluating and comparing the systemic and mucosal immuno-inflammatory status, the female sexual function (FSF) and generalized anxiety in HIV+ women on successful HAART with healthy women (HW).

Methods
We enrolled 53 subjects (23 HIV+ women on successful HAART and 30 Healthy women (HW)) with no statistical differences in age. The figure (named: methodology) below explains the methods applied:
Cytometry and Kit ELISA were used to estimate lymphocytes and all cytokines. Women were also tested for co-morbidities such as diabetes, blood pressure, HCV, cervical cancer etc. Statistical analysis was performed using PRISM 8.0.
Methodology

Results
Higher CD4 and CD8 cell count was observed in HW compared to HIV+ women (p=0.02,p=0.004).Plasma levels of sCD 163, CXCL-10, IL-1, IL-6 and IL-8 were significantly higher in HIV women as compared to HW(p&lt; 0.001), while IL-6 and IL8 were lower in the VL of HIV women. An ASCUS in HW was found for PAP Test. CXCL-10 was correlated to estradiol levels (r=0.8, p=0.02). 57% reported FSD and 43% had a FSFI score≤10. A significant difference between the two groups in the FSFI score (p=0.007) was found, particularly in sexual desire, arousal and pain. A positive correlation between level of testosterone and FSFI score was found only in HIV+ women (p=0.02; r= 0.74). 17% of women presented an anxiety disorder. Z-index was associated with orgasm domains (p=0.01; r=-0.4) and CD4+ T cells (p=0.02; r=-0.45).

Conclusion
Higher plasma levels of the cytokines despite successful antiretroviral therapy were observed. At the mucosal level evaluating the balance within pro anti-inflammatory cytokines and micro-biome will be interesting to study. FSD is detected in more than half of HIV infected women and seems to be related to testosterone levels. The comparison with uninfected women underlying a persistent gap in quality of life of young HIV women should be bridged.

Disclosures
All Authors: No reported disclosures