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  5. FP1420 : Real time PCR for HIV RNA in Aqueous Aspirate in Diagnosis and Management...

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Dr. Jayapratha, J20210, Dr. Sudharshan S

ABSTRACT

 

METHOD: Case Series

PURPOSE: Intraocular (IO) inflammation in HIV patients can be due to opportunistic infections, drugs, Immune Recovery Uveitis or a primary manifestation of HIV. We studied the role of RT-PCR for HIV RNA in the diagnosis and management of HIV induced uveitis.

RESULT: 4 patients (age: 24-43 years), with HIV disease duration-1 day to 10 years and CD4 range-164 to 412 μL with IO inflammation were included. Common inflammatory and infective causes were ruled out. Aqueous tap PCR for MTB, CMV, HSV, VZV and Toxoplasma were negative. RT- PCR for HIV RNA was positive in 3 patients-range (62 to 1, 64,773 IU/ml).All were treated with HAART. 1 patient (25%), post treatment aqueous viral load dropped by 81.91% correlating to blood values. At 3 month follow-up, complete resolution of inflammation and visual acuity improvement was noted.

CONCLUSION: RT-PCR for HIV is a reliable tool for detection and quantification of the HIV RNA in aqueous and blood and is of diagnostic value in inconclusive situations.

REAL TIME POLYMERASE CHAIN REACTION (PCR) FOR HIV RNA IN AQUEOUS ASPIRATE IN DIAGNOSIS AND MANAGEMENT OF HIV UVEITIS

Intraocular inflammation in HIV patients can be due to opportunistic infections, drugs, Immune recovery uveitis (IRU) or a primary manifestation of the HIV infection itself. We studied the role of RT-PCR for HIV RNA in the diagnosis and management of HIV induced Uveitis.

INTRODUCTION

After the introduction of Highly active Anti-Retroviral Therapy (HAART),the  incidence of Immune recovery uveitis (IRU),ocular toxic and allergic reactions have increased while the prevalence of opportunistic infections has decreased. HIV itself is a suspected cause of intraocular inflammation. Several cases are reported in the literature, where the HIV 1 viral load was increased in HIV patients presenting with anterior uveitis, in whom all the other causes of inflammation and infection were ruled out. (1,2)

We report a series of four patients in whom Real Time PCR for HIV RNA was used in the diagnosis and management of HIV induced uveitis.

Case 1

A thirty six year old male presented with history of two months of gradual diminution of vision in both eyes. On examination his best corrected visual acuity in the right eye was 6/6,N6 and in the left eye 6/18, N6. Slit lamp examination showed medium sized keratic precipitates diffusely distributed on the corneal endothelium in both eyes. The anterior chamber had 2+ cells and 2+ flare. There were no peripheral synechiae. The media was hazy due to vitritis (2+). Disc evaluation was normal. There was no vasculitis, exudates or clinically evident edema. Chest X-ray was normal .The blood sampled for evaluation of possible collagen vascular diseases, auto immune diseases and infective causes for intraocular inflammation were negative. RA factor, ANA were not detected, serum ACE was within normal limits and RPR, TPHA, ELISA for Toxoplasma, Quantiferon TB Gold test were all negative.Quantification PCR of HIV RNA by RT-PCR detected 13,404 IU/ml and 261 IU/ml of RNA in aqueous and blood sample respectively. An Anterior chamber aspirate was negative for PCR mycobacteria, CMV, HSV1&2, VZV and toxoplasma. Quantiferon TB Gold test was negative.High resolution Computed Tomography of Chest showed infiltrates in the lungs.Blood samples tested for HIV were positive. Considering a possible diagnosis of HIV induced uveitis, based on the increased viral load, both in blood and aqueous, the patient was started on topical Prednisolone acetate eye drops 3 hourly with weekly tapering, along with Homatropine 1% eye drops. Anti-tuberculosis treatment was started in consultation with a physician. Combination anti-retroviral therapy was started in consultation with an AIDS care physician after two weeks of starting the ATT. After 2 weeks of ART, the patient was started on weekly tapering schedule of systemic steroids and was closely monitored clinically. At 8 weeks, after completion of systemic steroid therapy, the patient was on still on ART, the aqueous samples were taken for microbial evaluation, which showed no HIV RNA detected.At three months follow up visit, he had best corrected visual acuity of 6/6; N-6, respectively for distance and near, in both eyes. The aqueous and vitreous were quiet and intraocular pressure were normal (12mm Hg OU).

CASE 2

A twenty one year old male presented with painless progressive diminision of vision in left eye for one year. He is a known case of HIV for the past one month, not on HAART and his CD4 count was 412 microliter. On examination, his best corrected visual acuity was 6/9,N6 in right eye and 6/12, N6 in left eye. Slit lamp examination showed 1+ cells and 1+ flare in anterior chamber with medium sized grey-white Keratic precipitates on corneal endothelium and posterior sub-capsular cataract in left eye. The media was hazy due to 1+ vitritis. There was no peripheral synechiae. After ruling out  possible causes of infection and inflammation using appropriate investigations, anterior chamber tapping was done and tested for PCR of Mycobacteria,CMV, HSV 1 & 2, VZV, and TOXOPLASMA AND HIV RNA. RT PCR for HIV RNA in aqueous aspirate detected 1, 64,773 IU/ml and blood sample tested for RT PCR for HIV RNA detected 1,50,729 IU/ml of HIV RNA. Considering a possible diagnosis of HIV induced uveitis, based on increased viral load, the patient was started on 1 %  topical Prednisolone acetate drops 3 hourly with weekly tapering along with Homatropine 1 % eyedrops. Combination anti-retroviraltherapy (Tenofovir+ lamivudine + Efavirenz) was started in consultation with AIDS care physician. After 2 weeks of HAART, the patient was started on weekly tapering schedule of systemic steroids. At 3 months follow up visit, the patient was asymptomatic with best corrected visual acuity of 6/7.5, N6 in right eye and 6/9, N6 in left eye. The aqueous and vitreous were quiet and intraocular pressure normal (10,10 mm of Hg).

CASE 3

A forty year old male presented with complaints of redness and pain in left eye for one day. His best corrected visual acuity was 6/6, N6 in both eyes. Slit lamp examination revealed old Keratic precipitates in right eye ,circum-corneal congestion, 4+ cells and hypopyon of 1 mm in left eye. The intra ocular pressure was 12 and 10 mm of Hg in right and left eyes respectively. The fundus examination of right eye was within normal limits and left eye was hazily viewed due to anterior chamber reaction. He is a known case of HIV on HAART (Darunavir, Raltegravir and Ritonavir). There was no vasculitis, exudates or clinically evident edema. Chest X-ray was normal .The blood sampled for evaluation of possible collagen vascular diseases, auto immune diseases and infective causes for intraocular inflammation were negative. RA factor, ANA were not detected, serum ACE was within normal limits and RPR, TPHA, ELISA for Toxoplasma, Quantiferon TB Gold test were all negative. The CD count was 105 cells /mm3. An Anterior chamber aspirate was negative for PCR mycobacteria, CMV, HSV1&2, VZV,toxoplasma and HIV RNA. Quantiferon TB Gold test was negative. The patient was started on 1 % topical Prednisolone acetate drops   3 hourly with weekly tapering along with Homatropine 1 % eyedrops and HAART was continued. At 2 weeks follow up, hypopyon resolved in left eye and anterior chamber reaction reduced. At three months follow up visit, he was asymptomatic with  best corrected visual acuity of 6/6, N6 in both eyes and intraocular pressure was 12 mm of Hg in both eyes.

CASE 4

A twenty nine year old haemophilia patient, who acquired HIV infection through blood transfusion presented with complaints of blurring of vision in both eyes for the past six months. His best corrected visual acuity was 6/7.5, N6 in right eye and 6/6, N6 in left eye. Slit lamp examination showed 1+ cells in both eyes. Fundus examination in both eyes were within normal limits. Chest X-ray was normal .The blood sampled for evaluation of possible collagen vascular diseases, auto immune diseases and infective causes for intraocular inflammation were negative. RA factor, ANA were not detected, serum ACE was within normal limits and RPR, TPHA, ELISA for Toxoplasma, Quantiferon TB Gold test were all negative. RT PCR for HIV RNA detected 62 IU/ml and 121 IU/ml in right and left eyes respectively. The patient was started on 1 % topical Prednisolone acetate drops 3 hourly with weekly tapering along with Homatropine 1 % eye drops and HAART was continued. He was started on weekly tapering schedule of systemic steroids. At three months follow up visit, he was asymptomatic with best corrected visual acuity of 6/6, N6 in both eyes and intraocular pressure was 12 mm of Hg in both eyes.

DISCUSSION

HIV replicates in CD4 lymphocytes. Intra –thecal persistence of HIV in spite of HAART and reduced plasma viral loads has been demonstrated (3 to 6), due to compartmentalization and replication of HIV in central nervous system. The CD4 receptors have been identified in helper T cells, activated monocytes, macrophages and glial cells (7). HIV -1 has been located in retinal vascular cells and endothelium of chorio-capillaries (8). HIV -1 antigen has been found in all layers of sensory retina (9). It has been found that HIV-1 can be cultured from all the layers of pigment epithelium of retina (10).

PCR is an adjunct in the diagnosis and treatment of uveitis. Factors like prevalence of the disease in the population, sample collection and transportation play a role in the sensitivity and specificity of a testing strategy.

Harper et al showed a 61%true positivity and 25% true negativity in demonstrating infectious agent in aqueous sample (11).Biswas et alreported 4 % prevalence of anterior uveitis in an AIDS population(12). Anterior chamber paracentesis is a relatively safe procedure which can aid in the diagnosis and management of anterior, posterior, intermediate and pan uveitis. PCR testing of aqueous aspirate has reportedly altered the treatment plan in posterior uveitis(11,13).

It has been demonstrated that HIV induced uveitis was observed in six out of fifty six HIV patients who presented with uveitis(14).HIV infection in the eye and positive HIV RNA in intraocular fluids are associated with high plasma HIV RNA loads(15 to 17).

The sporadic cases of intraocular inflammation observed in patients without opportunistic infections who exhibited high intraocular fluid: plasma HIV RNA load, the uveitis was tentatively attributed to HIV.

Currently, uveitis due to HIV is a diagnosis of exclusion and no reliable approach to diagnosis has been established. In HIV patients, where a complete history and review of systems is inconclusive in establishing the cause for intraocular inflammation, an aqueous tap for RT PCR can be helpful in diagnosis. Demonstration of a decreasing trend of the HIV load in aqueous tap may have prognostic value.

Anterior segment manifestations in  HIV infected individuals are varied. They can be vision threatening. More commonly, they can affect the quality of life which ultimately hampers the rehabilitation of the individual which is the ultimate goal of treatment. They may present with minimal inflammatory reactions and hence the clinician should be aware of the atypical presentations(18).

Most HIV patients with Uveitis are treated on the basis of clinical features. Most of the uveitis is attributed to the opportunistic infections; partly because HIV induced uveitis does not cause sudden and severe clinical signs. Presence of another undetected pathogen cannot be ruled out(14).

In contrast to IRU where the HAART increases the symptoms and flares up the inflammation, HAART brings down the inflammation in HIV induced uveitis.

Hence HIV induced uveitis should be suspected in all HIV naïve patients who have not received HAART after ruling out  opportunistic infections and other possible causes of inflammation. RT PCR for HIV RNA in those patients will not only serve as a diagnostic tool, but can be a flawless guide of treatment deciding the systemic steroid course, which is a potential cause of morbidity in all uveitis patients.

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