Criteria Specification (CSpec) Registry is intended to provide access to the Criteria Specifications used and applied by ClinGen Variant Curation Expert Panels and biocurators in the classification of variants.
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Criteria & Strength Specifications
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PVS1 | ||||
Original ACMG Summary
Null variant (nonsense, frameshift, canonical +/−1 or 2 splice sites, initiation codon, single or multi-exon deletion) in a gene where loss of function (LOF) is a known mechanism of disease.
Caveats: • Beware of genes where LOF is not a known disease mechanism (e.g. GFAP, MYH7). • Use caution interpreting LOF variants at the extreme 3’ end of a gene. • Use caution with splice variants that are predicted to lead to exon skipping but leave the remainder of the protein intact. • Use caution in the presence of multiple transcripts. Stand Alone
Very Strong
Null variant in a gene where loss of function is a known mechanism of disease.
Modification Type:
Gene-specific
Strong
Null variant in a gene where loss of function is a known mechanism of disease.
Modification Type:
Gene-specific
Moderate
Supporting
Instructions:
See attached GUCY2D-specific PVS1 Decision Tree file, which has been modified from Walker et al., 202317 Not Applicable
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PS1 | ||||
Original ACMG Summary
Same amino acid change as a previously established pathogenic variant regardless of nucleotide change.
Example: Val->Leu caused by either G>C or G>T in the same codon. Caveat: Beware of changes that impact splicing rather than at the amino acid/protein level. Stand Alone
Very Strong
Strong
Same amino acid change as a previously established Pathogenic variant regardless of nucleotide change.
Same predicted splicing impact as a previously classified Pathogenic variant.
Specific combinations are found in GUCY2D-specific PVS1 Decision Tree part (b) (Table 2 from Walker 2023).
Modification Type:
Gene-specific
Moderate
Same amino acid change as a previously established Likely Pathogenic variant regardless of nucleotide change.
Same predicted splicing impact as previously classified Likely Pathogenic variant.
Specific combinations are found in GUCY2D-specific PVS1 Decision Tree part (b) (Table 2 from Walker 2023).
Modification Type:
Gene-specific,Strength
Supporting
Specific combinations are found in GUCY2D-specific PVS1 Decision Tree part (b) (Table 2 from Walker 2023).
Modification Type:
Gene-specific,Strength
Instructions:
Not Applicable
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PS2 | ||||
Original ACMG Summary
De novo (both maternity and paternity confirmed) in a patient with the disease and no family history.
Note: Confirmation of paternity only is insufficient. Egg donation, surrogate motherhood, errors in embryo transfer, etc. can contribute to non-maternity. Stand Alone
Very Strong
De novo (both maternity and paternity confirmed) in a patient with the disease and no family history.
Modification Type:
Gene-specific
Strong
De novo (both maternity and paternity confirmed) in a patient with the disease and no family history.
Modification Type:
Gene-specific
Moderate
De novo (both maternity and paternity confirmed) in a patient with the disease and no family history.
Modification Type:
Gene-specific
Supporting
De novo (both maternity and paternity confirmed) in a patient with the disease and no family history.
Modification Type:
Gene-specific
Instructions:
Use point table from SVI Recommendation for De Novo Criteria (PS2 & PM6) - Version 1.1 12 , file attached Individuals must have 2 variants to consider scoring one for de novo. To determine the appropriate strength level to apply for de novo occurrence(s), each proband with a de novo variant is awarded a point value based upon phenotypic consistency and confirmed or assumed parental relationships (PS2/PM6 file -Table 1). The combined point value of all de novo occurrences is then compared to Table 2 to determine the applicable evidence strength level. For “Phenotypic consistency” category, consider whether the proband meets PP4:
Note that all probands being considered for any pathogenic phenotype codes (i.e. – PP1, PP4, PM3, PM6, PS2) at any strength must have the following phenotype characteristics: Absent or severely decreased rod electroretinogram response OR congenital night blindness/nyctalopia OR a diagnosis of Leber congenital amaurosis/eoRD/cone-rod dystrophy. Not Applicable
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PS3 | ||||
Original ACMG Summary
Well-established in vitro or in vivo functional studies supportive of a damaging effect on the gene or gene product.
Note: Functional studies that have been validated and shown to be reproducible and robust in a clinical diagnostic laboratory setting are considered the most well-established. Stand Alone
Very Strong
Strong
Well-established in vitro or in vivo functional studies supportive of a damaging effect. See attached table for acceptable functional studies. Eligible results include Gucy2e−/−Gucy2f−/− knockout mice subretinally injected with adeno-associated virus-packaged constructs encoding a GUCY2D variant (Boye et al., 2016,10) or mice with knock-in of a GUCY2D variant (not yet available in the published literature). Results should include flat or nearly flat scotopic and photopic electroretinogram responses in comparison to wild-type injected control, absent or near-absent guanylate cyclase activity in comparison to wild-type injected control, and/or absent or near absent GUCY2D expression at the protein level despite approximately wild-type expression at the RNA level.
Modification Type:
Gene-specific,Strength
Moderate
Supporting
Well-established in vitro or in vivo functional studies supportive of a damaging effect. Not applicable for splicing effects (replaced by PVS1_Strength (RNA)) See attached table for acceptable functional studies. For studies reporting guanylate cyclase activity, cutoff is ≤10% of wild-type control.
Modification Type:
Gene-specific,Strength
Instructions:
Studies included in attached table: Boye et al., 2016 10 Daich Varela et al., 2023 18 Duda et al.,1999 9 Duda et al., 2011 20 Jacobson et al., 2013 5 Jacobson et al., 2021 2 Jacobson et al., 2022 3 Peshenko et al., 2010 21 Peshenko et al., 2015 7 Peshenko et al., 2020 1 Tucker et al., 1998 8 Tucker et al., 2004 4 Wimberg et al., 2018 15 Zägel et al., 2014 19 Zulliger et al., 2015 6 Not Applicable
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PS4 | ||||
Original ACMG Summary
The prevalence of the variant in affected individuals is significantly increased compared to the prevalence in controls.
Note 1: Relative risk (RR) or odds ratio (OR), as obtained from case-control studies, is >5.0 and the confidence interval around the estimate of RR or OR does not include 1.0. See manuscript for detailed guidance. Note 2: In instances of very rare variants where case-control studies may not reach statistical significance, the prior observation of the variant in multiple unrelated patients with the same phenotype, and its absence in controls, may be used as moderate level of evidence. Stand Alone
Very Strong
Strong
Moderate
Supporting
Instructions:
Not Applicable
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PM1 | ||||
Original ACMG Summary
Located in a mutational hot spot and/or critical and well-established functional domain (e.g. active site of an enzyme) without benign variation.
Stand Alone
Very Strong
Strong
Moderate
Met by active site residues that bind the GTP substrate or the Mg2+ ion; Phe883, Asp885, Thr890, Leu905, Glu925, Ile927, Asp929, Met932, Arg976, Arg995, Cys997, Leu998, Phe999, Gly1000, Val1003, Asn1004, Arg1008, and Glu1010 13.
Modification Type:
Gene-specific
Supporting
Modification Type:
Gene-specific
Instructions:
Because these sites have been identified based on well-established functional domains rather than based on a known cluster of pathogenic or likely pathogenic variants, this criterion is not mutually exclusive with PM5.
Not Applicable
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PM2 | ||||
Original ACMG Summary
Absent from controls (or at extremely low frequency if recessive) in Exome Sequencing Project, 1000 Genomes or Exome Aggregation Consortium.
Caveat: Population data for indels may be poorly called by next generation sequencing. Stand Alone
Very Strong
Strong
Moderate
Supporting
Absent/rare from controls in an ethnically-matched cohort population sample.
Modification Type:
Disease-specific,Strength
Instructions:
Cutoff value of 4.0x10-4 is set just above the FAF of the most common pathogenic GUCY2D variant (3.6x10-4) and below the Whiffin-Ware calculation of 1.6x10-3 for the maximum credible population allele frequency for the disease. This rule should not be applied if variant would otherwise meet criteria for a benign classification, as rarity of the variant should not outweigh other types of benign evidence. Not Applicable
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PM3 | ||||
Original ACMG Summary
For recessive disorders, detected in trans with a pathogenic variant
Note: This requires testing of parents (or offspring) to determine phase. Stand Alone
Very Strong
Points are calculated using Table 1 from https://clinicalgenome.org/site/assets/files/3717/svi_proposal_for_pm3_criterion_-_version_1.pdf (or attached file “PM3 Tables”)
Modification Type:
Disease-specific
Strong
Points are calculated using Table 1 from https://clinicalgenome.org/site/assets/files/3717/svi_proposal_for_pm3_criterion_-_version_1.pdf (or attached file “PM3 Tables”)
Modification Type:
Disease-specific
Moderate
Points are calculated using Table 1 from https://clinicalgenome.org/site/assets/files/3717/svi_proposal_for_pm3_criterion_-_version_1.pdf (or attached file “PM3 Tables”)
Modification Type:
Disease-specific
Supporting
Points are calculated using Table 1 from https://clinicalgenome.org/site/assets/files/3717/svi_proposal_for_pm3_criterion_-_version_1.pdf (or attached file “PM3 Tables”)
Modification Type:
Disease-specific
Instructions:
Use SVI recommendations found in https://clinicalgenome.org/site/assets/files/3717/svi_proposal_for_pm3_criterion_-_version_1.pdf (or attached file “PM3 Tables”) To determine the appropriate strength level to apply for in trans occurrence(s), each proband is awarded a point value based upon phasing of the two variants in question (confirmed in trans versus unknown) and classification of the variant on the other allele (Table 1). The combined point value of all proband occurrences is then summed and compared to Table 2 to determine the applicable evidence strength level. Both variants must be classified using these rule specifications. Note that all probands being considered for any pathogenic phenotype codes (i.e. – PP1, PP4, PM3, PM6, PS2) at any strength must have the following phenotype characteristics: Absent or severely decreased rod electroretinogram response OR congenital night blindness/nyctalopia OR a diagnosis of Leber congenital amaurosis/eoRD/cone-rod dystrophy Not Applicable
Comments:
Not applicable for UBE3A.
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PM4 | ||||
Original ACMG Summary
Protein length changes due to in-frame deletions/insertions in a non-repeat region or stop-loss variants.
Stand Alone
Very Strong
Strong
Moderate
Protein length changes due to in-frame deletions/insertions in a non-repeat region or stop-loss variants.
Modification Type:
Gene-specific
Supporting
Protein length changes due to in-frame deletions/insertions in a non-repeat region or stop-loss variants.
Modification Type:
Gene-specific,Strength
Not Applicable
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PM5 | ||||
Original ACMG Summary
Novel missense change at an amino acid residue where a different missense change determined to be pathogenic has been seen before.
Example: Arg156His is pathogenic; now you observe Arg156Cys. Caveat: Beware of changes that impact splicing rather than at the amino acid/protein level. Stand Alone
Very Strong
Strong
Moderate
Missense change at an amino acid residue where a different missense change determined to be pathogenic has been seen before.
Modification Type:
None
Supporting
Missense change at an amino acid residue where a different missense change determined to be pathogenic has been seen before.
Modification Type:
Strength
Not Applicable
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PM6 | ||||
Original ACMG Summary
Assumed de novo, but without confirmation of paternity and maternity.
Stand Alone
Very Strong
Strong
Moderate
Supporting
Instructions:
Use point table from SVI Recommendation for De Novo Criteria (PS2 & PM6) - Version 1.1 1, file attached To determine the appropriate strength level to apply for de novo occurrence(s), each proband with a de novo variant is awarded a point value based upon phenotypic consistency and confirmed or assumed parental relationships (Table 1). The combined point value of all de novo occurrences is then compared to Table 2 to determine the applicable evidence strength level. For “Phenotypic consistency” category, use option 3: "Phenotype consistent with gene but not highly specific and high genetic heterogeneity" Note that all probands being considered for any pathogenic phenotype codes (i.e. – PP1, PP4, PM3, PM6, PS2) at any strength must have the following phenotype characteristics: Absent or severely decreased rod electroretinogram response OR congenital night blindness/nyctalopia OR a diagnosis of Leber congenital amaurosis/eoRD. Not Applicable
Comments:
Use the PS2 code in lieu of using this code for de novo variants.
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PP1 | ||||
Original ACMG Summary
Co-segregation with disease in multiple affected family members in a gene definitively known to cause the disease.
Note: May be used as stronger evidence with increasing segregation data. Stand Alone
Very Strong
Strong
Co-segregation with disease in multiple affected family members and evidence that this variant and another GUCY2D variant are in trans.
Modification Type:
Disease-specific,Strength
Moderate
Co-segregation with disease in multiple affected family members and evidence that this variant and another GUCY2D variant are in trans.
Modification Type:
Disease-specific,Strength
Supporting
Co-segregation with disease in multiple affected family members and evidence that this variant and another GUCY2D variant are in trans.
Modification Type:
Disease-specific,Strength
Instructions:
For moderate and strong codes, segregations can be added across multiple families, with each having a proband + at least one affected relative. Note that all probands being considered for any pathogenic phenotype codes (i.e. – PP1, PP4, PM3, PM6, PS2) at any strength must have the following phenotype characteristics: Absent or severely decreased rod electroretinogram response OR congenital night blindness/nyctalopia OR a diagnosis of Leber congenital amaurosis/eoRD/cone-rod dystrophy Not Applicable
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PP2 | ||||
Original ACMG Summary
Missense variant in a gene that has a low rate of benign missense variation and where missense variants are a common mechanism of disease.
Stand Alone
Very Strong
Strong
Moderate
Supporting
Not Applicable
Comments:
Not applicable for GUCY2D.
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PP3 | ||||
Original ACMG Summary
Multiple lines of computational evidence support a deleterious effect on the gene or gene product (conservation, evolutionary, splicing impact, etc.).
Caveat: As many in silico algorithms use the same or very similar input for their predictions, each algorithm should not be counted as an independent criterion. PP3 can be used only once in any evaluation of a variant. Stand Alone
Very Strong
Strong
Moderate
Modification Type:
Gene-specific,Strength
Supporting
Modification Type:
Gene-specific
Instructions:
PP3 should not be used to evaluate variants at canonical splice sites. For canonical splice sites, apply PVS1(splicing). For non-canonical sites, if SpliceAI score is ≥0.2, apply PP3 (splicing) instead. Score ranges are based on calculations found in the publication of the SVI Working Group “Calibration of computational tools for missense variant pathogenicity classification and ClinGen recommendations for PP3/BP4 criteria” Not Applicable
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PP4 | ||||
Original ACMG Summary
Patient’s phenotype or family history is highly specific for a disease with a single genetic etiology.
Stand Alone
Very Strong
Strong
Moderate
Modification Type:
Disease-specific,Strength
Supporting
Modification Type:
Disease-specific
Instructions:
A point system was developed to determine when there is enough information about a proband’s phenotype to qualify for use of this code (see below). Review eligible phenotype characteristics and add up points for each finding. This code can be used for a single proband. A proband must have two GUCY2D variants to consider applying PP4 (phase is not considered here). Caveat: PP4 should not be applied if variant meets either BA1 or BS1. Required for use of PP4 (0.5 points each)
Specific GUCY2D Phenotype Findings List
Consistent with GUCY2D Phenotype Findings List (0.5 or 1 point each)
Not Applicable
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PP5 | ||||
Original ACMG Summary
Reputable source recently reports variant as pathogenic, but the evidence is not available to the laboratory to perform an independent evaluation.
Not Applicable
This criterion is not for use as recommended by the ClinGen Sequence Variant Interpretation VCEP Review Committee.
PubMed : 29543229
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BA1 | ||||
Original ACMG Summary
Allele frequency is above 5% in Exome Sequencing Project, 1000 Genomes or Exome Aggregation Consortium.
Stand Alone
Use gnomAD Grpmax FAF if available, cutoff of >0.016
Modification Type:
Disease-specific
Very Strong
Strong
Moderate
Supporting
Instructions:
The BA1 value was derived by increasing the BS1 lower cutoff (> 0.0016) by one order of magnitude. Not Applicable
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BS1 | ||||
Original ACMG Summary
Allele frequency is greater than expected for disorder.
Stand Alone
Very Strong
Strong
Allele frequency greater than expected for disorder. Use gnomAD Grpmax FAF if available.
Modification Type:
Disease-specific
Moderate
Supporting
Instructions:
The maximum credible population allele frequency for the disease, based on the Whiffen-Ware calculator, is 1.6 x 10 -3. This assumes a population frequency of 1 in 2000 individuals, genetic heterogeneity = 20%, penetrance of 100%, allele heterogeneity of 1. Not Applicable
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BS2 | ||||
Original ACMG Summary
Observed in a healthy adult individual for a recessive (homozygous), dominant (heterozygous), or X-linked (hemizygous) disorder, with full penetrance expected at an early age.
Stand Alone
Very Strong
Strong
Variant is present in ≥ 3 homozygotes without any features of the phenotype. This rule applies to individuals found in the literature who have been well-phenotyped and are unaffected by age 40. Alternatively, this strength can be applied if the variant is present in ≥ 6 homozygotes in gnomAD v.4.1.0 or later.
Modification Type:
Disease-specific,Strength
Moderate
Supporting
Variant is present in ≥ 3 homozygotes in gnomAD v.4.1.0 or later.
Modification Type:
Disease-specific,Strength
Not Applicable
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BS3 | ||||
Original ACMG Summary
Well-established in vitro or in vivo functional studies show no damaging effect on protein function or splicing.
Stand Alone
Very Strong
Strong
Moderate
Supporting
Not applicable for splicing effects (replaced by BP7_Strong (RNA)). For studies reporting guanylate cyclase activity, cutoff is >50% of wild-type control. Studies included; Peshenko et al., 20201, Feng et al., 2020, Jacobson et al., 20212, Jacobson et al., 20203, Tucker et al., 20044, Jacobson et al., 20135, and Peshenko et al., 20157 See attached table of Approved PS3 Functional Assays for a list of acceptable functional studies including those that:
Modification Type:
Gene-specific,Strength
Instructions:
Extracellular domain variants (occurring between amino acids 52-462) are known to show no damaging effect in these assays and are excluded from BS3. Not Applicable
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BS4 | ||||
Original ACMG Summary
Lack of segregation in affected members of a family.
Caveat: The presence of phenocopies for common phenotypes (i.e. cancer, epilepsy) can mimic lack of segregation among affected individuals. Also, families may have more than one pathogenic variant contributing to an autosomal dominant disorder, further confounding an apparent lack of segregation. Stand Alone
Very Strong
Strong
Lack of segregation in affected members of a family.
Modification Type:
Clarification
Moderate
Supporting
Not Applicable
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BP1 | ||||
Original ACMG Summary
Missense variant in a gene for which primarily truncating variants are known to cause disease.
Stand Alone
Very Strong
Strong
Moderate
Supporting
Not Applicable
Comments:
Not applicable for GUCY2D.
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BP2 | ||||
Original ACMG Summary
Observed in trans with a pathogenic variant for a fully penetrant dominant gene/disorder or observed in cis with a pathogenic variant in any inheritance pattern.
Stand Alone
Very Strong
Strong
Moderate
Supporting
Observed in cis with a Pathogenic variant.
Modification Type:
Disease-specific
Not Applicable
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BP3 | ||||
Original ACMG Summary
In frame-deletions/insertions in a repetitive region without a known function.
Stand Alone
Very Strong
Strong
Moderate
Supporting
Not Applicable
Comments:
No repetitive regions with unknown function.
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BP4 | ||||
Original ACMG Summary
Multiple lines of computational evidence suggest no impact on gene or gene product (conservation, evolutionary, splicing impact, etc)
Caveat: As many in silico algorithms use the same or very similar input for their predictions, each algorithm cannot be counted as an independent criterion. BP4 can be used only once in any evaluation of a variant. Stand Alone
Very Strong
Strong
Moderate
For a missense variant use REVEL, requires a score of ≤0.183. In addition, highest SpliceAI delta score should also be below cutoff of 0.1.
Modification Type:
Gene-specific,Strength
Supporting
For a missense variant use REVEL, requires a score between 0.183 - 0.290. In addition, highest SpliceAI delta score should also be below cutoff of 0.1. For a silent / intronic variant outside the designated splice region (conservatively at or beyond positions +7/-21) and synonymous (silent) exonic variants located outside of the first and the last 3 bases of the exon, BP4 can be met if the highest of the four delta scores within SpliceAI is below the cutoff of ≤0.1. See GUCY2D-specific PVS1 Decision Tree part (a) for SpliceAI flowchart.
Modification Type:
Gene-specific
Instructions:
Only applicable if both the REVEL and SpliceAI scores are below cutoffs. Not Applicable
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BP5 | ||||
Original ACMG Summary
Variant found in a case with an alternate molecular basis for disease.
Stand Alone
Very Strong
Strong
Moderate
Supporting
Not Applicable
Comments:
Due to the high genetic heterogeneity and limited phenotypic specificities of retinal dystrophies, this rule should not be used.
Additionally, the presence of this variant could simply represent carrier status.
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BP6 | ||||
Original ACMG Summary
Reputable source recently reports variant as benign, but the evidence is not available to the laboratory to perform an independent evaluation.
Not Applicable
This criterion is not for use as recommended by the ClinGen Sequence Variant Interpretation VCEP Review Committee.
PubMed : 29543229
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BP7 | ||||
Original ACMG Summary
A synonymous variant for which splicing prediction algorithms predict no impact to the splice consensus sequence nor the creation of a new splice site AND the nucleotide is not highly conserved.
Stand Alone
Very Strong
Strong
BP7_Strong (RNA) Used to designate capture of splicing data (not BS3). See GUCY2D-specific PVS1 Decision Tree, file attached, for weighting and combining with other codes.
Modification Type:
Disease-specific
Moderate
Supporting
Use not only for synonymous variants but also for intronic variants located outside of the donor/acceptor +/- 1,2 dinucleotide positions. If SpliceAI score ≤0.1, apply BP4 followed by assessment of BP7. See GUCY2D-specific PVS1 Decision Tree part (a) for SpliceAI flowchart.
Modification Type:
Gene-specific
Instructions:
BP4 and BP7 can be added unless variant is in an excluded region. Evolutionary conservation is not considered informative for application of this code. Not Applicable
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