Atypical infectious mononucleosis in the cervical region mimicking lymphoma in an adult: a case-based literature review

Infectious mononucleosis (IM) is an infectious clinical entity commonly seen in adolescence and early adulthood. It is caused in the majority of cases by Epstein-Barr virus (EBV) and it presents as pharyngitis, fever, and lymphadenopathy. As the viruses mainly target the lymphocytes and reticuloendothelial system of the body, leading to the proliferation of the lymphatic tissues, it is often mistaken as lymphoma. Most cases of IM can be easily distinguished from lymphoma by clinical presentations and laboratory ndings. However, atypical clinical presentations or laboratory ndings occasionally occur, including the ages of patients over 30, generalized or isolated lymphadenopathy at unusual sites, negative heterophile antibody tests, absence of atypical lymphocytosis in the peripheral blood smear, etc. These atypical cases may lead to confused diagnosis. Therefore, IM should be differentiated from malignant lymphoma. In this paper, we reported a case of atypical IM in the cervical region of an adult, with elder age and longer disease course than typical IM.


Abstract
Background Infectious mononucleosis (IM) is an infectious clinical entity commonly seen in adolescence and early adulthood. It is caused in the majority of cases by Epstein-Barr virus (EBV) and it presents as pharyngitis, fever, and lymphadenopathy. As the viruses mainly target the lymphocytes and reticuloendothelial system of the body, leading to the proliferation of the lymphatic tissues, it is often mistaken as lymphoma. Most cases of IM can be easily distinguished from lymphoma by clinical presentations and laboratory ndings. However, atypical clinical presentations or laboratory ndings occasionally occur, including the ages of patients over 30, generalized or isolated lymphadenopathy at unusual sites, negative heterophile antibody tests, absence of atypical lymphocytosis in the peripheral blood smear, etc. These atypical cases may lead to confused diagnosis. Therefore, IM should be differentiated from malignant lymphoma. In this paper, we reported a case of atypical IM in the cervical region of an adult, with elder age and longer disease course than typical IM.

Case presentation
In this paper, we reported a case of IM in the cervical region of a 31-year-old man, and reviewed the recent literatures on the pathogenesis, as well as differential diagnosis of IM.

Conclusion
When facing enlarged tonsil or cervical lymph nodes, the pathologists should increase vigilance to the differential diagnosis between infectious diseases and lymphomas. Especially in ambiguous cases, it is essential to test for EBER by in situ hybridization to rule out IM before making a preliminary diagnosis of lymphoma.

Background
Infectious mononucleosis (IM) is a benign and self-limited lymphoproliferative disease mainly caused by Epstein-Barr virus (EBV) infection with a course of two to three weeks 1 . IM occurs mainly in adolescents and young adults 2 . Based on the current literature, 44% of IM patients present as triad of pharyngitis, cervical lymphadenopathy, and fever. Besides, some patients can also present hepatosplenomegaly (60.1%), tonsillar white coat (39.3%), headache (25.6%), cough (13.1%), abdominal tenderness (12.5%), abdominal pain (9.5%) and facial edema (8.3%) 3 . For most cases of IM, the diagnosis depends on clinical presentations and laboratory ndings 4 . Clinical signs that make the diagnosis more likely include triad of pharyngitis, fever, and lymphadenopathy 2 . Serological diagnosis requires either (1) a positive anti-viral capsid antigen (VCA)-immunoglobulin M (IgM) and anti-VCA IgG, or (2) a positive monospot test and anti-VCA IgG, or (3) a positive monospot and anti-VCA IgM 5 . These clinical manifestations and laboratory examinations can easily lead to the diagnosis of IM. While, some patients with atypical presentations, including large or asymmetric lymph nodes or tonsillar masses (frequently out of the sizes of pharyngitis/tonsillitis), may be biopsied to rule out lymphoma 6-8 . However, owing to the presence of extensive proliferation of immunoblasts and atypical Reed-Sternberg-like cells, IM is usually misdiagnosed as lymphoma in biopsies 9,10 . To prevent pitfalls during diagnosis and inappropriate treatments due to misdiagnosis, pathologists should consider the possibility of infectious mononucleosis, based on disease history, clinical presentations, pathologic ndings etc., before making a diagnosis of lymphoma.

Case Presentation
A 31-year-old man came to our hospital presenting with a mass in his left cervical region for 3 months.
Three months earlier, the patient had a routine physical examination, and a chest radiograph showed asymptomatic lymphadenopathy in his left submandibular region. Ten days earlier, he got a fever with the temperature of 37.6 ℃ (99.68 ℉), and sometimes he felt pain on his shoulder. These symptoms were relieved after anti-in ammatory treatment (the name of the medicine was unknown), but the size of the mass kept unchanged. The patient was in good status without splenomegaly, rashes, or hepatomegaly.
Physical examination revealed a 25 mm × 15 mm mass in the left submandibular region with clear boundaries and good mobility. The mass was in slight tenderness on palpation. Color Doppler revealed multiple lymph nodes in the left submandibular region and around the left internal jugular vein. An initial diagnosis of tuberculosis was made. The patient was then proceeded to surgery in our hospital. During the surgery, the mass was found in the left submandibular region, measuring 25 mm × 15 mm. The mass was sent for routine pathological examination.
Macroscopically, the mass was irregular in shape, measuring 28 mm × 15 mm × 10 mm, with intact capsule. On cross-section, it was solid, medium-textured and eshy. Microscopically, it was a lymph node with an intact capsule and proliferative lymphocytes ( Fig. 1a). At high magni cation, there was expansions of the interfollicular areas by polymorphous in ltrations of small-, medium-, and large-sized lymphocytes, and occasionally histiocytes, resulting in expansion and distortion, but not obliteration of the architecture of the lymph node. The in ltrated cells within the interfollicular zones exhibited mild abnormalities, with mitotic gures easily seen (Fig. 1b).
Differential diagnosis of lymphoproliferative diseases was considered, and a panel of immunohistochemical stainings were performed using EnVision system. T cells were positive for CD2, CD3, and CD5 (Fig. 2a, 2b and 2c). B cells were positive for CD20 (Fig 2d). The patterns of the positivities of CD3 and CD20 immunostainings demonstrated the mixed nature of small-, medium-, and large-sized lymphocytes, including transformed active form of immunoblasts and mature plasma cells. Germinal centers were positive for Bcl-6 ( Fig. 2e) and CD10 (Fig. 2f), but negative for Bcl-2 (Fig. 2g). Follicular dendritic cell (FDC) networks were delineated by the positivity of CD21 (Fig. 2h. More than 60% of the cells within the interfollicular regions were labeled by Ki-67 (Fig. 2i).
Due to the presence of enlarged T zones and T cells, differential diagnosis of T-cell lymphoma was made.
However, the architecture of the lymph node was not obliterated and T cell-associated antigens (CD2, CD3, CD5) was not lost, thus T-cell lymphoma could be excluded. Then, in situ hybridization (ISH) for EBV-encoded small RNAs (EBER) was performed to rule out IM. Positive signals of EBER could be detected, with 200 positive cells/HPF (Fig 3a). EBER+ cells were predominantly small lymphocytes and large immunoblasts located within the interfollicular regions, as well as small lymphocytes located within the germinal center beneath the capsule (Fig 3b and 3c).
Based on the normal structure of the lymph node, proliferative lymphocytes of mixed cell types, and positive signals of EBER in both large and small cells, a nal diagnosis of IM was made. giving a mottled appearance at low magni cation. Mitotic gures and nuclear fragments are commonly detected. Increased small blood vessels, as well as necrosis can be seen in a few cases. In a minority of cases, small sheets of monocyte-like B cells can be found 6 . Immunohistochemical pro les revealed that CD3 + T cells are the major small lymphocytes within the expanded interfollicular zone. Active lymphoidlike blasts and immunoblasts which are CD20 and CD30 positive are scatteredly distributed, and are partially positive for CD30 with variable intensity. In all EBV infectious cases, nuclear EBER positivity can be detected in large-, medium-and small-sized lymphocytes.

Discussion
Actually, the pathogenesis of IM is a continuous spectrum following EBV infection, and can be divided into four stages, i.e., latent stage, early stage, middle stage, and late stage.
(1) Latent stage: as EB viruses invade the body, they rst infect B cells, resulting in a massive proliferation of B cells and induction of humoral immune responses. To prevent the proliferation of B cells, cytotoxic T lymphocytes are activated. Thus, this stage presents as the proliferation of B lymphocytes in the lymphoid follicles and cytotoxic T lymphocytes in the interfollicular zone, with EBV-infected B lymphocytes less than 5%. Latent stage of IM is similar to toxoplasmic lymphadenitis. However, the features of toxoplasmic lymphadenitis not only include the presence of immunoblasts and plasma cells, but also include necrosis and Langerhans cells. PCR and serological tests can be helpful to distinguish the pathogens 14 .
(2) Early stage: this stage presents as the proliferation of the large-, medium-, and small-sized cells in the paracortex region, among which large B lymphocytes signi cantly proliferated (accounting for 5%~75%).
At T lymphocytes, such as T cell receptor beta-chain (TCR beta). Clonal rearrangement of genes can be helpful in the differential diagnosis 6, 18 .
As IM progresses, the interaction between T and B lymphocytes leads to enhanced activity of suppressor T cells and macrophages, which block the proliferation of B lymphocytes. Ultimately, both B and T lymphocytes are decreased. Thus, IM presents as a self-limited disease. The treatment strategy for IM includes supportive treatment, rest, and analgesics. Since the pathological changes of IM at different stages mimic HL or non-HL, misdiagnosis of IM as lymphomas is common, resulting in inappropriate treatments. Thus, the pathologists should consider the differential diagnosis of IM and lymphomas at different stages. It is believed that in situ hybridization using EBER probes is helpful for differential diagnosis of IM 2 .

Conclusion
In this study, we reported one IM arising in the left submandibular region, with elder age (31-year-old) and long disease course (3 months). A nal diagnosis of IM was made based on normal structure of the lymph node, proliferative lymphocytes of mixed cell types, and positive signals of EBER in both large and small cells by in situ hybridization. It suggested that when a lymph node is proliferative with mixed background, IM should be considered as differential diagnosis and in situ hybridization using EBER probes should be performed to rule out IM. The pathologists should avoid the misdiagnosis of lymphoma to prevent the patients suffering from unnecessary treatment and psychological burden.

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