一只犬耳廓病毒性斑块中检测到一种新的乳头瘤病毒序列 ...

王帆

Detection of a novel papillomaviral sequence in viral plaques confined to the pinna of a dog

一只犬耳廓病毒性斑块中检测到一种新的乳头瘤病毒序列

作者：John S. Munday | Geoff Orbell | Lynne Robinson

翻译：王思权 校对：王帆

 

Abstract

摘要

A raised plaque that contained histological evidence of papillomavirus infection and sequences from a novel papillomavirus type developed close to the ear canal of a 14- year- old West Highland white terrier. The plaque was excised, and further plaques developed within the same area of pinna.

在一只14岁的西高地白㹴犬耳道附近发现了一个凸起的斑块，组织学提示其中有乳头瘤病毒感染，并且是一种新型乳头瘤病毒序列。斑块被切除后，在耳廓的同一区域出现更多的斑块。

KEYWORDS

关键词

aural plaques, canine papillomavirus, dog, papillomaviruses, pigmented plaques, viral oncogenesis, warts

耳部斑块；犬乳头瘤病毒；犬；乳头瘤病毒；色素斑块；病毒性肿瘤；疣

 

INTRODUCTION

概述

Plaques resulting from papillomavirus (PV) infection are uncommon cutaneous lesions of dogs. Canine cutaneous plaques typically appear as slightly raised pigmented lesions that are most common on the ventrum and medial surfaces of the limbs, although plaques can become more widespread over the body. Unlike the commonly observed cutaneous or oral viral papillomas (warts) that are caused by PV types within either the Lambdapapillomavirus or Taupapillomavirus genera, plaques are caused by a number of closely related Chipapillomavirus PV types. It is thought that immune dysfunction could play a significant role in lesion development by allowing increased PV replication. Some breeds of dog, notably pugs, are predisposed to viral plaques, suggesting these breeds may have an inherited inability to mount an effective immune response against Chipapillomavirus infection.The development of plaques also may be influenced by immunosuppressive diseases or treatments.The present report describes plaques that were confined to one pinna of a dog. In addition to the unique location, the plaques also had unusual histological features and contained PV sequences from a putative novel Chipapillomavirus type.

由乳头瘤病毒(PV)感染引起的斑块是犬的不常见皮肤病变。犬皮肤斑块通常表现为轻微凸起的有色素病变，虽然最常见位于腹部和四肢内侧面，但斑块可以在全身广泛分布。与常见的由乳头瘤病毒-11或乳头瘤病毒-19属的PV型引起的皮肤或口腔病毒性乳头瘤(疣)不同，这种斑块是由许多与乳头瘤病毒-22PV密切相关的病毒引起的。人们认为由于免疫功能失调，让PV复制增加从而导致病变发展。一些品种的犬，特别是巴哥犬，易患病毒斑块，这表明这些品种可能遗传了无法对乳头瘤病毒-22感染产生有效免疫反应的能力。斑块的形成也可能受到免疫抑制疾病或治疗的影响。本报告描述了局限于一只犬的一个耳廓的斑块病例。除了独特的位置外，这些斑块还在组织学上具有不寻常的特征，并且推测其含有一种新的来自于乳头瘤病毒-22型的PV序列。

 

CASE REPORT

病例报告

A 14- year- old female spayed West Highland white terrier developed a small flat plaque close to the opening of the right external ear canal. The dog had a history of chronic otitis affecting this ear for the previous two years. Although the dog had undergone a bulla osteotomy approximately one year previously, she had continued to develop episodes of bacterial otitis which had been treated by flushing the ear and administering topical antimicrobials. The dog also previously had been diagnosed with allergic bronchitis and had been treated with 0.5 mg/kg prednisolone per os, once daily (Redipred; Aspen Pharmacare Australia) for approximately a year before presentation. An excisional biopsy of the lesion was taken for histological evaluation.

一只14岁的雌性已绝育西高地白㹴在右侧外耳道开口附近出现了一个小的扁平斑块。在过去的两年里，此犬的这只耳患有慢性耳炎病史。虽然此犬在大约一年前接受了鼓泡截骨手术，但她继续出现了细菌性耳炎，并通过冲洗耳道和外用抗菌药进行治疗。这只犬之前还被诊断为过敏性支气管炎，并在耳廓病变出现的一年前接受了每日一次口服0.5mg/kg泼尼松龙，治疗约一年。对病变部位进行切除活检以进行组织学评估。

 

Histological results revealed a well- demarcated area of variable epidermal hyperplasia that resulted in multiple raised plaques of marked hyperplasia separated by narrow areas with more mild changes. The epidermis was not folded. Cells within basilar areas appeared crowded; however, the majority of the thickening resulted from expansion of the granular layer due to the presence of keratocytes that had large quantities of clear or faintly granular basophilic cytoplasm. These cells also contained large central nuclei that occasionally were darkened and surrounded by a clear halo (koilocytes). While nucleoli often were prominent, intranuclear viral inclusions were not identified. The plaques were covered by a thick layer of parakeratosis. Increased lymphocytes and plasma cells were visible within the dermis underlying the plaque.

组织学结果显示了一个边界清晰、形成多个凸起斑块的可变表皮增生区，这些增生区被变化轻微的狭窄区域隔开。表皮未折叠。基底区细胞显得拥挤；然而，大多数增厚是由于角质形成细胞的存在导致颗粒层的扩张，角质形成细胞含有大量透明或微小颗粒的嗜碱性细胞质。这些细胞包含了偶尔较暗的，被清晰的光环（空泡细胞）包围的中央核。虽然核仁通常很突出，但核内未发现病毒包涵体。斑块处被一层很厚的角化不全细胞所覆盖。可见斑块下真皮层淋巴细胞和浆细胞增多。

 

The lesion was diagnosed as a probable viral hyperplastic plaque although, as a consequence of the orientation of the sample within the histological block, an early squamous cell carcinoma (SCC) could not definitively be excluded. Recurrence was observed in the same location around two weeks after the first surgery with a larger (0.5 cm diameter) central mass surrounded by smaller, less raised, plaques. The large mass continued to grow and three months after the initial surgery, this lesion was 2 cm in diameter and raised 0.5 cm from the surrounding skin of the pinna. The masses remained confined to the skin surrounding the opening of the external ear canal (Figure 1) and no additional plaques were visible elsewhere on the body. Owing to the possibility that the lesion could be an SCC, an additional sample was taken. Histological results revealed similar lesions as before (Figure 2), although clear visualisation of the basement membrane allowed exclusion of an SCC.

尽管病变被诊断为拟病毒性增生性斑块，但由于这是样本在组织学倾向的改变结果（orientation），不能明确排除早期鳞状细胞癌(SCC)。首次手术后约两周，斑块在同一部位复发，中心肿块较大(直径0.5 cm)，周围斑块较小，凸起程度较小。首次术后的3个月，肿物继续增大，直径2cm，距耳廓周围皮肤隆起0.5 cm。肿块仍局限于外耳道开口处周围的皮肤(图1)，身体其他部位未见其他斑块。由于病变可能是鳞状细胞癌，因此又取了一个样本。尽管这次基底膜清晰可见可以排除SCC，但组织学上仍显示了与之前相似的病变(图2)。

 

 

 

 

FIGURE 1 Cutaneous viral plaques. The plaques appear as multiple raised lesions on the inside surface of the pinna (arrows). Plaques were restricted to the skin adjacent to the opening of the external ear canal.

图1. 皮肤病毒斑块。斑块表现为耳廓内表面的多个凸起病变（箭头）。斑块局限于外耳道开口处附近的皮肤。

 

 

 

 

FIGURE 2 Cutaneous viral plaque, histological results. (a) The plaque consists of irregularly thickened epidermis covered by parakeratosis. Expansion of the epidermis is most marked within the stratum spinosum with keratinocytes enlarged by increased cytoplasm. (b) Keratinocytes within the plaques have increased qualities of vacuolated or granular grey-blue or clear cytoplasm. Cell nuclei are enlarged, central and prominent. Some cells contain darkened nuclei surrounded by a clear halo (koilocytes). Mitotic figures are visible within the hyperplastic epidermis (arrows).

图2. 皮肤病毒斑块，组织学结果。 (a) 斑块包含被角化不全细胞覆盖的不规则增厚表皮。表皮的扩张在棘层内最为明显，角质形成细胞因细胞质增多而增大。 (b) 斑块内的角质形成细胞具有更多的空泡化或颗粒状灰蓝色或透明的细胞质。细胞核增大、居中且突出。一些细胞含有被透明光晕包围的深色细胞核（空泡细胞）。增生表皮内可见有丝分裂（箭头）。

 

As a consequence of the histological evidence of PV infection, total DNA was extracted from formalin- fixed, paraffin- embedded samples of both excised plaques and the MY09/11 and CP4/5 consensus PCR primers were used to amplify PV DNA.6Both primer sets amplified PV DNA from both plaques. When the DNA that had been amplified by the MY09/11 primers was sequenced, the PV DNA sequences were identical from the initial plaque and the recurrent lesion. This sequence was compared to other sequences in the GenBank database using the Blast tool. This revealed that the 271 bp sequence amplified from both plaques had the greatest (85.9%) similarity to CPV15, and second highest similarity to CPV8 (71.9%). The novel sequence was deposited in GenBank under accession number OP645388.

根据PV感染的组织学证据，从福尔马林固定的和石蜡包埋的两种斑块标本中提取总DNA，并使用MY09/11和CP4/5共有PCR 引物从两个斑块组织中扩增PV DNA。两个引物均在两个斑块病变组织中扩增PV DNA。当对MY09/11引物扩增的DNA进行测序时，发现来自初始斑块病变和复发斑块病变的PV DNA序列相同。使用Blast工具将该序列与GenBank数据库中的其他序列进行比对。结果显示，从两个斑块中扩增出的271 bp序列与CPV15的相似性最高(85.9%)，与CPV8的相似性第二高(71.9%)。该新序列保存在GenBank中，登录号为OP645388。

 

The plaques continued to grow slowly in the four months since the last sample was taken, and, although the large central mass was around 3 cm in diameter at the time of writing, the masses did not appear to cause irritation to the dog.

在最后一次采样后的四个月里，斑块继续缓慢增长，但在撰写本文时，中心肿块的直径约为3厘米，但肿块似乎没有对犬引起刺激。

 

DISCUSSION

讨论

The cutaneous plaques in the present case contained a DNA sequence from a novel PV type. Detection of a PV sequence in a lesion does not confirm that the PV caused the lesion. However, the plaques contained histological evidence supporting a PV aetiology. Additionally, the same novel PV DNA sequence was detected in samples of both the initial and the recurrent plaques. Furthermore, no other PV type was amplified from the plaques in this case. Therefore, while additional cases are required, the putative novel PV type appears likely to be the cause of the plaques described herein. Definitive classification of a new PV type is only possible when the complete L1 sequence is known; however, the putative novel PV type amplified from the plaques appears most likely to be within the Chipapillomavirus genus with the sequence much less similar to PV types of other PV genera.

本病例的皮肤斑块含有一种新型PV型的DNA序列。在病变中检测到PV序列不能确定是PV引起病变。但是，斑块病变的组织学包含了支持PV是病因的证据。并且，在初始斑块和复发斑块的样本中均检测到相同的新型PV DNA序列。此外，在本病例中，没有从斑块中扩增出其他PV类型。因此，即使需要更多的病例来完善确诊，但拟定的新型PV类型似乎可能是本文所述斑块形成的原因。只有在完整的L1序列已知的情况下，才能明确分类新的PV类型。然而，从斑块中扩增出的拟新型PV型似乎最有可能属于乳头瘤病毒-22型，其序列与其他PV属的PV型不太相似。

 

Cutaneous viral plaques are well- recognised in dogs, the present case is unique as plaques have not previously been reported to be confined to the ear of a dog. Why the plaques developed at this location cannot be determined. However, the chipapillomaviruses are thought to cause viral plaques as a result of the immune system being unable to inhibit viral replication.In the present case, the dog had otitis which resulted in chronic discharge from the affected ear. This discharge, along with the probable self- trauma to the ear, could have damaged the skin in this area, and may therefore have reduced the normal skin defences against PV infection. This reduced skin defence then could have allowed more rapid PV replication, resulting in the development of a visible plaque. Supporting a link between otitis and plaque development in this dog was the restriction of plaques to the skin surrounding the opening of the affected ear canal. However, to the best of the authors' knowledge, areas of chronic inflammation have not previously been reported to be predisposed to plaque formation in dogs or other species.

皮肤病毒斑块在犬上很容易识别，但因为以前没有报道过局限于犬的耳部的斑块病变，所以本病例是独特的。斑块仅在这个部位形成的原因还不能确定。然而，当免疫系统无法抑制病毒复制时，乳头瘤病毒-22型被认为是形成病毒斑块的病因。在本病例中，由于犬患有耳炎，导致其耳部长期存在分泌物。这种分泌物，连同可能对耳部的自我损伤，也许损坏了该区域的皮肤，因此可能降低了正常皮肤对PV感染的防御能力。这种皮肤防御能力的降低可能会导致PV复制更快，从而导致可见斑块的形成。在这只犬耳炎和斑块发展之间的关联是，斑块局限在患病耳道开口周围的皮肤上。然而，据作者所知，以前没有报道过犬或其他物种的慢性炎症区域易形成斑块。

 

In humans, the development of cutaneous plaques is a well- recognised risk of immunosuppressive medications such as those used after organ transplantation. Likewise in dogs, there are rare reports of plaques developing in association with immunosuppressive treatments or diseases. Therefore, in addition to the local skin inflammation, the development of the plaques in the present case also could have been influenced by the chronic administration of low doses of corticosteroids. However, as many dogs are treated with corticosteroids without developing PV- induced lesions, the significance of the corticosteroids in the present case is uncertain.

在人类中，广泛认为免疫抑制药物(如器官移植后使用的药物)的使用会增加斑块形成的风险。同样在犬中，很少有与免疫抑制治疗或疾病相关的斑块形成的报道。因此，除了局部皮肤炎症外，本病例中斑块的形成发展也可能受到长期服用低剂量皮质类固醇的影响。然而，由于许多犬接受了皮质类固醇治疗，也没有出现PV引发的病变，因此在本病例中，皮质类固醇的意义尚不明确。

 

The lesions in the present case were classified as plaques rather than warts owing to the clinical presentation, histological appearance of the lesions and the presumptive aetiology.9The clinical features supporting a viral plaque rather than wart included the recurrence after excision, the slow progression of the lesions and the failure to spontaneous resolve. Additionally, the lesions were only mildly exophytic in the present case rather than being filiform or papillary as is more often seen with warts. Histologically, the lesions were formed as a result of moderate epidermal expansion. This is in contrast to the massive epidermal thickening and folding typically present in a viral wart. However, the presently described lesions did not contain pigment within the underlying dermis or the typical ‘scalloped appearance’ that usually is present within a cutaneous viral plaque. Additionally, the aural lesions contained marked PV- induced cytological changes. Such changes are to the result of PV replication in the lesion and are observed only rarely in cutaneous viral plaques in dogs, although they typically are visible in warts. Finally, a diagnosis of viral plaque was supported by the detection of a putative Chipapillomavirus PV type within the lesion. Viral plaques are associated with Chipapillomavirus PV types while warts are caused by PV types within the Lambdapapillomavirus or Taupapillomavirus genera.

由于病变的临床表现、组织学表现和拟定的病因，本病例中的病变被分类为斑块而不是疣。支持是病毒斑块而不是疣的临床特征包括其切除后复发、病变发展缓慢和未能自发消退。此外，在本病例中，病变只是轻微的外部生长，而不是常见于疣类病变的丝状或乳头状外观。组织学上，病变是由于适度的表皮扩张而形成的。这与病毒疣病变中通常出现的巨大的表皮增厚和折叠形成鲜明对比。此外，耳部病变含有明显的PV诱导的细胞学变化。这种变化是病变中PV增殖的结果，但它们在犬的皮肤病毒斑块中很少观察到，而通常在疣中可见。最后，通过在病变中检测到拟定的的乳头瘤病毒-22PV型，支持了病毒斑块的诊断。病毒斑块与乳头瘤病毒-22PV型有关，而疣则由乳头瘤病毒-11或乳头瘤病毒-19属的PV型引起。

 

Although 13 different Chipapillomavirus types have been detected in previous studies of canine viral plaques, the putative novel PV type amplified from the pinnal plaques has not been reported previously. This could suggest that the novel PV type is a very uncommon cause of viral plaques in dogs. Alternatively, some PV types have been shown to have a strong tropism for specific areas of the body. As this is the first report of plaques being confined to the pinna of a dog, it is possible that the novel PV type may preferentially infect skin in this area and so be more likely to cause plaques close to the ear than elsewhere on the body. Overall, the results from this case add to the clinical manifestations of PV- induced skin disease of dogs, as well as adding to the number of different PV types that can cause disease in dogs.

尽管在先前对犬病毒斑块的研究中已经发现了13种不同的乳头瘤病毒-22病毒类型，但这种从耳廓斑块分离的拟新型PV还没有报道过。这可能表明，这种新型的PV是造成犬病毒斑块的一类非常罕见的原因。或者，一些类型的PV对机体的特定区域有强烈的趋向性。由于这是首次报道局限于犬耳廓的斑块病变，可能是这种新型的PV型病毒优先感染这一区域的皮肤，因此更有可能导致病变斑块靠近耳部而不是身体其他部位。总体而言，本病例的探究结果增加了PV病毒诱导犬皮肤病变的临床表现，也增加了可引起犬产生疾病的PV类型的数量。

 

 

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