Diagnostic and disease severity determination criteria for hydroa vacciniforme lymphoproliferative disorders and severe mosquito bite allergy

Hydroa vacciniforme lymphoproliferative disorder (HV‐LPD) and severe mosquito bite allergy (SMBA) are both cutaneous forms of Epstein–Barr virus (EBV)‐associated T/natural killer (NK) cell LPDs and are closely related to chronic active EBV disease (CAEBV) and EBV‐associated hemophagocytic lymphohistiocytosis (EBV‐HLH). HV‐LPD is further divided into classic HV, a benign subtype mediated by EBV‐positive γδT cells, and systemic HV, another life‐threatening subtype mainly associated with EBV‐positive αβT or γδT cells. The vast majority of patients with SMBA have increased numbers of EBV‐infected NK cells in the blood. Clinical symptoms of HV‐LPD and SMBA often overlap in the same patient and may progress to more serious disease conditions equivalent to the systemic form of CAEBV. To define the disease spectrum of HV‐LPD and SMBA, we propose the diagnostic criteria and the determination criteria for disease severity. The proposed diagnostic criteria are consistent with those for CAEBV and EBV‐HLH in the guidelines for the management for CAEBV and related disorders 2023.


| INTRODUC TI ON
Hydroa vacciniforme lymphoproliferative disorder (HV-LPD) and severe mosquito bite allergy (SMBA) represent two cutaneous variants of Epstein-Barr virus (EBV)-positive T/natural killer (NK) cell LPDs. Given that both diseases occasionally overlap and progress to the serious condition comparable with chronic active EBV disease (CAEBV), they have also been classified into the cutaneous form of CAEBV. 1 Before the terms HV-LPD and SMBA were introduced, various diagnostic names had been used in previous studies. [2][3][4] Furthermore, both diseases still have no consensus diagnostic criteria that can distinguish them clearly from CAEBV. In 2018, a consensus meeting on the disease spectrum and terminology for HV-LPD was held in Madison, USA. 5 Thereafter, the 5th edition of the World Health Organization (WHO) classification adopted the newly proposed nomenclatures and defined the spectrum of HV-LPD, SMBA and "systemic" CAEBV separately. 6 The term systemic CAEBV (ICD-O coding: 9725/1) is synonymous with the term CAEBV used in this report.
In line with the consensus terminology and definition, 6,7 the Ministry of Health, Labour and Welfare (MHLW) research team for CAEBV and related disorders in Japan redefined the disease spectrum of HV-LPD and SMBA, and proposed the diagnostic criteria and the determination criteria for disease severity. A full set of guidelines for the management of CAEBV, HV-LPD, SMBA, and EBV-associated hemophagocytic lymphohistiocytosis (EBV-HLH) has been published elsewhere. 8 In this report, we summarize the diagnostic criteria and the disease severity determination criteria for HV-LPD and SMBA.

| MATERIAL S AND ME THODS
For the diagnostic and determination criteria for disease severity, we conducted a literature review on etiologically related disorders, including HV-LPD, SMBA, CAEBV, HLH, and EBV-related lymphoma/ leukemia. Through expert meetings, the disease spectrum and nomenclature were revised to be consistent with those adopted in the 5th edition of the WHO classification 6 and the newly launched terminologies and definitions for CAEBV and related diseases. 7

| HV-LPD: D IS E A S E S PEC TRUM AND D IAG NOS TI C CRITERIA
Hydroa vacciniforme (HV) was first described by EH Bazin in 1862 as a benign photosensitivity dermatosis of childhood, characterized by herpetic vesicles on sun-exposed areas. 9 In 1999, EBV-positive T cells were detected in skin lesions with classic HV, 10 and then EBVpositive γδT cells were found to be increased in the patient's blood ( Figure 1). [11][12][13] The term "classic" HV is used in this report to represent the certain type of HV-LPD mentioned above.
have been reported in Eastern Asia and Central America under different names: recurrent necrotic papulovesicles of the face, severe or systemic HV, and HV-like lymphoma. 2,3,14,15 Many of those patients have an increased number of EBV-positive αβT or γδT cells in the peripheral blood, and may overlap with SMBA or have a history of SMBA associated with an increased number of EBV-positive NK cells. Those patients often progress to severe disease conditions such as CAEBV and fatal EBV-related lymphomas.
Based on the similarity of cutaneous manifestations, the term hydroa vacciniforme-like LPD has long been used to encompass classic HV and severe forms. 1,5,16 In the 5th edition of the WHO classification, 6 the term hydroa vacciniforme LPD (HV-LPD) has been adopted instead because EBV infection can be confirmed to be present in all HV lesions, including the classic form. [10][11][12] HV-LPD is classified into classic HV (or classic HV-LPD), a major subtype occurring among Caucasians, and systemic HV (or systemic HV-LPD), which is mainly encountered in Asians and Hispanics (Table 1). 16 Classic HV is a self-limiting skin disease without systemic symptoms or extracutaneous organ involvement, whereas systemic HV may exhibit severe cutaneous manifestations and systemic symptoms, and extracutaneous organ involvement which often leads to an unfavorable outcome. [15][16][17][18][19][20] From therapeutic and prognostic points of view, classic HV must be clearly distinguished from the systemic HV, although approximately 10% of patients with an initial diagnosis of classic HV may progress to the systemic type. 15 (1) Herpetic vesicles and papules/crusts on sun-exposed areas including face, auricles, and dorsal surfaces of the hands.
(2) EBV-infected cells or the virus-gene products can be detected in the skin lesions.
(3) The number of EBV-positive T cells is increased in peripheral blood. (4) The level of EBV DNA load in the blood is high. (5) The following diseases are excluded: other known skin diseases, CAEBV, and SMBA. The definite case fulfills all of (1)-(5). The probable case fulfills (1) and has at least one of (2) and (3). The possible case fulfills (1) and (4).

Supplementary explanation
Classic hydroa vacciniforme (1) In addition to herpetic vesicles, cHV may be accompanied with conjunctival hyperemia and aphthous stomatitis of the lips and gingiva.
(2) Detection of EBER-positive cells in the skin biopsy samples is a standard method, but when biopsy is difficult, the blister roof or crusts can be used instead for EBER detection by RT-PCR (at a research level). (3) cHV usually occurs in childhood, associated with an increased number of EBV-infected γδT cells (>5% of all lymphocytes). sHV may occur even in young and old adults as well as in children, and include either EBV-infected αβT-cell-dominant or γδT-cell-dominant type. Flow cytometryfluorescent in-situ hybridization or immunomagnetic bead-aided quantitative PCR is performed at a research level to confirm EBV infection in the T-cell fraction. (4) The standardized value of elevated EBV DNA load in whole blood is >1 × 10E4 IU/mL.  showed that some patients with CAEBV, HV-LPD, and SMBA share similar somatic mutations such as DDX3X, as also observed in other EBV-associated T/NK-cell lymphomas, and that the cases carrying a driver mutation showed a significantly poor prognosis. 22 Therefore, patients with severe HV-LPD are closely related to CAEBV, and need careful follow-up and management for CAEBV ( Figure 2). 6,8,17,18

| S MBA : D IS E A S E S PEC TRUM AND D IAG NOS TI C CRITERIA
SMBA is characterized by a swollen erythematous skin reaction followed by deep ulcers on the sites of mosquito or insect bites and even vaccine injection ( Figure 1). Previously, SMBA was termed hypersensitivity to mosquito bites. 4,20,23 Presumably, mosquito bites induce an initial inflammation, which provokes the secondary severe cutaneous reactions due to the infiltration of activated EBV-positive NK cells. 23 The number of EBV-infected NK cells showing a cell type of large granular lymphocytes (LGL) is usually increased (>30% of lymphocytes) in the peripheral blood. 21,24 Even though the skin reaction is severe, an allergic reaction to mosquito bites without the involvement of EBV-positive NK/T cells should be excluded from SMBA's diagnostic criteria (Table 3).

| S MBA : D IS E A S E S E VERIT Y AND DIAG NOS TIC ALGORITHM
Patients with definite SMBA who have been in remission for more than 1 year, with or without residual hematological abnormalities, are classified as mild SMBA (Table 4) underlying diseases are excluded. The definite case fulfills all of (1)-(5). The probable case fulfills (1) and has at least one of (2) and (4). The possible case fulfills (1) and (3).

Supplementary explanation
(1) SMBA is mostly induced by mosquito bites, but it can also be caused by other insect bites and vaccine injection. Patients may present with transient systemic symptoms including fever, lymphadenopathy, and liver dysfunctions. Scar formation resulting from previous episodes of SMBA can be present. (2) Detection of EBER-positive cells using skin biopsy samples is a standard method, but when biopsy is difficult, EBER can be detected in the crusts and lesional skin tissue by RT-PCR at a research level. (3) The standardized value of elevated EBV DNA load in whole blood is >1 × 10E4 IU/mL. (4) Flow cytometric analysis reveals that most patients with SMBA have an increased total number of NK cells (≥30% of the total lymphocytes). Flow cytometry-fluorescent in-situ hybridization or immunomagnetic bead-aided quantitative PCR is performed at a research level to confirm EBV infection in the NK cell fraction. Some patients may have an EBV-infected T-celldominant type of SMBA. (5) Diseases to be excluded: blisters, redness, swelling, and generalized rash related to common insect bite allergy, and pathergy reactions associated with neutrophilic dermatoses including Behçet's disease, Sweet disease, and pyoderma gangrenosum. The overlapping of CAEBV should be considered in line with the diagnostic algorithm if patients have the following symptoms: infectious mononucleosis-like symptoms for >3 months, persistent fever, and organ involvement (see the CAEBV algorithm). SMBA may overlap HV-LPD associated with systemic symptoms, thereby being included in the category of cutaneous CAEBV. The cutaneous form, however, is not the same as CAEBV defined in the present diagnostic criteria. SMBA may progress to ENKTL-NT, and aggressive NK-cell leukemia.
Abbreviations: CAEBV, chronic active Epstein-Barr virus disease; EBER, EBV-encoded small nuclear RNA; EBV, Epstein-Barr virus; ENKTL-NT, extranodal NK/T-cell lymphoma, nasal type; HV-LPD, hydroa vacciniforme lymphoproliferative disorder; LGL, large granular lymphocyte; NK, natural killer; NK/T, natural killer cell, or T cell; RT-PCR, reverse transcription PCR; SMBA, severe mosquito bite allergy.  Other prognostic indicators include an onset age over 9 years (over 8 years in Okuno et al. 22 ), the mRNA expression of BZLF1, a reactivation marker, 19,25 and the presence of a somatic driver mutation such as DDX3X. 22 These prognostic indicators, however, are excluded from the present criteria because the onset age is not determined clearly by medical interview alone, and the detection of molecular and genetic markers has not widely been used in a clinical setting.

TA B L E 4 Determination criteria for the disease severity of SMBA
The elevated EBV DNA load (>1 × 10E4 IU/mL) in the whole peripheral blood sample is useful for a diagnosis of HV-LPD and SMBA, but not applicable as a prognostic indicator. However, extremely high levels of EBV DNA load in the plasma are found to be related to hemophagocytic syndrome complications. 26 In the present diagnostic criteria, classic HV can be separated from systemic HV and classified into the mild form of HV-LPD in the disease severity determination criteria because of its favorable prognosis. 16

| PROS PEC TIVE
Consensus diagnostic criteria are important to perform the patient's registry, statistical analysis, and therapeutic effect evaluation. We hope that the present diagnostic criteria can contribute to these purposes.

ACK N OWLED G M ENTS
The authors would like to thank the members of the MHLW Research Team in Japan for their contribution to the review process of the current diagnostic criteria.

FU N D I N G I N FO R M ATI O N
This study was supported by the grant (20FC1004 and 22FC1004) from the Ministry of Health, Labour and Welfare (MHLW), Japan.

CO N FLI C T O F I NTER E S T S TATEM ENT
None.