Avaliação de dois testes sorológicos comerciais para diagnóstico das infecções pelo FIV e pelo FeLV / Evaluation of two point-of-care tests to diagnosis of FIV and FeLV infections

FIV e FeLV são retrovírus associados principalmente com neoplasias. Dois testes rápidos são disponibilizados no Brasil para o diagnóstico dessas infecções: um kit de imunocromatografia de fluxo bidirecional (SNAP® Combo IDEXX) e um kit de imunocromatografia de fluxo lateral unidirecional (ALERE/BIONOTE Anigen Rapid). O objetivo deste estudo foi comparar o teste SNAP® com o teste ALERE. Amostras de sangue de 178 gatos foram testadas utilizando-se ambos os kits. A reação em cadeia de polimerase em tempo real (qPCR) foi empregada como método confirmatório para todos os resultados. O teste SNAP® apresentou sensibilidade e especificidade de 100% para FIV; a sensibilidade e a especificidade do teste ALERE foram de 96,15% e 98,68%, respectivamente. A sensibilidade e a especificidade para o FeLV foram de 93,02% e 96,30% para o teste SNAP® e de 90,70% e 97,78% para o teste ALERE. Ainda em relação ao FeLV, três amostras com resultado positivo na qPCR obtiveram resultado falso-negativo em ambos os testes. Não houve diferença estatisticamente significante entre os métodos. Considerando a qPCR como padrão-ouro, o teste SNAP® apresentou maior sensibilidade e especificidade para o FIV, e o teste ALERE apresentou maior especificidade para o FeLV. Os resultados mostraram uma boa correlação entre os testes.(AU)

FIV and FeLV are Retrovirus associated mainly with feline neoplasms. Two point-of-care tests are commercially available in Brazil for diagnosis of these infections: a bidirectional flow immunochromatography kit (IDEXX SNAP ® Combo) and a lateral unidirectional flow immunochromatography kit (ALERE/BIONOTE Anigen Rapid). The aim of this study was to compare SNAP ® and ALERE tests. Blood samples obtained from 178 cats were evaluated using both tests. Quantitative real-time polymerase chain reaction (qPCR) was used as confirmatory test for all samples. The sensitivity and specificity of SNAP ® test was 100% for FIV, and for ALERE test was 96.15% and 98.68%, respectively. The sensitivity and specificity for FeLV was 93.02% and 96.30% for SNAP ® test and 90.70% and 97.78% for ALERE test. Three samples with a qPCR positive result for FeLV obtained a false negative result in both SNAP ® and ALERE tests. There was no statistically significant difference between the two methods. Considering qPCR as gold standard method, the SNAP® test showed higher sensitivity and specificity for FIV, and the ALERE test presented higher specificity for FeLV. The results showed good agreement among the tests.(AU)

Merkel cell carcinoma in an immunosuppressed patient

Abstract Merkel cell carcinoma is an uncommon neuroendocrine carcinoma with a rising incidence and an aggressive behavior. It predominantly occurs in older patients, with onset occurring at a mean age of 75-80 years. Recognized risk factors are ultraviolet sunlight exposure, immunosuppression, and, more recently, Merkel cell polyomavirus. We report a case of Merkel cell carcinoma in a young HIV positive patient with Merkel Cell polyomavirus detected in the tumor.

Screening for BK virus nephropathy in kidney transplant recipients: comparison of diagnostic tests / Desempenho de métodos diagnósticos no rastreio de nefropatia pelo vírus BK em pacientes transplantados renais

Abstract Urine cytology and qPCR in blood and urine are commonly used to screen renal transplant recipients for polyomavirus-associated nephropathy (PVAN). Few studies, however, have directly compared these two diagnostic tests, in terms of their performance to predict PVAN. This was a systematic review in which adult (≥ 18 years old) renal transplant recipients were studied. A structured Pubmed search was used to identify studies comparing urine cytology and/or qPCR in urine and plasma samples for detecting PVAN with renal biopsy as the gold standard for diagnosis. From 707 potential papers, there were only twelve articles that matched the inclusion criteria and were analyzed in detail. Among 1694 renal transplant recipients that were included in the review, there were 115 (6.8%) patients with presumptive PVAN and 57 (3.4%) PVAN confirmed. In this systematic review, the qPCR in plasma had better performance for PVAN compared to urine cytopathology.

Resumo A citologia urinária e a reação da cadeia da polimerase em tempo real (qPCR) em amostras de sangue e/ou urina são comumente utilizados para rastrear nefropatia associada ao polyomavirus (PVAN), em pacientes transplantados renais. Entretanto, poucos estudos comparam diretamente esses testes diagnósticos quanto ao desempenho para predizer esta complicação. Aqui realizamos uma revisão sistemática na qual foram estudados pacientes transplantados renais adultos (≥ 18 anos). Uma pesquisa estruturada Pubmed foi utilizada para identificar estudos comparando citologia urinária e/ou qPCR em amostras de urina e plasma para detectar PVAN, utilizando a biópsia renal como padrão-ouro para o diagnóstico. Dentre os 707 artigos em potencial, apenas 12 atendiam aos critérios de inclusão e foram analisados em maior detalhe. Foram incluídos 1694 pacientes transplantados renais, entre os quais 115 (6,8%) classificados com PVAN presuntivo e 57 (3,4%) PVAN confirmado. Nessa revisão sistemática, o qPCR no plasma tive melhor desempenho para PVAN em comparação com citopatologia urinária.

Treatment of BK virus-associated hemorrhagic cystitis with low-dose intravenous cidofovir in patients undergoing allogeneic hematopoietic cell transplantation

BACKGROUND/AIMS: BK virus (BKV) has been associated with late-onset hemorrhagic cystitis (HC) in recipients of hematopoietic stem cell transplantation (HSCT). Cidofovir has been used at higher doses (3 to 5 mg/kg/wk) with probenecid prophylaxis; however, cidofovir may result in nephrotoxicity or cytopenia at high doses. METHODS: Allogeneic HSCT recipients with BKV-associated HC are treated with 1 mg/kg intravenous cidofovir weekly at our institution. A microbiological response was defined as at least a one log reduction in urinary BKV viral load, and a clinical response was defined as improvement in symptoms and stability or reduction in cystitis grade. RESULTS: Eight patients received a median of 4 weekly (range, 2 to 11) doses of cidofovir. HC occurred a median 69 days (range, 16 to 311) after allogeneic HSCT. A clinical response was detected in 7/8 patients (86%), and 4/5 (80%) had a measurable microbiological response. One patient died of uncontrolled graft-versus-host disease; therefore, we could not measure the clinical response to HC treatment. One microbiological non-responder had a stable BKV viral load with clinical improvement. Only three patients showed transient grade 2 serum creatinine toxicities, which resolved after completion of concomitant calcineurin inhibitor treatment. CONCLUSIONS: Weekly intravenous low-dose cidofovir without probenecid appears to be a safe and effective treatment option for patients with BKV-associated HC.

Risk factors in the progression of BK virus-associated nephropathy in renal transplant recipients

BACKGROUND/AIMS: BK virus-associated nephropathy (BKVAN) is an important cause of allograft dysfunction in kidney transplant recipients. It has an unfavorable clinical course, and no definite treatment guidelines have yet been established. Here, we report our center's experience with biopsy-proven BKVAN and investigate factors associated with its progression. METHODS: From January 2004 to April 2013, 25 patients with BKVAN were diagnosed by biopsy at Seoul St. Mary's Hospital. Of the 25 patients, 10 were deceaseddonor transplant recipients and 15 were living-donor transplant recipients. Three of the patients underwent retransplantation. The primary immunosuppressant used was tacrolimus in 17 patients and cyclosporine in eight patients. RESULTS: BKVAN was observed at a mean duration of 22.8 ± 29.1 months after transplantation. The mean serum creatinine level at biopsy was 2.2 ± 0.7 mg/dL. BKVAN occurred with acute rejection in eight patients (28%). Immunosuppression modification was performed in 21 patients (84%). Additionally, leflunomide and intravenous immunoglobulin were administered to 13 patients (52%) and two (8%), respectively. Allograft loss occurred in five patients (27.8%) during the follow- up period at 0.7, 17.1, 21.8, 39.8, and 41.5 months after the BKVAN diagnosis. Advanced stages of BKVAN, increased creatinine levels, and accompanying acute rejection at the time of BKVAN diagnosis increased the risk of allograft failure. CONCLUSIONS: The clinical outcomes in patients with biopsy-proven BKVAN were unfavorable in the present study, especially in patients with advanced-stage BKVAN, poor renal function, and acute allograft rejection.

JC polyomavirus infection in candidates for kidney transplantation living in the Brazilian Amazon Region

This study evaluated the relative occurrences of BK virus (BKV) and JC virus (JCV) infections in patients with chronic kidney disease (CKD). Urine samples were analysed from CKD patients and from 99 patients without CKD as a control. A total of 100 urine samples were analysed from the experimental (CKD patients) group and 99 from the control group. Following DNA extraction, polymerase chain reaction (PCR) was used to amplify a 173 bp region of the gene encoding the T antigen of the BKV and JCV. JCV and BKV infections were differentiated based on the enzymatic digestion of the amplified products using BamHI endonuclease. The results indicated that none of the patients in either group was infected with the BKV, whereas 11.1% (11/99) of the control group subjects and 4% (4/100) of the kidney patients were infected with the JCV. High levels of urea in the excreted urine, low urinary cellularity, reduced bladder washout and a delay in analysing the samples may have contributed to the low prevalence of infection. The results indicate that there is a need to increase the sensitivity of assays used to detect viruses in patients with CDK, especially given that polyomavirus infections, especially BKV, can lead to a loss of kidney function following transplantation.

Prevalence of human papillomavirus and Epstein-Barr virus DNA in penile cancer cases from Brazil

Penile cancer is a potentially mutilating disease. Although its occurrence is relatively rare worldwide, penile cancer rates can be high in developing countries. A few studies have been conducted on the involvement of human papillomavirus (HPV) in penile carcinoma, which have found HPV present in 30-70 percent of penile malignant lesions, with a higher prevalence of HPV 16 and 18. It has been assumed that cofactors, such as Epstein-Barr virus (EBV) infections, may play a role in the progression of penile neoplasia. The aim of this study was to determine HPV and EBV prevalence in 135 penile malignant lesions from Brazilian men through the use of MY09/11 polymerase chain reaction (PCR), type-specific PCR and restriction fragment length polymorphism analysis. HPV prevalence among the men tested was 60.7 percent. Of the men who tested positive, 27 presented with HPV 16 (29.7 percent), five with HPV 18 (5.5 percent), 21 with HPV 45 (23.1 percent) and nine with HPV 6 (9.9 percent). Seven mixed infections were detected (9.2 percent), while 11 cases remained untyped (13.4 percent). Regarding EBV positivity, 46.7 percent of the samples contained EBV DNA with EBV-1 as the most prevalent type (74.6 percent). More than 23 percent of the men were co-infected with both HPV and EBV, while 35 percent presented exclusively with HPV DNA and 20 percent presented only with EBV DNA. Penile carcinoma aetiology has not been fully elucidated and the role of HPV and EBV infections individually or synergistically is still controversial. Hence, more studies are needed to determine their possible role in carcinogenesis.

Polyoma BK virus: an emerging opportunistic infectious agent of the human central nervous system

BK virus, a double-stranded DNA virus, is a member of the Polyomaviridae family which is known to infect humans. Clinical evidence of disease is mostly encountered in immunosuppressed individuals such as AIDS patients or those who undergo renal or bone marrow transplantation where complications associated with BKV infection manifest commonly as a polyomavirus nephropathy or hemorrhagic cystitis, respectively. Recent evidence suggests that in addition to the JC virus (the other member of the same family known to be strongly neurotropic and responsible for the progressive multifocal leukoencephalopathy), BK virus can infect and cause clinically relevant disease in the human central nervous system. In this mini-review, an analysis of the literature is made. A special focus is given to alert clinicians to the possibility of this association during the differential diagnosis of infections of the central nervous system in the immunocompromised host.

BKV-infection in kidney graft dysfunction

INTRODUCTION: BKV nephropathy (BKN) causes kidney graft loss, whose specific diagnosis is invasive and might be predicted by the early detection of active viral infection. OBJECTIVE: Determine the BKV-infection prevalence in late kidney graft dysfunction by urinary decoy cell (DC) and viral DNA detection in urine (viruria) and blood (viremia; active infection). METHODS: Kidney recipients with >1 month follow-up and creatinine >1.5 mg/dL and/or recent increasing >20 percent (n = 120) had their urine and blood tested for BKV by semi-nested PCR, DC searching, and graft biopsy. PCR-positive patients were classified as 1+, 2+, 3+. DC, viruria and viremia prevalence, sensitivity, specificity, and likelihood ratio (LR) were determined (Table 2x2). Diagnosis efficacy of DC and viruria were compared to viremia. RESULTS: DC prevalence was 25 percent, viruria 61.7 percent, and viremia 42.5 percent. Positive and negative patients in each test had similar clinical, immunossupressive, and histopathological characteristics. There was no case of viremia with chronic allograft nephropathy and, under treatment with sirolimus, patients had a lower viruria prevalence (p = 0.043). Intense viruria was the single predictive test for active infection (3+; LR = 2.8).1,6-4,9 CONCLUSION: DC, BKV-viruria and -viremia are commun findings under late kidney graft dysfunction. Viremia could only be predicted by intense viruria. These results should be considered under the context of BKN confirmation.

Neuropatía por virus BK post trasplante renal diagnostico y seguimiento por PCR en tiempo real / BK virus nephropathy after renal transplantation: Diagnosis and prognosis by real time PCR

La nefropatía producida por el virus BK puede llevar a la pérdida del trasplante renal. El diagnóstico etiológico es importante debido a que la clínica no permite diferenciar entre nefropatía por virus BK y rechazo agudo, en donde los tratamientos de estas dos entidades son diametralmente opuestos. El desarrollo reciente de métodos moleculares muy sensibles y específicos como PCR y PCR en tiempo real para virus BK permiten un diagnóstico de certeza en forma rápida y cuantificar la carga viral presente. El diagnóstico de nefropatía por virus BK se realiza por inmunohistoquímica en una biopsia renal, pero dada la naturaleza multifocal de las lesiones, la sensibilidad no siempre es del 100%. Los nuevos métodos de PCR para detectar virus BK en sangre y orina contribuyen al diagnóstico de nefropatía de una manera más normatizada y menos invasiva. Más aún, la cuantificación del virus BK en sangre por PCR en tiempo real, ha demostrado ser útil en el diagnóstico y monitoreo de esta enfermedad. En este trabajo se presenta el caso de una paciente transplantada renal con nefropatía por virus BK y el desarrollo de un método de PCR en tiempo real para la detección de virus BK en sangre y orina. Esta nueva metodología confirmó el diagnóstico de nefropatía por virus BK lo que permitió un cambio en el esquema de inmunosupresión y la instauración de un tratamiento que pudo ser monitorizado utilizando la carga viral.

BK virus nephropathy may lead to kidney transplant failure. BK infection and acute rejection are clinically undistinguishable, therefore diagnosis of these entities is critical to establish the correct treatment. The new molecular methods using PCR and real time PCR have significantly contributed to the rapid and sensitive diagnosis of BK virus. Furthermore, viral load determination in plasma has significantly been associated with BK virus nephropathy. Definite diagnosis of nephropathy requires renal biopsy, although due to the multifocal nature of the disease sensitivity may be less than 100%. BK detection in blood and urine by PCR has contributed to the diagnosis of nephropathy in a more standardized and less invasive way. Recently, quantification of BK virus in plasma has been used for the diagnosis and monitoring of this disease. In the present study, we describe the validation of a real time PCR method for BK virus detection in plasma and urine and its application for diagnosis and monitoring in a renal transplant patient with nephropathy.

Diagnóstico y terapia del virus papiloma humano / Diagnosis and treatment of human papilloma virus

Tanto el diagnóstico como el tratamiento de la infección producida por el virus papiloma humano y el cáncer asociado a este virus, nos plantean uno de los mayores desafíos en la última década. Las principales dificultades radican en la identificación del genotipo viral, la ausencia de una terapia antiviral efectiva y las altas tasas de recurrencia y persistencia a pesar de la terapia empleada. Se presenta un resumen de la terapia disponible en la actualidad.

The identification and treatment of human papillomavirus (HPV) infections and HPV-associated neoplasm are complex. Difficulties in diagnosis and treatment of HPV-associated diseases arise from inabilities to detect HPV efficiently, the lack of specific antiviral drugs active against HPV and the high rates of recurrence and persistence of HPV infections after treatment. We present a review of therapies for HPV infections.

Importância da aplicaçäo de critérios morfológicos näo-clássicos para o diagnóstico citológico de papilomavírus humano / Application of non-classical morphological criteria in the cytological diagnosis of human papillomavirus

Existe atualmente grande preocupaçäo com a melhoria no diagnóstico citológico de HPV através da introduçäo de critérios näo-clássicos para o seu diagnóstico, tendo em vista a elevada freqüência da infecçäo viral, bem como o seu potencial carcinogênico. O presente trabalho objetivou uma releitura de 65 casos positivos para HPV por captura híbrida (cujos critérios inicialmente utilizados foram apenas os clássicos), e a introduçäo dos critérios näo-clássicos. Os critérios citológicos mais observados foram os näo-clássicos, sendo bi ou multinucleaçäo o mais freqüente, com n = 39 (60 por cento), seguido de halo perinuclear, com n = 37 (56,9 por cento) e disceratose leve e núcleo hipercromático, ambos com n = 32 (49,2 por cento). Para NIC I predominou a disceratose, com n = 20 (100 por cento); para inflamaçäo, disceratose leve, com n = 35 (87,5 por cento) e para Ascus e Agus, halo perinuclear, núcleo hipercromático, células gigantes e bi ou multinucleaçäo, todos com n = 4 (100 por cento). Todos os 20 casos de NIC I, dez dos 40 casos de inflamaçäo e todos os casos de Ascus (n = 1) e Agus (n = 3) foram considerados positivos para HPV com a utilizaçäo de critérios näo-clássicos, passando a 52,3 por cento dos casos positivos pela captura híbrida a apresentarem a infecçäo viral pela citologia, em detrimento de 23,1 por cento só com os critérios clássicos. Um número razoável de amostras diagnosticadas inicialmente como inflamatórias passaram a apresentar a infecçäo viral, podendo representar a variedade subclínica do vírus, para a qual o diagnóstico molecular apresentou-se como melhor metodologia. A utilizaçäo de critérios näo-clássicos de HPV pela citologia parece muito importante para maximizar a eficiência diagnóstica

Citología vaginal: la importancia de la zona de transformación y cómo obtener una muestra adecuada / Vaginal cytology: the importance of the transformation zone and how to obtain an adequate sample

Cytology smear is the test most frequently used to detect cancer of the uterine cervix. Much is written concerning it in its three phases, analytical, post-analytical, and preanalytical; the later implies sampling and preservation and is the purpose of this review. According to the national literature, result of the sampling is deficient in 64 of cases. Many instruments have been developed to take good samples, and statistically there is no difference between these in terms of achieving the goal of taking a good sample; despite this in Mexico bad samples persist. In this paper it is analyzed the source of these errors that are principally from ignoring the anatomy of the cervix, the handling and fixation of the sample, among others, and some observations are made that are not described in other articles in the same field. Some of the most popular instruments available in Mexico to take samples are the endocervical brush, spatula, and brush (Pappette) here described; their advantages and disadvantages are included, according to the type of cervix in which is going to be used, stressing some details concerning the transformation zone and how to select the best instrument. The results improved with these observations from 45 to 77 in taking good samples.

Diagnóstico citológico de Ascus: sua importância na conduta clínica / Cytological diagnosis of Ascus: its importance in clinical conduct

O termo "atipia de células escamosas de significado indeterminado" (Ascus) foi introduzido, em 1988, pelo Sistema Bethesda. A conduta clínica nesses casos é controversa. Outra dificuldade, é a falta de uniformização de critérios citológicos dessa anormalidade, resultando em taxas de ocorrência discrepantes. Neste trabalho, estudamos 111 casos, incluídos nesta categoria citológica, concorrendo para uma taxa de 2,4 por cento detectada num universo de 4.634 exames citológicos no período de um ano. Todas as pacientes foram avaliadas colposcopicamente, concomitantemente à colheita do material citológico cervicovaginal. Destas, 80 (72 por cento) apresentavam anormalidades ao exame colposcópico, sendo realizadas 38 biópsias dirigidas (34 por cento). A histopatologia mostrou cervicite crônica e/ou hiperplasia simples do epitélio malpighiano em 39 por cento dos exames, com os 61 por cento restantes apresentando lesões intraðepiteliais escamosas, variando de baixo grau em 20 casos (52 por cento) a alto grau em três (9 por cento). Predominaram os casos de infecção pelo HPV (39 por cento). Estas taxas comprovam a estreita relação de Ascus com lesões préðneoplásicas, inclusive aquelas de alto grau. Diante do significado clínico de Ascus estabelecido neste estudo, consideramos ser indicada biópsia dirigida em todos os casos de anormalidade colposcópica concomitante, sendo a histopatologia o meio mais fidedigno na avaliação destes casos

Analysis of cervico-vaginal (Papanicolaou) smears, in girls 18 years and under

This study was conducted retrospectively at the Queen Elizabeth Hospital and a private laboratory in Barbados to determine the types of epithelial abnormalities in cervico-vaginal Papanicolaou (Pap)-stained smears, and their clinical implications in Barbadian girls, 18 years and under, during the five-year period January 1995 to December 1999. Two hundred and sixty-five Pap smears from 236 patients were examined and the gynaecological history, initial and repeat Pap smear diagnoses, and histology reports of these patients were analyzed. Of the 236 first-visit smears, 94 (39.8) were abnormal with 36 (15.3) displaying cytologic features of squamous intra-epithelial lesions (SIL), (33 low grade and 3 high grade). A diagnosis of atypical squamous cells of undetermined significance (ASCUS) was reported in the remaining 58 (24.5) abnormal smears, of which 35 (60.3) were suspected to be related to human papillomavirus (HPV) infection. Twenty-two (23.4) of these 94 patients, who had abnormal smears of either ASCUS or low grade squamous intra-epithelial lesions (LSIL) were re-evaluated within six to twelve months of the initial abnormal Pap smear diagnosis. Eight of these 22 patients (36.4) had histological diagnosis of LSIL inclusive of cervical intra-epithelial neoplasia grade 1 (CIN 1) and condylomata. High-risk HPV DNA types were detected in two of these eight patients (25). The study confirms that sexually active teenage girls are at risk of developing SIL and high-risk HPV infection. Screening of sexually active teenaged girls by Pap smears followed by other appropriate investigative procedures is recommended.

Multiple papilloma larynx.

Multiple papilloma of larynx is caused by human papilloma virus. We treated sixteen such cases (10 males and six females) in the last 10 years. All presented with hoarseness while six presented with difficulty in respiration. Three patients needed tracheostomy, all had difficult decanulation, and one developed laryngotracheal stenosis and could not be decanulated. All were treated by surgical excision; ten had recurrence. Four patients were treated with post operative Acyclovir with no recurrence in three cases.