Correction: Lyons-Weiler, J., et al. Relative Incidence of Office Visits and Cumulative Rates of Billed Diagnoses along the Axis of Vaccination. Int. J. Environ. Res. Public Health 2020, 17, 8674.

There were two errors in the original article [...].

Relative Incidence of Office Visits and Cumulative Rates of Billed Diagnoses Along the Axis of Vaccination.

We performed a retrospective analysis spanning ten years of pediatric practice focused on patients with variable vaccination born into a practice, presenting a unique opportunity to study the effects of variable vaccination on outcomes. The average total incidence of billed office visits per outcome related to the outcomes were compared across groups (Relative Incidence of Office Visit (RIOV)). RIOV is shown to be more powerful than odds ratio of diagnoses. Full cohort, cumulative incidence analyses, matched for days of care, and matched for family history analyses were conducted across quantiles of vaccine uptake. Increased office visits related to many diagnoses were robust to days-of-care-matched analyses, family history, gender block, age block, and false discovery risk. Many outcomes had high RIOV odds ratios after matching for days-of-care (e.g., anemia (6.334), asthma (3.496), allergic rhinitis (6.479), and sinusitis (3.529), all significant under the Z-test). Developmental disorders were determined to be difficult to study due to extremely low prevalence in the practice, potentially attributable to high rates of vaccine cessation upon adverse events and family history of autoimmunity. Remarkably, zero of the 561 unvaccinated patients in the study had attention deficit hyperactivity disorder (ADHD) compared to 0.063% of the (partially and fully) vaccinated. The implications of these results for the net public health effects of whole-population vaccination and with respect for informed consent on human health are compelling. Our results give agency to calls for research conducted by individuals who are independent of any funding sources related to the vaccine industry. While the low rates of developmental disorders prevented sufficiently powered hypothesis testing, it is notable that the overall rate of autism spectrum disorder (0.84%) in the cohort is half that of the US national rate (1.69%). The practice-wide rate of ADHD was roughly half of the national rate. The data indicate that unvaccinated children in the practice are not unhealthier than the vaccinated and indeed the overall results may indicate that the unvaccinated pediatric patients in this practice are healthier overall than the vaccinated.

Prediction of severity and subtype of fibrosing disease using model informed by inflammation and extracellular matrix gene index.

Fibrosis is a chronic disease with heterogeneous clinical presentation, rate of progression, and occurrence of comorbidities. Systemic sclerosis (scleroderma, SSc) is a rare rheumatic autoimmune disease that encompasses several aspects of fibrosis, including highly variable fibrotic manifestation and rate of progression. The development of effective treatments is limited by these variabilities. The fibrotic response is characterized by both chronic inflammation and extracellular remodeling. Therefore, there is a need for improved understanding of which inflammation-related genes contribute to the ongoing turnover of extracellular matrix that accompanies disease. We have developed a multi-tiered method using Naïve Bayes modeling that is capable of predicting level of disease and clinical assessment of patients based on expression of a curated 60-gene panel that profiles inflammation and extracellular matrix production in the fibrotic disease state. Our novel modeling design, incorporating global and parametric-based methods, was highly accurate in distinguishing between severity groups, highlighting the importance of these genes in disease. We refined this gene set to a 12-gene index that can accurately identify SSc patient disease state subsets and informs knowledge of the central regulatory pathways in disease progression.

Impact of catch-up vaccination on aluminum exposure due to new laws and post social distancing.

BACKGROUND: The COVID-19 pandemic has placed significant stressors on the medical community and on the general public. Part of this includes patients skipping well-child visits to reduce risk of exposure to SARS-CoV-2 virus. Published estimates of the duration of whole-body aluminum (Al) toxicity from vaccines in infants from birth to six months indicate that CDC's recommended vaccination schedule leads to unacceptably long periods of time in which infants are in aluminum toxicity (as measured by %AlumTox). METHODS: We utilize these established clearance and accumulation models to calculate expected per-body-weight whole-body toxicity of aluminum from vaccines considering for children of all ages under CDC's Catch-Up schedule from birth to ten years, assuming social distancing for 6 months. Our updated Pediatric Dose Limit (PDL) model assumes a linear improvement in renal function from birth to two years. RESULTS: Our results indicate that due diligence in considering alternative spacing and use of non-aluminum containing vaccines when possible will reduce whole body toxicity and may reduce risk of morbidity associated with exposure to aluminum. CONCLUSIONS: While reduction or elimination of aluminum exposure from all sources is always a good idea, our results indicate that careful consideration of expected aluminum exposures during regular and Catch-Up vaccination is found to be especially important for infants and children below 2 years of age. We urge caution in the mass re-starting of vaccination under CDC's Catch-Up schedule for children under 12 months and offer alternative strategies to minimize per-day/week/month exposure to aluminum hydroxide following the COVID-19 period of isolation.

Pathogenic priming likely contributes to serious and critical illness and mortality in COVID-19 via autoimmunity.

Homology between human and viral proteins is an established factor in viral- or vaccine-induced autoimmunity. Failure of SARS and MERS vaccines in animal trials involved pathogenesis consistent with an immunological priming that could involve autoimmunity in lung tissues due to previous exposure to the SARS and MERS spike protein. Exposure pathogenesis to SARS-CoV-2 in COVID-19 likely will lead to similar outcomes. Immunogenic peptides in viruses or bacteria that match human proteins are good candidates for pathogenic priming peptides (similar to the more diffuse idea of "immune enhancement"). Here I provide an assessment of potential for human pathogenesis via autoimmunity via exposure, via infection or injection. SAR-CoV-2 spike proteins, and all other SARS-CoV-2 proteins, immunogenic epitopes in each SARS-CoV-2 protein were compared to human proteins in search of high local homologous matching. Only one immunogenic epitope in a SARS-CoV-2 had no homology to human proteins. If all of the parts of the epitopes that are homologous to human proteins are excluded from consideration due to risk of pathogenic priming, the remaining immunogenic parts of the epitopes may be still immunogenic and remain as potentially viable candidates for vaccine development. Mapping of the genes encoding human protein matches to pathways point to targets that could explain the observed presentation of symptoms in COVID-19 disease. It also strongly points to a large number of opportunities for expected disturbances in the immune system itself, targeting elements of MHC Class I and Class II antigen presentation, PD-1 signaling, cross-presentation of soluble exogenous antigens and the ER-Phagosome pathway. Translational consequences of these findings are explored.

Acute exposure and chronic retention of aluminum in three vaccine schedules and effects of genetic and environmental variation.

Like the mechanisms of action as adjuvants, the pharmacodynamics of injected forms of aluminum commonly used in vaccines are not well-characterized, particularly with respect to how differences in schedules impact accumulation and how factors such as genetics and environmental influences on detoxification influence clearance. Previous modeling efforts are based on very little empirical data, with the model by Priest based on whole-body clearance rates estimated from a study involving a single human subject. In this analysis, we explore the expected acute exposures and longer-term whole-body accumulation/clearance across three vaccination schedules: the current US Centers for Disease Control and Prevention (CDC) schedule, the current CDC schedule using low aluminum or no aluminum vaccines, and Dr. Paul Thomas' "Vaccine Friendly Plan" schedule. We then study the effects of an implicit assumption of the Priest model on whether clearance dynamics from successive doses are influenced by the current level of aluminum or modeled by the assumption that a new dose has its own whole-body dynamics "reset" on the day of injection. We model two additional factors: variation (deficiency) in aluminum detoxification, and a factor added to the Priest equation to model the potential impact of aluminum itself on cellular and whole-body detoxification. These explorations are compared to a previously estimated pediatric dose limit (PDL) of whole-body aluminum exposure and provide a new statistic: %alumTox, the (expected) percentage of days (or weeks) an infant is in aluminum toxicity, reflecting chronic toxicity. We show that among three schedules, the CDC schedule results in the highest %alumTox regardless of model assumptions, and the Vaccine Friendly Plan schedule, which avoids >1 ACV per office visit results in the lowest (expected) %alumTox. These results are conservative, as the MSL is derived from data used by FDA to estimate safety of aluminum in adult humans. These results demonstrate high potential utility of modeling variation in patient responses to aluminum. More empirical data from individuals who are suspected of being intolerant of aluminum from vaccines, evidenced by high aluminum retention, neurodevelopmental disorders and/or a myriad of chronic illnesses would help answer questions on whether the model predictions can be used to estimate parameter values tied to genetic factors including genomic sequence variation and family history of chronic illnesses tied to aluminum exposure.

Reconsideration of the immunotherapeutic pediatric safe dose levels of aluminum.

FDA regulations require safety testing of constituent ingredients in drugs (21 CFR 610.15). With the exception of extraneous proteins, no component safety testing is required for vaccines or vaccine schedules. The dosing of aluminum in vaccines is based on the production of antibody titers, not safety science. Here we estimate a Pediatric Dose Limit that considers body weight. We identify several serious historical missteps in past analyses of provisional safe levels of aluminum in vaccines, and provide updates relevant to infant aluminum exposure in the pediatric schedule considering pediatric body weight. When aluminum doses are estimated from Federal Regulatory Code given body weight, exposure from the current vaccine schedule are found to exceed our estimate of a weight-corrected Pediatric Dose Limit. Our calculations show that the levels of aluminum suggested by the currently used limits place infants at risk of acute, repeated, and possibly chronic exposures of toxic levels of aluminum in modern vaccine schedules. Individual adult exposures are on par with Provisional Tolerable Weekly Intake "limits", but some individuals may be aluminum intolerant due to genetics or previous exposures. Vaccination in neonates and low birth-weight infants must be re-assessed; other implications for the use of aluminum-containing vaccines, and additional limitations in our understanding of neurotoxicity and safety levels of aluminum in biologics are discussed.

Interleukin-17 limits hypoxia-inducible factor 1α and development of hypoxic granulomas during tuberculosis.

Mycobacterium tuberculosis (Mtb) is a global health threat, compounded by the emergence of drug-resistant strains. A hallmark of pulmonary tuberculosis (TB) is the formation of hypoxic necrotic granulomas, which upon disintegration, release infectious Mtb. Furthermore, hypoxic necrotic granulomas are associated with increased disease severity and provide a niche for drug-resistant Mtb. However, the host immune responses that promote the development of hypoxic TB granulomas are not well described. Using a necrotic Mtb mouse model, we show that loss of Mtb virulence factors, such as phenolic glycolipids, decreases the production of the proinflammatory cytokine IL-17 (also referred to as IL-17A). IL-17 production negatively regulates the development of hypoxic TB granulomas by limiting the expression of the transcription factor hypoxia-inducible factor 1α (HIF1α). In human TB patients, HIF1α mRNA expression is increased. Through genotyping and association analyses in human samples, we identified a link between the single nucleotide polymorphism rs2275913 in the IL-17 promoter (-197G/G), which is associated with decreased IL-17 production upon stimulation with Mtb cell wall. Together, our data highlight a potentially novel role for IL-17 in limiting the development of hypoxic necrotic granulomas and reducing disease severity in TB.

Altered DNA methylation in PAH deficient phenylketonuria.

While phenylalanine (PHE) is the toxic insult in phenylketonuria (PKU), mechanisms underlying PHE toxicity remain ill-defined. Altered DNA methylation in response to toxic exposures is well-recognized. DNA methylation patterns were assessed in blood and brain from PKU patients to determine if PHE toxicity impacts methylation. Methylome assessment, utilizing methylated DNA immunoprecipitation and paired-end sequencing, was performed in DNA obtained from brain tissue of classical PKU patients, leukocytes from poorly controlled PKU patients, leukocytes from well controlled PKU patients, and appropriate control tissues. In PKU brain tissue, expression analysis determined the impact of methylation on gene function. Differential methylation was observed in brain tissue of PKU patients and expression studies identified downstream impact on gene expression. Altered patterns of methylation were observed in leukocytes of well controlled and poorly controlled patients with more extensive methylation in patients with high PHE exposure. Differential methylation of noncoding RNA genes was extensive in patients with high PHE exposure but minimal in well controlled patients. Methylome repatterning leading to altered gene expression was present in brain tissue of PKU patients, suggesting a role in neuropathology. Aberrant methylation is observed in leukocytes of PKU patients and is influenced by PHE exposure. DNA methylation may provide a biomarker relating to historic PHE exposure.

Expression of three topologically distinct membrane proteins elicits unique stress response pathways in the yeast Saccharomyces cerevisiae.

Misfolded membrane proteins are retained in the endoplasmic reticulum (ER) and are subject to ER-associated degradation, which clears the secretory pathway of potentially toxic species. While the transcriptional response to environmental stressors has been extensively studied, limited data exist describing the cellular response to misfolded membrane proteins. To this end, we expressed and then compared the transcriptional profiles elicited by the synthesis of three ER retained, misfolded ion channels: The α-subunit of the epithelial sodium channel, ENaC, the cystic fibrosis transmembrane conductance regulator, CFTR, and an inwardly rectifying potassium channel, Kir2.1, which vary in their mass, membrane topologies, and quaternary structures. To examine transcriptional profiles in a null background, the proteins were expressed in yeast, which was previously used to examine the degradation requirements for each substrate. Surprisingly, the proteins failed to induce a canonical unfolded protein response or heat shock response, although messages encoding several cytosolic and ER lumenal protein folding factors rose when αENaC or CFTR was expressed. In contrast, the levels of these genes were unaltered by Kir2.1 expression; instead, the yeast iron regulon was activated. Nevertheless, a significant number of genes that respond to various environmental stressors were upregulated by all three substrates, and compared with previous microarray data we deduced the existence of a group of genes that reflect a novel misfolded membrane protein response. These data indicate that aberrant proteins in the ER elicit profound yet unique cellular responses.

Direct regulation of diurnal Drd3 expression and cocaine reward by NPAS2.

BACKGROUND: Circadian gene disruptions are associated with the development of psychiatric disorders, including addiction. However, the mechanisms by which circadian genes regulate reward remain poorly understood. METHODS: We used mice with a mutation in Npas2 and adeno-associated virus-short hairpin RNA mediated knockdown of Npas2 and Clock in the nucleus accumbens (NAc). We performed conditioned place preference assays. We utilized cell sorting quantitative real-time polymerase chain reaction, and chromatin immunoprecipitation followed by deep sequencing. RESULTS: Npas2 mutants exhibit decreased sensitivity to cocaine reward, which is recapitulated with a knockdown of neuronal PAS domain protein 2 (NPAS2) specifically in the NAc, demonstrating the importance of NPAS2 in this region. Interestingly, reducing circadian locomotor output cycles kaput (CLOCK) (a homologue of NPAS2) in the NAc had no effect, suggesting an important distinction in NPAS2 and CLOCK function. Furthermore, we found that NPAS2 expression is restricted to Drd1 expressing neurons while CLOCK is ubiquitous. Moreover, NPAS2 and CLOCK have distinct temporal patterns of DNA binding, and we identified novel and unique binding sites for each protein. We identified the Drd3 dopamine receptor as a direct transcriptional target of NPAS2 and found that NPAS2 knockdown in the NAc disrupts its diurnal rhythm in expression. Chronic cocaine treatment likewise disrupts the normal rhythm in Npas2 and Drd3 expression in the NAc, which may underlie behavioral plasticity in response to cocaine. CONCLUSIONS: Together, these findings identify an important role for the circadian protein, NPAS2, in the NAc in the regulation of dopamine receptor expression and drug reward.

Differential hippocampal gene expression and pathway analysis in an etiology-based mouse model of major depressive disorder.

We have recently reported the creation and initial characterization of an etiology-based recombinant mouse model of a severe and inherited form of Major Depressive Disorder (MDD). This was achieved by replacing the corresponding mouse DNA sequence with a 6-base DNA sequence from the human CREB1 promoter that is associated with MDD in individuals from families with recurrent, early-onset MDD (RE-MDD). In the current study, we explored the effect of the pathogenic Creb1 allele on gene expression in the mouse hippocampus, a brain region that is altered in structure and function in MDD. Mouse whole-genome profiling was performed using the Illumina MouseWG-6 v2.0 Expression BeadChip microarray. Univariate analysis identified 269 differentially-expressed genes in the hippocampus of the mutant mouse. Pathway analyses highlighted 11 KEGG pathways: the phosphatidylinositol signaling system, which has been widely implicated in MDD, Bipolar Disorder, and the action of mood stabilizers; gap junction and long-term potentiation, which mediate cognition and memory functions often impaired in MDD; cardiac muscle contraction, insulin signaling pathway, and three neurodegenerative brain disorders (Alzheimer's, Parkinson's, and Huntington's Diseases) that are associated with MDD; ribosome and proteasome pathways affecting protein synthesis/degradation; and the oxidative phosphorylation pathway that is key to energy production. These findings illustrate the merit of this congenic C57BL/6 recombinant mouse as a model of RE-MDD, and demonstrate its potential for highlighting molecular and cellular pathways that contribute to the biology of MDD. The results also inform our understanding of the mechanisms that underlie the comorbidity of MDD with other disorders.

The effect of environmental enrichment on substantia nigra gene expression after traumatic brain injury in rats.

Experimental investigations into the effects of traumatic brain injury (TBI) have demonstrated significant alterations in dopaminergic systems. Dopaminergic fibers originating within the substantia nigra and ventral tegmental area (VTA) are important for reward learning, addiction, movement, and behavior. However, little is known about the effect of TBI on substantia nigra and VTA function. Environmental enrichment (EE) has been shown to improve functional outcome after TBI, and a number of studies suggest that it may exert some benefits via dopaminergic signaling. To better understand the role of dopamine in chronic TBI pathophysiology and the effect of EE, we examined the mRNA expression profile within the substantia nigra and VTA at 4 weeks post-injury. Specifically, three comparisons were made: 1) TBI versus sham, 2) sham+EE versus sham+standard (STD) housing, and 3) TBI+EE versus TBI+STD. There were differential expressions of 25, 4, and 40 genes in these comparisons, respectively. Chronic alterations in genes post-injury within the substantia nigra and VTA included genes important for cellular membrane homeostasis and transcription. EE-induced gene alterations after TBI included genes important for signal transduction, in particular calcium signaling pathways, membrane homeostasis, and metabolism. Elucidation of these alterations in gene expression within the substantia nigra and VTA provides new insights into chronic changes in dopamine signaling post-TBI, and the potential role of EE in TBI rehabilitation.

Variations in discovery-based preeclampsia candidate genes.

Preeclampsia is a common and potentially lethal pregnancy disorder with lifelong increased risk of cardiovascular disease in survivors. Our prior global gene expression microarray analysis led to a novel set of 36 candidates in first trimester placentas of women who subsequently developed preeclampsia. In this report, we present preliminary studies demonstrating biomarkers of genotype and methylation variations in a subset of these candidate genes in maternal leukocyte and fetoplacental DNA of 28 case and 27 control dyads. We tested 84 single nucleotide polymorphisms (SNPs) using MassArray iPLEX and 50 CpG sites using EpiTYPER assays. Promising prediction modeling was identified with 25 SNPs selected using Fisher's exact tests (p ≤ 0.05) and 20 CpG sites selected on fold change. Genotype Distribution Analysis identified SNP variations that differed between nine paired cases versus paired controls. The findings validate the examined candidate genes and support feasibility of methods for further biomarker development. The integrative approach that was implemented begins to translate the 36 candidates toward clinical utility as a screening modality for preeclampsia.

Profiling molecular changes induced by hydrogen treatment of lung allografts prior to procurement.

BACKGROUND: We previously demonstrated that donor treatment with inhaled hydrogen protects lung grafts from cold ischemia/reperfusion (I/R) injury during lung transplantation. To elucidate the mechanisms underlying hydrogen's protective effects, we conducted a gene array analysis to identify changes in gene expression associated with hydrogen treatment. METHODS: Donor rats were exposed to mechanical ventilation with 98% oxygen and 2% nitrogen or 2% hydrogen for 3 h before harvest; lung grafts were stored for 4h in cold Perfadex. Affymetrix gene array analysis of mRNA transcripts was performed on the lung tissue prior to implantation. RESULTS: Pretreatment of donor lungs with hydrogen altered the expression of 229 genes represented on the array (182 upregulated; 47 downregulated). Hydrogen treatment induced several lung surfactant-related genes, ATP synthase genes and stress-response genes. The intracellular surfactant pool, tissue adenosine triphosphate (ATP) levels and heat shock protein 70 (HSP70) expression increased in the hydrogen-treated grafts. Hydrogen treatment also induced the transcription factors C/EBPα and C/EBPß, which are known regulators of surfactant-related genes. CONCLUSION: Donor ventilation with hydrogen significantly increases expression of surfactant-related molecules, ATP synthases and stress-response molecules in lung grafts. The induction of these molecules may underlie hydrogen's protective effects against I/R injury during transplantation.

Mechanism of transfer of functional microRNAs between mouse dendritic cells via exosomes.

Dendritic cells (DCs) are the most potent APCs. Whereas immature DCs down-regulate T-cell responses to induce/maintain immunologic tolerance, mature DCs promote immunity. To amplify their functions, DCs communicate with neighboring DCs through soluble mediators, cell-to-cell contact, and vesicle exchange. Transfer of nanovesicles (< 100 nm) derived from the endocytic pathway (termed exosomes) represents a novel mechanism of DC-to-DC communication. The facts that exosomes contain exosome-shuttle miRNAs and DC functions can be regulated by exogenous miRNAs, suggest that DC-to-DC interactions could be mediated through exosome-shuttle miRNAs, a hypothesis that remains to be tested. Importantly, the mechanism of transfer of exosome-shuttle miRNAs from the exosome lumen to the cytosol of target cells is unknown. Here, we demonstrate that DCs release exosomes with different miRNAs depending on the maturation of the DCs. By visualizing spontaneous transfer of exosomes between DCs, we demonstrate that exosomes fused with the target DCs, the latter followed by release of the exosome content into the DC cytosol. Importantly, exosome-shuttle miRNAs are functional, because they repress target mRNAs of acceptor DCs. Our findings unveil a mechanism of transfer of exosome-shuttle miRNAs between DCs and its role as a means of communication and posttranscriptional regulation between DCs.

Lung tissues in patients with systemic sclerosis have gene expression patterns unique to pulmonary fibrosis and pulmonary hypertension.

OBJECTIVE: Pulmonary complications, including pulmonary fibrosis (PF) and pulmonary arterial hypertension (PAH), are the leading cause of mortality in patients with systemic sclerosis (SSc). The aim of this study was to compare the molecular fingerprint of lung tissue and matching primary fibroblasts from patients with SSc with that of lung tissue and fibroblasts from normal donors, patients with idiopathic pulmonary fibrosis (IPF), and patients with idiopathic pulmonary arterial hypertension (IPAH). METHODS: Lung tissue samples were obtained from 33 patients with SSc who underwent lung transplantation. Tissues and cells from a subgroup of SSc patients with predominantly PF or PAH were compared to those from normal donors, patients with IPF, and patients with IPAH. Microarray data were analyzed using efficiency analysis for determination of the optimal data-processing methods. Real-time polymerase chain reaction and immunohistochemistry were used to confirm differential levels of messenger RNA and protein, respectively. RESULTS: Consensus efficiency analysis identified 242 and 335 genes that were differentially expressed in lungs and primary fibroblasts, respectively. SSc-PF and IPF lungs shared enriched functional groups in genes implicated in fibrosis, insulin-like growth factor signaling, and caveolin-mediated endocytosis. Gene functional groups shared by SSc-PAH and IPAH lungs included those involved in antigen presentation, chemokine activity, and interleukin-17 signaling. CONCLUSION: Using microarray analysis on carefully phenotyped SSc and comparator lung tissues, we demonstrated distinct molecular profiles in tissues and fibroblasts from patients with SSc-associated lung disease compared to idiopathic forms of lung disease. Unique molecular signatures were generated that are disease specific (SSc) and phenotype specific (PF versus PAH). These signatures provide new insights into the pathogenesis and potential therapeutic targets of SSc-related lung disease.

Evaluation of the Consensus of Four Peptide Identification Algorithms for Tandem Mass Spectrometry Based Proteomics.

The availability of different scoring schemes and filter settings of protein database search algorithms has greatly expanded the number of search methods for identifying candidate peptides from MS/MS spectra. We have previously shown that consensus-based methods that combine three search algorithms yield higher sensitivity and specificity compared to the use of a single search engine (individual method). We hypothesized that union of four search engines (Sequest, Mascot, X!Tandem and Phenyx) can further enhance sensitivity and specificity. ROC plots were generated to measure the sensitivity and specificity of 5460 consensus methods derived from the same dataset. We found that Mascot outperformed individual methods for sensitivity and specificity, while Phenyx performed the worst. The union consensus methods generally produced much higher sensitivity, while the intersection consensus methods gave much higher specificity. The union methods from four search algorithms modestly improved sensitivity, but not specificity, compared to union methods that used three search engines. This suggests that a strategy based on specific combination of search algorithms, instead of merely 'as many search engines as possible', may be key strategy for success with peptide identification. Lastly, we provide strategies for optimizing sensitivity or specificity of peptide identification in MS/MS spectra for different user-specific conditions.