Direct Immunofluorescence Research Articles

Dynamics of parainfluenza virus among hospitalized children with acute respiratory tract infection under two-child policy and COVID-19 pandemic in Hubei Province, China, 2014-2022.

To analyze changes in the detection of parainfluenza virus (PIV) in children hospitalized with acute respiratory tract infection (ARTI) during 2014-2022 in Hubei Province, and explore the impact of the universal two-child policy and the public health measures against COVID-19 epidemic on the prevalence of PIV in China. The study was conducted at the Maternal and Child Health Hospital of Hubei Province. Children aged <18 years with ARTI admitted from January 2014 to June 2022 were enrolled. The infection of PIV was confirmed by the direct immunofluorescence method in nasopharyngeal specimens. Adjusted logistic regression models were used to analyze the influence of the universal two-child policy implementation and public health measurements against COVID-19 on PIV detection. Totally 75 128 inpatients meeting the criteria were enrolled in this study from January 2014 to June 2022 with an overall PIV positive rate of 5.5%. The epidemic seasons of PIV prevalence lagged substantially in 2020. A statistically significant higher positive rate of PIV was observed in 2017-2019 compared to that in 2014-2015 (6.12% vs 2.89%, risk ratio = 2.12, p < 0.001) after the implementation of the universal two-child policy in 2016. A steep decline occurred in PIV positive rate during the COVID-19 epidemic in 2020 (0.92% vs 6.92%, p < 0.001) and it rebounded during the regular epidemic prevention and control period in 2021-2022 (6.35%, p = 0.104). In Hubei Province, the implementation of the universal two-child policy might have led to an increase of PIV prevalence, and public health measures during the COVID-19 epidemic might have influenced the fluctuation in PIV detection since 2020.

The features of regulatory T-blood cells subpopulation and the level of CD25 expression in patients with Graves’ disease in dynamics after radioactive iodine therapy

BACKGROUND: The content of regulatory T cells (Treg) at different stages in formation of effector subpopulations and the level of CD25 expression on the membrane of their various fractions in Graves’ disease can determine the long-term autoimmune process persistence and be the target of immunotropic therapy of the disease.AIM: To study the features of regulatory T-blood cells subpopulation and the level of CD25 expression in patients with Graves’ disease in dynamics after radioactive iodine therapy (RIT) to identify the specific Treg subpopulations for potential immunotropic therapy targets of the disease.MATERIALS AND METHODS: A single-center, prospective, cohort, open, controlled study was conducted with the participation of women with laboratory-confirmed Graves’ disease. The features of regulatory T-blood cells subpopulation and the level of expression (MFI) CD25 surface receptor were studied by flow cytometry using direct immunofluorescence using monoclonal antibodies.RESULTS: The study included 36 women with recurrent Graves’ disease, middle age 46.34±14.32 years. In patients with Graves’ disease before and during the entire period after RIT a low percentage of naive (CD45R0-CD62L+) and terminally differentiated (CD45R0-CD62L-) Treg was established relative to the control, and on 3 and 6 months after RIT a significant decrease of cells with this phenotype was revealed relative to the values detected in patients before and 1 month after RIT (p&lt;0.001). Against the background of compensated hypothyroidism the most significant changes of expression CD25 receptor in patients with Graves’ disease were found on 3 and 6 months after RIT: reduced levels of MFI CD25 on surface of naive and terminally differentiated Treg.CONCLUSION: A decrease in the level of naive Treg was found (apparently due to a violation of differentiation processes in thymus) and terminally differentiated Tregs (due to maturation and survival processes), which are supplemented by a reduced expression of the CD25 receptor on the surface of these cells and do not depend on hyperthyroidism compensation, the titer of TSH receptor antibodies, previous conservative therapy with thiamazole and RIT. The obtained new data reveal the role of naive and terminally differentiated Treg subpopulations in immunopathogenesis and help to outline further ways to develop approaches for immunotropic therapy.

Epidermolysis bullosa in a twins infant: a rare case

Background: Epidermolysis bullosa (EB) is a rare hereditary genodermatosis characterized by blisters due to trauma and temperature. Cases of EB in twin infants are rare. This report will discuss EB in twin infants to improve our knowledge about this genodermatosis. Case: A baby boy with a twins history aged 4 months was consulted by the Pediatric Department with complaints of fluid-filled blisters that have been present since birth. During examination, bullae appeared on the upper and lower extremities. The gram examination and culture results showed Staphylococcus aureus infection and gentamicin sensitivity. The histopathology results showed a subepidermal blister with the dermis layer showed lymphocytic infiltration, which was in accordance with EB. The baby was hospitalized for 5 days and then came back to the outpatient unit with his twin, who had the same complaint. Examination of the second infant revealed multiple erosions and hypopigmented macules on the superior and inferior extremities. Both babies were born at term, normal, adequate weight, and are the first twins. Direct immunofluorescence did not show immunoglobulin G (IgG) and complement C3 deposits in the basement membrane zone. Both infants received symptomatic therapy. Conclusion: Epidermolysis bullosa is a rare case, especially in twins. Electron microscopy is a gold standard for determining EB type. Symptomatic treatment is the main therapy in this population of EB.

The Concentration of PCSK9-Lp(a) Complexes and the Level of Blood Monocytes in Males with Coronary Atherosclerosis.

In this study we analyzed the concentration of lipoprotein(a) (Lp(a)), PCSK9-Lp(a) complexes and the circulating monocyte subsets in coronary atherosclerosis. For this study, 257 patients with coronary atherosclerosis and 68 patients without stenotic atherosclerosis in the coronary, carotid and lower extremity arteries (control group) were enrolled. The monocyte subpopulations (classical CD14++CD16-, intermediate CD14++CD16+ and non-classical CD14+CD16++) were analyzed by direct immunofluorescence and flow cytometry. The Lp(a) and PCSK9-Lp(a) complexes in the serum were detected by ELISA. The concentration of Lp(a) was higher in the coronary atherosclerosis group compared with the controls (23.0 (9.1; 73.3) mg/dL versus 10.7 (4.7; 25.0) mg/dL, p < 0.05). No correlations between the level of Lp(a) and the concentration of the PCSK9-Lp(a) complexes, nor between the level of Lp(a) or PCSK9 and the total number of monocytes, were observed in either group. A slight positive correlation between the concentration of PCSK9-Lp(a) complexes and the absolute level of monocytes was obtained (r = 0.20, p = 0.002) in the patients with atherosclerosis due to the intermediate monocyte subsets (r = 0.33, p = 0.04). According to regression analysis, both the PCSK9-Lp(a) complexes concentration and BMI were related to the absolute number of blood monocytes in patients with atherosclerosis. Further studies are required to determine the pathogenetic contribution of PCSK9-Lp(a) complexes to the development of atherosclerosis.

Mechanisms of Long-Term Persistence of Mycoplasmas in Children with Asthma.

Improving the management of children with asthma associated with mycoplasma infection is important. Aim: To study the duration of the persistence of antigens, and DNA in a free state, in the structures of circulating immune complexes (CICs) and living cells of Mycoplasma pneumoniae (Mpn) and Mycoplasma hominis (Mh) in children with asthma. In total, 205 children with asthma from 1 to 14 years were observed. The reaction of aggregate-hemagglutination (AHAA), the direct immunofluorescence reaction (DIF), the reaction of the polymerase chain reaction (PCR), and the culture method were used. In addition, 47 children were re-examined 1.5 months after the treatment of mycoplasma infection with azithromycin. The number of samples positive for antigens and DNA in the free state and in the structures of CICs significantly decreased. Then, 50 blood serum samples containing Mh antigens, and 50 samples containing Mpn antigens were analyzed by culture method. Mh was isolated in 21 (65.5%) of 32 samples containing DNA. Mpn was isolated from antigen-positive samples in nine cases. The presented data indicate the long-term persistence of antigens, and DNA of mycoplasma cells in the free state, in the structure of CICs, as well as in the form of "microcolonies". A high level of CICs can be used to predict the course of the disease and the response to therapy.

P19 Diagnostic conundrums in paraneoplastic pemphigus

Abstract Paraneoplastic pemphigus (PNP) is a subset of pemphigus, a group of autoimmune blistering skin disorders. A 51-year-old man presented with paraneoplastic pemphigus associated with a hepatic neoplasm. He described an epigastric mass and associated mucositis affecting the orogenital mucosae, as well as the sclerae. Both computed tomography and magnetic resonance imaging demonstrated a 14-cm mass arising from the left lobe of the liver. A liver biopsy showed features of an atypical spindle-cell neoplasm, but a definite diagnosis was not possible. A diagnostic skin biopsy was taken from his outer lip mucosae. Histology showed surface ulceration with an acute inflammatory infiltrate, lichenoid lymphoplasmacytic infiltrate with focal basal vacuolar change, degenerate keratinocytes and adjacent suprabasal acantholysis and spongiosis. Perilesional direct immunofluorescence (DIF) was negative; however, indirect immunofluorescence (IIF) showed intercellular IgG antibodies on transitional epithelium with a high titre of 1 : 800 to rat bladder. These finding were consistent with the suspected clinical diagnosis of paraneoplastic pemphigus. The patient was commenced on oral prednisolone 1 mg kg−1 daily. Topical therapy included budesonide mixed with sweetener applied to the oral mucosae. Following this, a trial of 3 days of intravenous methylprednisolone along with 2% cocaine mouthwash, triple mouthwash (betamethasone, doxycycline and nystatin) and Dermovate® ointment was commenced, with improvement noted in pain, haemorrhage and ulceration. Intravenous immunoglobulin was administered preoperatively. The patient was subsequently transferred to a specialist centre for hepatobiliary surgery and listed for debulking of the liver mass. Postoperatively, he had plasmapheresis, with a noted reduction in mucosal ulceration and significant improvement in pain. Chemotherapy with doxorubicin and ifosfamide was initiated thereafter. PNP is a severe autoimmune blistering disorder characterized clinically by flaccid blisters and erosions. Although much is yet to be understood regarding the pathophysiology of PNP, studies show that the development of autoantibodies to the plakin family and desmogleins 1 and 3 play a particular role in its pathogenesis. In addition to autoantibodies, CD8+ T cells have also been shown to contribute to the development of PNP. In adults, the majority of cases are associated with an underlying haematological malignancy and, rarely, a solid-organ tumour. DIF findings include IgG and C3 deposition along the intercellular and basement membrane zones, while IIF detects autoantibodies to proteins found in transitional or stratified squamous epithelium. As in our case, reports have shown that the DIF can be negative, with one study showing a positive DIF in only 41% of cases. This case highlights the complex diagnostic and treatment challenges practitioners face when dealing with PNP and the patients it affects.

003 Utility of plucked hair direct immunofluorescence as a possible replacement for conventional skin biopsy direct immunofluorescence in immunological activity monitoring and prediction of relapse in pemphigus

Abstract Pemphigus vulgaris (PV) is a potentially lethal chronically relapsing autoimmune bullous disorder with intraepithelial lesions affecting the skin and mucous membranes. The prediction of clinical relapse has been studied with conventional skin biopsy and plucked hair direct immunofluorescence (DIF; Daneshpazhooh M, Naraghi ZS, Ramezani A et al. Direct immunofluorescence of plucked hair for evaluation of immunologic remission in PV. J Am Acad Dermatol 2011; 65:173–7) which can be of use in timing treatment withdrawal and helping with better management. The aims of our study were to determine the association of plucked hair root DIF positivity status at baseline with clinical relapse within the study duration, and to determine the correlation between skin biopsy and plucked hair DIF. Seventy-eight consecutive patients diagnosed with PV in clinical remission on therapy, on minimal therapy or off therapy, were included after obtaining informed consent. Three-monthly follow-up was carried out for 12 months. Immunological monitoring with plucked hair DIF was done at baseline and repeated every 3 months for 1 year or a relapse was experienced, whichever occurred earlier. Conventional skin biopsy DIF was performed at baseline, at 3 months and at the time of relapse. In patients receiving adjuvant therapy during their present remission episode, discontinuation of therapy was done at inclusion, at 3 months and at 6 months, and occurrence of relapse in relation to the time point of discontinuation was assessed. Of 78 patients, 22 had disease relapse and 56 remained in complete remission. Clinical relapse during the study duration was significantly more frequent in patients who had higher mean Autoimmune Bullous Skin Disorder Intensity Score during the preceding active disease episode (P = 0.033), pruritus during the present clinical remission episode (P = 0.007) and early discontinuation (within 3 months of study inclusion) of concomitant adjuvant therapy (P = 0.007). Immunological parameters associated with clinical relapse were baseline IgG (P = 0.005) and C3 positivity (P = 0.047) in plucked hair, greater intensity of IgG and C3 positivity at baseline in plucked hair (IgG, P = 0.002; C3, P = 0.033) and skin biopsy specimens (IgG, P = 0.004; C3, P = 0.0.002), and positive hair DIF at the time of adjuvant therapy withdrawal (P = 0.017). Plucked hair DIF is a reliable technique to assess immunological remission in patients with PV who are in clinical remission and helps to guide treatment according to immunological remission status.

DP09 The lupus band test: a review of the sensitivity and specificity in the diagnosis of lupus erythematosus

Abstract The lupus band test (LBT) is a direct immunofluorescence (DIF) technique which shows a band of localized immunoglobulins at the dermoepidermal junction in lesional, non-sun-exposed skin of patients with both systemic (SLE) and cutaneous lupus erythematosus and in the perilesional skin of patients with SLE. However, low sensitivity and poor concordance between histological and clinical diagnoses warrant a review of the application of the LBT in the diagnosis of lupus erythematosus (LE). The aim of this study was to assess the sensitivity and specificity of the LBT in diagnosing LE following clinicopathological correlation (CPC). All cases sent to our pathology department between 2011 and 2018 for DIF with a clinical query of LE were reviewed. Data collection included demographic details, pathology requests, histology and DIF reports, clinical reports and diagnoses and serology. Of 256 histology requests, 9.0% (n = 23) had a positive LBT. This was discordant with the prevalence of LE diagnosis, as 46.3% were diagnosed with LE following CPC. The sensitivity and specificity of the LBT for LE were 17.6% and 98.8%, respectively, with a positive predictive value of 92.9% and negative predictive value of 58.2%. The LBT is not a sensitive diagnostic test for LE, but is highly specific and should be considered as a supportive diagnostic tool for LE. This is the largest reported case series to evaluate the efficacy of the LBT in the diagnosis of LE.

CPC02 Delayed widespread lichenoid drug reaction with radioisotopic response in a patient treated with nivolumab

Abstract The introduction of immune checkpoint inhibitors for the treatment of a wide variety of malignancies has led to a rise in cutaneous immune-related adverse events (irAEs), including autoimmune and autoinflammatory skin conditions, which typically present within the first 16 weeks of treatment—on average, 3.6 weeks after drug initiation. Cutaneous irAEs occur in approximately 8.7% of patients receiving antiprogrammed cell death type 1 therapy (PD-1). Nivolumab is an antibody against PD-1 widely used for various cancers. As the application of these therapies expands, understanding of their adverse cutaneous effects is essential. We present a 76-year-old woman with metastatic renal cell carcinoma, who developed a widespread pruritic eruption 18 months after commencing nivolumab. Examination revealed a papular and vesicular rash affecting the arms, legs, anterior chest, face, hands and feet. There was a confluent well-demarcated rectangular area of grouped papules on the right side of her back; this site had been treated with radiotherapy for localized rib metastases 1 month prior to onset of rash. There was no mucosal involvement and no systemic symptoms. Three skin biopsies were taken, which showed vacuolar alteration of the basal layer and a band-like inflammatory cell infiltrate at the dermoepidermal junction, with focal areas of subepidermal separation. Direct immunofluorescence was negative, and skin swabs were negative for bacterial and viral infection. A diagnosis of delayed bullous lichenoid drug reaction secondary to nivolumab was made. The area involving the previous radiotherapy field was considered to be a radioisotopic response rather than a radiation recall response as she was already on nivolumab when it occurred. She discontinued nivolumab and commenced oral prednisolone and her skin settled over 5 weeks. Lichenoid dermatitis is a known adverse reaction associated with PD-1 therapy, with improved survival outcomes noted in patients with lichenoid irAEs on PD-1 therapy. Typically, it occurs within 16 weeks, with a documented range of 1–266 days. This patient developed a delayed lichenoid eruption at approximately 550 days, with concurrent radioisotopic response at a previous radiotherapy site. Radiotherapy-induced lichen planus is rare, with only one case report documenting involvement outside the irradiated field. We found no previous reports of nivolumab causing lichenoid drug eruption after such a protracted period. This case adds to the growing literature around PD-1 inhibitor-associated cutaneous eruptions.

DP14 Going against the grain: eosinophilic dermatosis presenting as erythema gyratum repens

Abstract Erythema gyratum repens (EGR) is a rare, paraneoplastic rash with a characteristic woodgrain appearance. It is heavily associated with malignancies, including lung, oesophageal and breast cancers. However, other conditions can mimic EGR, including leucocytoclastic vasculitis, immunobullous and papulosquamous conditions. We report an unusual presentation of eosinophilic dermatosis with an EGR-like pattern, not associated with malignancy. A 67-year-old woman presented with a 2-week history of a pruritic eruption on her legs. It initially started with three small circular erythematous macules on the right leg and one on the left leg. Over the course of a few days, the three macules evolved into large concentric annular patches simulating woodgrain, which raised the clinical suspicion of EGR. After a week, the patches began to blister with accentuation of some of the blisters on the edge of the annular patches. She denied any previous history of skin problems or changes in medication and was otherwise well with no systemic symptoms, including fever. In view of the almost 70% chance of an underlying malignancy, she was thoroughly investigated. Her full blood count, eosinophil count, liver function test, renal profile and serum C3 and C4 were normal. Skin basement membrane/desmosome antibodies and Borrelia burgdorferi IgG and IgM antibodies were also negative. Histology revealed prominent epidermal spongiosis with some corneal and intraepidermal vesicles that were cell-poor and contained only red blood cells. The underlying dermis showed prominent papillary dermal oedema with a brisk interstitial and superficial/deep perivascular mixed inflammatory infiltrate containing numerous eosinophils but no obvious flame figures. There was also eosinophil infiltration into the underlying subcutaneous fat. Direct immunofluorescence was negative. The histological pattern was that of an eosinophil-rich dermatitis with vesiculation; the history and morphology of the eruption were not compatible with bullous insect bites. An eosinophilic dermatosis such as eosinophilic cellulitis presenting with an EGR-like pattern was considered to be the most likely possibility for her presentation. Computed tomography of the neck, thorax, abdomen and pelvis, bidirectional endoscopy and mammography revealed no malignancies. She was treated with prednisolone 30 mg daily for 10 days with Elocon® ointment, and the eruption improved remarkably leaving postinflammatory colour changes that subsequently resolved. We would like to raise the awareness that an EGR-like pattern can be found in several other dermatoses, including eosinophilic dermatosis, and woodgrain pattern is not always equivalent to EGR or a high risk of associated malignancies (Table 1).Table 1Erythema gyratum repens-like dermatosesLeucocytoclastic vasculitisImmunobullousPapulosquamousBullous pemphigoidPsoriasisEpidermolysis bullosa acquisitaPityriasis rubra pilarisLinear IgA dermatoses

P13 A rare case of Sjögren syndrome in a White woman with Kikuchi–Fujimoto disease

Abstract A 36-year-old White woman was seen in May 2022 for a rash on her upper back, shoulders, cheeks, forehead and extensor surfaces of the upper limbs. It had developed following sun exposure on the beach despite regularly applying SPF50. Over time, the rash became sore but exhibited no signs of vesicles or oozing. Of note, the patient had been recently diagnosed with Kikuchi–Fujimoto disease (KFD) following a lymph node biopsy. She gave history of Raynaud phenomenon since childhood, and arthralgia primarily affecting the small joints of hands. She also experienced episodic dry eyes, dry mouth, peripheral oedema, dyspnoea, abdominal pain and constipation. For the previous month, she had felt fatigued with some undocumented weight loss. Clinical examination revealed an erythematous rash with discrete nonblanching macules and plaques of varying sizes, oval-to-circular in shape, on the involved areas. Lymph nodes were not palpable. Differential diagnosis included cutaneous lupus, other connective tissue disorders, and—less likely—polymorphic light eruption. Skin biopsy showed focal compact hyperkeratosis and parakeratosis with focal subtle spongiosis, intraepidermal neutrophils and lymphocyte exocytosis, with the presence of perivascular and periappendageal chronic inflammation. There was mild basement membrane thickening with focal deposition of fibrinogen at the basement membrane zone on direct immunofluorescence. She was positive for anti-Ro antibodies, with a titre of 84 kU L−1 (normal &amp;lt; 11 kU L−1). A diagnosis of Sjögren syndrome with KFD was made. The patient was advised on photoprotection with a high-potency topical steroid, and referred to rheumatology for review of her joint problems. After 4 weeks, the patient had only some improvement with topical therapy and was started on oral hydroxychloroquine. At the 12-week follow-up, there was significant improvement. Her skin was clear, with improvement in joint pain and fatigue. KFD, also known as necrotizing lymphadenitis, is a benign and self-limited disorder, characterized by regional cervical lymphadenopathy with tenderness, usually accompanied with mild fever and night sweats. Rarely, the skin and joints are involved. There is an established association between KFD and connective tissue disorders, especially Sjögren syndrome (Gates GA, Li Y, Magyar C, Sarantopoulos GP. Kikuchi–Fujimoto disease with unusual cutaneous findings. Am J Dermatopathol 2021; 43:e213–17). This is more prevalent among Asians and rarely reported in White people (Perry AM, Choi SM. Kikuchi-Fujimoto disease: a review. Arch Pathol Lab Med 2018; 142:1341–6), where it may cause diagnostic difficulties. This case highlights the importance of lateral thinking while formulating a differential diagnosis, despite the time pressures of a busy clinic. Although the reported prevalence of KFD among White people has been negligible (Perry et al.), it should not be dismissed solely based on a patient’s ethnicity. Timely and adequate management of rare conditions can reduce patient morbidity and improve their quality of life.

DP12 Painful oral keratosis and skin lesions: a paraneoplastic diagnostic conundrum

Abstract A 70-year-old man was referred to oral medicine with extensive, painful oral hyperkeratosis affecting the tongue, buccal mucosa and palate. Medical history included a previous diagnosis of prostate cancer. Histology from multiple oral biopsy specimens showed only hyperparakeratosis and acanthosis, with no fungi or dysplasia. Owing to the absence of oral inflammation, the pain was treated as neuropathic. The patient then developed skin lesions and dermatology review was requested. Skin examination revealed marked hyperkeratotic, verrucous plaques affecting the extensor surfaces of the limbs and dorsal hands, as well as the plantar feet bilaterally. The nails were unaffected. There was violaceous discolouration of the pinnae. Given the history of prostate cancer, a paraneoplastic process was suspected. Two incisional biopsies were taken, which showed epidermal acanthosis with necrotic and dyskeratotic keratinocytes throughout the epidermis. There was a dense eosinophilic inflammatory infiltrate. Direct and indirect immunofluorescence was negative. The appearances were suggestive of a paraneoplastic phenomenon, including acrokeratosis paraneoplastica. Haematological investigations identified an IgG monoclonal paraproteinaemia with raised kappa:lambda ratio. Following further investigation, including bone marrow biopsy and positron emission tomography computed tomography, a diagnosis of monoclonal gammopathy of undetermined significance was made by haematology. A repeat bone marrow biopsy 7 months later confirmed evolution to myeloma. As CRAB criteria (hypercalcaemia, renal insufficiency, anaemia and bone lesions) were absent, a period of observation followed. A trial of oral steroids improved the cutaneous eruption, but he developed significant side-effects. Oral mycophenolate, methotrexate and acitretin were used as steroid-sparing agents, with a poor clinical response. The skin lesions progressed to a widespread psoriasiform eruption on the trunk and lower limbs with the formation of bullae. Further biopsies showed an eosinophil-rich infiltrate with negative immunofluorescence, raising the possibility of an eosinophilic dermatosis of haematological malignancy. After 2 years, given the worsening skin symptoms, it was decided to commence bortezomib, cyclophosphamide and dexamethasone (VCD) chemotherapy. Despite four cycles of VCD treatment, the skin lesions worsened and as the paraprotein levels did not reduce, treatment was stopped. Subsequently, further elevations in serum free light chains prompted treatment with lenalidomide. This resulted in a marked improvement in the skin and reduced pain from the oral lesions, as well as a reduction in serum free light chains. Acrokeratosis paraneoplastica (Bazex syndrome) is characterized by digital hyperkeratosis and paronychia, which may progress to involve the limbs and trunk. Eosinophilic dermatosis of haematological malignancy is characterized by a pruritic, papular skin eruption with a vesiculobullous component. This case presents a diagnostic conundrum, with evolving features of two different paraneoplastic conditions.

P09 A case of suspected COVID-19 vaccine-induced antiphospholipid syndrome

Abstract A 50-year-old man presented with an acute onset, painful leg ulcer affecting his right calf. This developed 2 weeks after his second COVID-19 vaccine. It started with a painful erythematous plaque, which evolved slowly into a deep-seated 5-cm ulcer over a 6-week period. A thorough review was unremarkable. He had no significant medical history and was a keen athlete and nonsmoker. His father had an unspecified stroke in his 80s. Examination revealed no signs of chronic venous insufficiency or discoloration to his foot, and his neurovascular examination was unremarkable. Investigations for antineutrophil cytoplasm antibodies, antinuclear antibody, complement, paraprotein, HIV and hepatitis were all unremarkable. Biopsy demonstrated nonspecific inflammatory changes, and wound swabs were unremarkable. Initial treatment with oral antibiotics, regular dressings, compression, topical emollients, Dermovate® and prednisolone (0.5 mg kg−1) was only partially effective. He subsequently developed additional painful superficial erosions and ulcers affecting the same limb involving the calf and shin, with a surrounding erythematous livedoid-like rash involving his shin. Rebiopsy demonstrated superficial and deep dermal and subcutaneous thrombosis without vasculitis. Direct immunofluorescence showed the presence of fibrinogen within dermal vessels. Further investigations demonstrated the presence of lupus anticoagulant. Factor V Leiden, protein C and S, anti-B2-GP1 and anticardiolipin antibodies, antithrombin III, homocysteine and coagulation profile were all unremarkable. Histological confirmation of thrombosis and the presence of serum lupus anticoagulant confirmed the diagnosis of antiphospholipid syndrome (APS). He was started on warfarin and the ulcers improved. APS is a rare autoimmune disorder characterized by recurrent miscarriage, thrombophilia and the presence of antiphospholipid autoantibodies (either serum lupus anticoagulant, anti-B2-GP1 or anticardiolipin autoantibodies). Positivity for APS antibodies can be found in up to 5% of the general population. An individual must have the clinical features and antibody positivity to be diagnosed with APS. The temporal relationship with his second COVID-19 vaccine raises the possibility of vaccine-induced APS. COVID-19 vaccine-induced APS is limited to just a few case reports (Moreno-Torres V, Gutiérrez A, Valdenbro M et al. Catastrophic antiphospholipid syndrome triggered by mRNA COVID-19 vaccine. Clin Exp Rheumatol 2022; 40:1054–5). It is postulated that mRNA vaccines may induce a rare complex immunogenic response characterized by a type 1 interferon reaction. This results in a prothrombotic cascade in individuals with pre-existing APS autoantibody positivity (Talotta R, Robertson E. Antiphospholipid antibodies and risk of post-COVID 19 vaccination thrombophilia: the straw that breaks the camel’s back? Cytokine Growth Factor Rev 2021; 60:52–60).

PA04 (P05) A rare presentation of blistering in an adolescent

Abstract A previously well 16-year-old male presented with florid and widespread tense bullae and erosions. He gave a 2-week history of an erythematous rash consisting of crusted erosions that started on his chest and face. He denied any new medications or over-the-counter preparations. There was no preceding systemic illness, and no history of cold sores or sexual contacts. In the 24 h prior to his admission, his rash had become widespread, involving his trunk, limbs, hands, ears, face, lips, oral and genital mucosa. He had started to feel systemically unwell and was in significant pain. On examination, he had a combination of heavily crusted erosions and extensive tense bullae containing straw-coloured fluid on a background of inflamed skin. The morphology was herpetiform in some areas. There was notable extension of bullae onto the palms and penis, as well as intraoral and labial erosions. Viral skin swabs were negative, and his biopsy subsequently demonstrated subepidermal blistering with neutrophil-rich inflammation and positive direct immunofluorescence (IMF) for C3 and IgG only. Serum autoantibodies were negative. Childhood bullous pemphigoid (BP) was initially the working diagnosis, but indirect IMF showed localization to the base of the blister and anticollagen VII antibodies later proved positive, consistent with a diagnosis of epidermolysis bullosa acquisita (EBA). Given the clinical features, it was classified as BP-like EBA. He was initially treated with high-dose oral prednisolone, superpotent topical steroids and analgesia as an inpatient. Mycophenolate mofetil (MMF) was initiated, and his corticosteroid dose was slowly tapered. His condition was completely controlled with MMF 500 mg twice daily at 4 months. The patient then elected to stop taking MMF and remains in complete remission at 7 months. The EBA is rare in adults and even more so in children. The mean age of onset is 44–54 years, but up to 5% of reported cases are in children/adolescents. Four subtypes are described: classic mechanobullous, BP-like, mucous membrane pemphigoid-like and linear IgA bullous dermatosis-like. The BP-like EBA is characterized by widespread vesicles and tense bullae, commonly on an inflammatory background and lacking skin fragility, scarring or milia. Oral and genital mucosae are frequently affected in paediatric cases of EBA. Various steroid-sparing treatment modalities are described, including dapsone, colchicine and MMF. The prognosis in the paediatric population is favourable compared with that of adults: the majority achieve complete remission or complete control on treatment.

DP17 A very rare presentation of linear IgA disease during pregnancy

Abstract We present a very rare immunobullous dermatosis, linear IgA disease (LAD), that developed during pregnancy, to highlight its clinical features, management and important safety considerations for the neonate. During her first pregnancy, a 32-year-old Japanese woman of 27 weeks’ gestation developed an intensely itchy blistering rash on her limbs, trunk and face without periumbilical predilection. The blisters were tense, clustering in an annular configuration, with subsequent development of blistering and erosions involving the oral and genital mucosae. Medications included a pregnancy multivitamin only. A lesional skin biopsy revealed a subepidermal blister containing neutrophils and eosinophils. Direct immunofluorescence of perilesional skin demonstrated linear deposition of IgA along the basement membrane zone. Linear IgA disease was diagnosed. Initial treatment consisted of erythromycin and daily clobetasol ointment. Owing to inadequate symptom control and blister progression, dapsone 50 mg daily was initiated with prompt improvement in her condition and cessation of new blistering. A postpartum flare necessitated a dose increase to 75 mg daily. Her baby’s glucose-6-phosphate dehydrogenase (G6PD) level was found to be normal at birth. She proceeded to breastfeed her son, and monitoring of his haemoglobin level was normal. Although her clinical presentation initially raised the possibility of pemphigoid gestationis (PG), there was no periumbilical involvement, and the morphology of blistering in conjunction with mucosal involvement was more suggestive of LAD. The immunofluorescence results also helped to distinguish LAD from PG. The new onset of LAD during pregnancy is very unusual and has been rarely reported (Ikkaku N, Tateishi N, Oda Y et al. Linear immunoglobulin A bullous dermatosis developing during late pregnancy. J Dermatol 2017; 44:e44–5). Pre-existing LAD typically improves during pregnancy, although postpartum flares are recognized. Pemphigoid gestationis has been associated with an increased risk of prematurity and intrauterine growth restriction, but this has not been reported with LAD during pregnancy, possibly due to its rarity. Both the diagnosis of LAD and its mainstay treatment—dapsone—have important potential implications for the neonate. As for other immunobullous diseases, transplacental transfer of pathogenic antibodies may result in transient, usually mild, neonatal blistering. Although dapsone can be safely used during pregnancy and breastfeeding, the risk of neonatal haemolysis and methemoglobinaemia necessitates the assessment of the G6PD level and haemoglobin monitoring in the neonate. Multidisciplinary input from the dermatology, obstetric and paediatric teams is crucial to ensure appropriate counselling and care of the affected mother and baby.

P05 A rare presentation of blistering in an adolescent

Abstract A previously well 16-year-old male presented with florid and widespread tense bullae and erosions. He gave a 2-week history of an erythematous rash consisting of crusted erosions that started on his chest and face. He denied any new medications or over-the-counter preparations. There was no preceding systemic illness, and no history of cold sores or sexual contacts. In the 24 h prior to his admission, his rash had become widespread, involving his trunk, limbs, hands, ears, face, lips, oral and genital mucosa. He had started to feel systemically unwell and was in significant pain. On examination, he had a combination of heavily crusted erosions and extensive tense bullae containing straw-coloured fluid on a background of inflamed skin. The morphology was herpetiform in some areas. There was notable extension of bullae onto the palms and penis, as well as intraoral and labial erosions. Viral skin swabs were negative, and his biopsy subsequently demonstrated subepidermal blistering with neutrophil-rich inflammation and positive direct immunofluorescence (IMF) for C3 and IgG only. Serum autoantibodies were negative. Childhood bullous pemphigoid (BP) was initially the working diagnosis, but indirect IMF showed localization to the base of the blister and anticollagen VII antibodies later proved positive, consistent with a diagnosis of epidermolysis bullosa acquisita (EBA). Given the clinical features, it was classified as BP-like EBA. He was initially treated with high-dose oral prednisolone, superpotent topical steroids and analgesia as an inpatient. Mycophenolate mofetil (MMF) was initiated and his corticosteroid dose was slowly tapered. His condition was completely controlled with MMF 500 mg twice daily at 4 months. The patient then elected to stop taking MMF and remains in complete remission at 7 months. EBA is rare in adults and even more so in children. The mean age of onset is 44–54 years, but up to 5% of reported cases are in children/adolescents. Four subtypes are described: classic mechanobullous, BP-like, mucous membrane pemphigoid-like and linear IgA bullous dermatosis-like. BP-like EBA is characterized by widespread vesicles and tense bullae, commonly on an inflammatory background and lacking skin fragility, scarring or milia. Oral and genital mucosae are frequently affected in paediatric cases of EBA. Various steroid-sparing treatment modalities are described, including dapsone, colchicine and MMF. The prognosis in the paediatric population is favourable compared with that of adults: the majority achieve complete remission or complete control on treatment.

P12 A flare of dermatitis herpetiformis precipitated by etanercept

Abstract We report the case of a 79-year-old man who presented with an acute, intensely pruritic, widespread rash, which, on examination, consisted of symmetrical, haemorrhagic nodules and vesicles with some desquamation affecting the extensor surfaces of the arms, flexor surfaces of the legs and the natal cleft. He had a history of coeliac disease (for which he had adhered to a strict gluten-free diet since 1997) and rheumatoid arthritis. He had his first dose of the anti-tumour necrosis factor (TNF) agent etanercept prior to this episode to manage his arthritis. A punch biopsy for histopathology showed subepidermal blisters with a predominance of neutrophils. Eosinophils were also present to the level of the eccrine coils within the dermis, with eosinophilic flame figures but no eosinophil spongiosis at the edges of the blister. Direct immunofluorescence showed granular IgM and IgA in the dermal papillae, and a similar reaction was seen with fibrinogen. These histological findings were strongly suggestive of dermatitis herpetiformis (DH). Owing to diagnostic uncertainty on presentation, initial treatment was with high-dose topical steroids and emollient. and a yellow card was completed. Since this presentation, he has had no further flares and has not required dapsone. Multidisciplinary team discussion decided that the patient should avoid other anti-TNF treatments, but he has found relief for multiple months with rituximab and has had no cutaneous adverse events on this agent. The aetiology of DH following anti-TNF treatment is not immediately clear. Increased immunogenicity and cytokine release have been hypothesized to be part of the pathology of many adverse reactions to anti-TNF agents. It is conceivable that the cytokine release ordinarily seen in DH is replicated by the administration of anti-TNF agents. There have been many cases of anti-TNF agents leading to worsening eczema and psoriasis. Interestingly, these conditions have been found to upregulate TNF-α, a feature that is also seen in DH. This appears to be the first case described in the literature of etanercept-induced DH. A reaction with infliximab was reported in 2008, with a similar rash and histology consistent with DH, but it differed as the patient had no prior diagnosis of coeliac disease and presented after their second dose (Marakli SS, Uzun S, Ozbek S, Tuncer I. Dermatitis herpetiformis in a patient receiving infliximab for ankylosing spondylitis. Eur J Dermatol 2008; 18:88–9).

Experimental Epizotology of Low-Virulent Variants of African Swine Fever Virus

African swine fever (ASF) remains an urgent problem of pig farming in Ukraine, the solution of which is possible only on the basis of deep scientific knowledge about the specific driving forces of the epizootic in its specific nozoareal. This is necessary in order to target anti-epizootic measures on the most vulnerable link of the epizootic chain in a specific nozoareal. The aim of the work was to develop a low-budget methodological base for experimental epizootology of low-virulent ASFV variants in Ukraine, in particular, to study the mechanisms of the formation of enzootic areas, quality control of anti-epizootic measures, and evaluation of the effectiveness of the antiviral drugs against them in Ukraine. Methods. Experimental and epizootological studies in the adaptation of suckling piglets to keeping in the biosecurity-level BSL-3 for laboratory animals (Patent UA No. 133248 dated 03/25/2019) were conducted at the laboratory base of the Odesa branch of NSC «IECVM». All procedures with infectious active biological materials in the current order were carried out in the BSL-3 module, built and certified with the assistance of the US Government in UAPRI (Odesa). The ASF agent strain «IECVM/Ternopil/2017» (infectious activity 4.0—7.5lg HAdU50/cm 3) circulating in the Ternopil region in 2017—2020 was used as a test virus. The presence of low-virulence variants of the ASFV pathogen in the studied samples was determined by a bioassay on suckling piglets, followed by three consecutive passages on a stable Vero line of the baby green monkey kidney cells. The isolated ASF virus was identified according to the methods and reagents recommended by the OIE Manual. Results. Intermittent passages «by the founder’s method» of dilutions 10-1 and 10-2 of the ASF virus strain «IECVM/Ternopil/2017» on piglets (n=20) and the culture of porcine alveolar macrophages («ASFVPAM») allowed us to identify highly-, moderately-, and low-virulent variants/clones in its composition. Verifi cation by bioassay on suckling piglets (n=5) of low-virulent clones of the agent, which were stabilized in Vero cell culture («ASFVVero»), showed that after intraperitoneal infection at a dose of 4.25 lgHAdU50/cm 3, they are capable of causing only a non-lethal (within 2 weeks) viral infection with a maximum daily rectal temperature of 39.4±0.22 °C and duration of fever on average 1.6±0.14 days (5 of 5 piglets). Clones with greater virulence («ASFVPAM») under similar conditions were able to cause a lethal infection with a maximum temperature of 40.7±0.37°C and duration of fever on average 3.9±0.27 days (17 of 20 piglets). Low-virulent clones were revealed by direct immunofluorescence in pulmonary and spleen smears of clinically healthy piglets on days 14 and 17 post-infection (p.i.); their antigens were visualized in Vero cells by indirect immunoperoxidase method after 48 h p.i. at dose about 0.01 lg HAdU50/cm 3. They caused «crumbly» hemadsorption of infected Vero cells and their virions had typical for Asfarvirus view and size (210—220 nm). The obtained data served as the basis for analysis of the mechanism of rooting ASF agents in West Podillia enzootic foci, as well as for implication of the concept of low-budget quality control of anti-epizootic measures and evaluation of antiviral drugs’ activities against ASF. Conclusions. Low-cost operational procedures have been developed that allow one to use a vivarium of laboratory animals for ASF bioassay and meet principal requirements for science-based research in important aspects of experimental ASF epizootology. With their help, confirmation of the heterogeneity of the population of the ASF virus circulating in endemic foci of the Ukrainian Western Podillia was obtained (p&lt;0.05, n=25)&gt;&lt;0.05, n=25). The developed methodological approach is suitable for the study of fundamental issues of ASF epizootology, as well as for the quality control of anti-epizootic measures against ASF. In particular, it is advisable to use it to improve the biosecurity of agricultural export programs in Ukraine, a country that is disadvantaged by ASF.

Fosfomycin modifies the replication kinetics of bovine alphaherpesvirus-1 and reduces the timing of its protein expression on bovine (MDBK) and human (SH-SY5Y) cell lines.

Bovine alphaherpesvirus 1 (BoAHV-1) predisposes cattle to respiratory secondary bacterial infections, which can be treated with the broad-spectrum antibiotic fosfomycin. This drug also suppresses NF-kB activity and pro-inflammatory responses. Therefore, cattle may be exposed to an interaction between the virus and the antibiotic which may have effects on it. The aim of this study was to determine the effect of calcium fosfomycin (580µg/mL) on BoAHV-1 (moi = 0.1) replication. Two cell lines (MDBK and SH-SY5Y) were used in this study. Our results show that fosfomycin has novel properties. By MTT assay we have shown that it is non-cytotoxic for any of the cell lines. Extracellular and intracellular viral titers demonstrated that fosfomycin has a cell-type and time-dependent effect on BoAHV-1 replication. By direct immunofluorescence it was shown that it reduces the timing of BoAHV-1 protein expression, and by qPCR, we found that its effect on NF-kB mRNA expression depends on the cell type.

PLASMA CELL GINGIVITIS TRIGGERED BY TOOTHPASTE

Plasma cell gingivitis is a rare benign condition characterized by the inflammatory infiltration of plasma cells, secondary to a hypersensitivity reaction to allergens found in oral hygiene products. This study aimed to present the case of a 48-year-old female, who was admitted to our clinic complaining of an intense gingival discomfort for the past six months, with the onset of symptoms occurring after using a new toothpaste. Clinical examination revealed generalized gingival erythema and edema. An incisional biopsy was performed, which, through direct immunofluorescence, revealed connective tissue densely permeated by interstitial lymphoplasmohistiocytic inflammatory cells, a condition that must be distinguished from other autoimmune diseases. The diagnosis of plasma cell gingivitis was concluded with a detailed history, differential diagnoses, and histopathology. The elimination of the etiologic factor, namely the toothpaste, and the use of topical corticosteroids improved both the lesions and the symptoms. Plasma cell gingivitis is a rare benign condition characterized by the inflammatory infiltration of plasma cells, secondary to a hypersensitivity reaction to allergens found in oral hygiene products. This study aimed to present the case of a 48-year-old female, who was admitted to our clinic complaining of an intense gingival discomfort for the past six months, with the onset of symptoms occurring after using a new toothpaste. Clinical examination revealed generalized gingival erythema and edema. An incisional biopsy was performed, which, through direct immunofluorescence, revealed connective tissue densely permeated by interstitial lymphoplasmohistiocytic inflammatory cells, a condition that must be distinguished from other autoimmune diseases. The diagnosis of plasma cell gingivitis was concluded with a detailed history, differential diagnoses, and histopathology. The elimination of the etiologic factor, namely the toothpaste, and the use of topical corticosteroids improved both the lesions and the symptoms.