Enhanced U.S. Army HIV Diagnostic Algorithm Used to Diagnose Acute HIV Infection in a Deployed Soldier

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1 MILITARY MEDICINE, 177, 5:609, 2012 Enhanced U.S. Army HIV Diagnostic Algorithm Used to Diagnose Acute HIV Infection in a Deployed Soldier Shilpa Hakre, DrPH, MPH*; LTC Robert M. Paris, MC USA ; MAJ Julie E. Brian, AN USAR ; CDR Jennifer Malia, USPHS ; Eric E. Sanders-Buell, BS ; Sodsai Tovanabutra, PhD ; LTC Bryan C. Sleigh, MC USA ; COL James E. Cook, MC USA**; COL Nelson L. Michael, MC USA ; Paul T. Scott, MD, MPH ; COL Dan R. Deuter, MC USA ; LTC Steven B. Cersovsky, MC USA ; Sheila A. Peel, MSPH, PhD ABSTRACT Antibody screening alone may fail to detect human immunodeficiency virus (HIV) in recently infected individuals. By U.S. Army regulation, HIV-infected soldiers are not permitted to deploy to areas of conflict, including Iraq and Afghanistan. We report here the first case of acute HIV infection (AHI) in a soldier in a combat area of operation detected by an enhanced U.S. Army HIV testing algorithm and discuss features of the tests which aided in clinical diagnosis. We tested the sample from the AHI case with a third generation HIV-1/HIV-2 plus O enzyme immunoassay, HIV-1 Western Blot, and a qualitative HIV-1 ribonucleic acid molecular diagnostic assay. Risk factors for HIV acquisition were elicited in an epidemiologic interview. Evaluation of the blood sample for AHI indicated an inconclusive serologic profile and a reactive HIV-1 ribonucleic acid result. The main risk factor for acquisition reported was unprotected sexual intercourse with casual strangers in the U.S. while on leave during deployment. The clinical diagnosis of AHI in a combat area of operation is important. Diagnosis of HIV is key to preventing adverse effects to the infected soldier from deployment stressors of deployment and further transmission via parenteral or sexual exposures. INTRODUCTION Acute human immunodeficiency virus (HIV) infection (AHI), known as primary HIV infection or acute retroviral syndrome, is the period between HIV acquisition and antibody detection and can be up to 22 days in duration depending on the serologic test used. 1 Intense replication of HIV and the host *United States Military HIV Research Program, Henry M. Jackson Foundation for the Advancement of Military Medicine, 6720-A Rockledge Drive, Suite 400, Bethesda, MD Department of Medicine, Infectious Diseases Service Clinic, Walter Reed National Military Medical Center-Bethesda, 8901 Wisconsin Avenue, Bethesda, MD th Battle Command Training Division, Building 5520 Nashville Street, Fort Dix, NJ United States Military HIV Research Program, Walter Reed Army Institute of Research, 13 Taft Court, Suite 100, Rockville, MD kunited States Military HIV Research Program, Henry M. Jackson Foundation for the Advancement of Military Medicine, 503 Robert Grant Avenue, Rockville, MD HQ Army Medical Directorate, FASC, Rm 129, Royal Military Academy at Sandhurst, United Kingdom **Preventive Medicine Department, Madigan Army Medical Center, Building 9920A, Ramp 3, Tacoma, WA United States Military HIV Research Program, Walter Reed Army Institute of Research, 503 Robert Grant Avenue, Silver Spring, MD United States Military HIV Research Program, Walter Reed Army Institute of Research, 6720-A Rockledge Drive, Suite 400, Bethesda, MD Medical Command, Building 2748, 3151 Scott Road, Suite 1334, Fort Sam Houston, TX kkunited States Army Public Health Command Epidemiology & Disease Surveillance, ATTN: MCHB-TS-D, 5158 Blackhawk Road, Aberdeen Proving Ground, MD The views expressed are those of the authors and should not be construed to represent the positions of the U.S. Department of Defense, the U.S. Government, or any of its agencies. immune response during this period result in high levels of detectable biomarkers (ribonucleic acid [RNA] and p24 antigen) in blood and semen 2 and in symptoms such as fever, fatigue, rash, headache, and sore throat in up to 89% of patients. 3 Diagnosis of AHI in patients seeking medical care for symptoms can result in improved outcomes for HIVinfected individuals 4 and provide opportunities for public health authorities to prevent further transmission. 5 However, the U.S. Centers for Disease Control and Prevention estimated that at the end of 2006, 21% of the U.S. population was unaware of their infection status. 6 Moreover, approximately 8.6 to 11.4% of new HIV infections in the United States may be acquired from individuals with AHI. 7,8 Detection and diagnosis are key first steps to HIV care and prevention. U.S. Military personnel on active duty are mandated to undergo biennial HIV antibody screening. 9 In 1986, HIV surveillance policies were instituted by the U.S. Military to ensure the safety of combat blood supply during urgent blood collections within combat casualty resuscitation settings. Currently, Military personnel must screen negative for HIV within 120 days of deployment to U.S. Central Command Areas of Operation (CENTCOM AOR), inclusive of Iraq and Afghanistan. 10 HIV-infected personnel are not permitted to deploy. An estimated 1.64 million Military personnel have deployed to combat operations in Iraq and Afghanistan. 11 The capability to detect and diagnose HIV at a single time point (i.e., from a single sample) is critical for a highly mobile population such as the U.S. Military with unique operational requirements. The standard HIV diagnostic algorithm, repeatedly reactive enzyme immunoassay (EIA) followed by confirmation with HIV-1 Western Blot (WB), used within the United States since 1989, is seriously limited MILITARY MEDICINE, Vol. 177, May

2 by the diagnostic window period of the gold standard WB which is less sensitive than current third and fourth generation EIAs, and fails to detect AHI and late stage infection. 12 To address these limitations, earlier serological detection methods incorporating p24 antigen have been employed, as well as HIV-1 RNA for AHI diagnosis, a model-based score for targeted screening, and pooled HIV-1 RNA nucleic acid test (NAT) of antibody-negative samples. 5,13 On December 1, 2009, the U.S. Army HIV Diagnostic Reference Laboratory (HDRL, Rockville, Maryland) implemented an enhanced screening algorithm to address limitations of the standard serological HIV algorithm by incorporating a highly sensitive qualitative HIV-1 RNA diagnostic assay in conjunction with serological testing. Through June 30, 2011, 1,149,773 soldiers in the U.S. Army Active, National Guard, and Reserve components have been tested using this algorithm. 14 We report here the first case of AHI detected in a combat theater of operation and discuss features of the tests introduced in the U.S. Army AHI testing algorithm that aided in clinical diagnosis. The diagnosis of the case was in support of the deployed U.S. Military Command staff in theater and was not considered research by the Walter Reed Army Institute of Research. CASE REPORT In early 2010, a 46-year-old senior enlisted white male soldier presented to a combat support hospital (CSH) in Afghanistan with chief complaints of headache, rectal pain, itching, and discomfort and requested an HIV test. Upon further questioning, he reported that 18 days earlier he had participated in high-risk behavior during a 3-week Rest and Recuperation leave (R&R) in Florida. He reported having fever, sore throat, malaise, and skin rash while returning from R&R, and had been treated en route for presumed streptococcal pharyngitits. His last seronegative HIV test was 7 months before this visit. Physical examination was unremarkable. Initial laboratory work-up at the CSH, which did not include an HIV rapid test, demonstrated a positive rapid plasma reagin for syphilis, which was confirmed by fluorescent treponemal antibody. A serum sample, drawn the same day as the medical visit to the CSH, was shipped to HDRL for HIV testing. Time from acquisition of the specimen to report of results was 27 days. Evaluation of the sample by the Army algorithm (Fig. 1) confirmed patient and clinical suspicions of AHI. Index specimen results summary included a repeat reactive EIA (Genetic Systems HIV-1/HIV-2 Plus O EIA, BioRad Laboratories, Redmond, Washington), faint FIGURE 1. Enhanced U.S. Army diagnostic algorithm for diagnosis of HIV. 610 MILITARY MEDICINE, Vol. 177, May 2012

3 antigen reactivity at p24 and gp160 on HIV-1 WB (Genetic Systems HIV-1 Western Blot, BioRad Laboratories), reactive Aptima HIV-1 RNA Qualitative Assay Signal to cutoff (S/CO = 23; >1.0 = reactive) (Gen-Probe, San Diego, California), and a nonreactive MultiSpot HIV-1/HIV-2 Rapid Test (BioRad Laboratories) (see sample number 1, Table I). Partial length sequencing of the reverse transcriptase, protease, and envelope regions of the virus from the second sample indicated HIV-1 subtype B infection. The soldier was treated for syphilis and evacuated to Landstuhl Regional Medical Center (Landstuhl, Germany) for follow-up. The second independent specimen collected at Landstuhl and referred to HDRL for testing was EIA repeat reactive, HIV-1 WB positive, and Multispot rapid test reactive. The HIV-1 viral load was 360,088 copies/ml (see sample number 2, Table I). He was referred to a Military Medical Treatment Facility in the United States for Infectious Diseases specialty evaluation and management, where the third independent specimen was collected in accordance with Army Regulation TABLE I. Laboratory Tests Performed for Diagnosis and Care of HIV in a Soldier Deployed to Afghanistan, 2010 Sample Number HIV Test 1 a 2 b 3 c 4 d Genetic Systems HIV-1/HIV-2 Plus O TNP EIA, Initial EIA, Repeat EIA, Repeat EIA Final Interpretation e Reactive Reactive Reactive WB, Initial TNP p p p p gp p p p gp gp WB Final Interpretation Indeterminate Positive Positive BioRad MultiSpot Nonreactive Reactive TNP TNP Aptima HIV-1 RNA Reactive TNP TNP TNP Qualitative Assay Roche Cobas Ampliprep/ Cobas TaqMan HIV-1 Test (copies/ml) f TNP 360,088 TNP 431 TNP, test not performed. a Collected at an initial visit to a CSH in Afghanistan for flu-like illness. b Collected at a Military Treatment Facility in Landstuhl, Germany, between initial visit and 34 days after sample 1. c Collected at a Military Treatment Facility in Landstuhl, Germany, 34 days after sample 1. d Collected at a Military Treatment Facility in the United States, 68 days after sample 1; Genotype resistance testing was not performed because of insufficient virus in sample. e Signal to cutoff ratio ³ = Reactive. f Linear range of quantification = 48 to 10,000,000 copies/ml. for diagnosis of HIV infection by two independent HIV-1 WB positive specimens (see sample number 3, Table I). A fourth specimen was acquired for HIV-1 resistance genotype that was not performed as HIV-1 viral load, 431 copies/ml, was below the assay requirement (>1,000 copies/ml). An epidemiologic public health interview conducted at initial notification of the positive HIV test result by a physician and public health nurse, revealed a history of: (1) unprotected sexual intercourse with four different female casual partners during his R&R; (2) inconsistent use of a condom with two of the casual partners; (3) consumption of more than 10 alcoholic drinks a day while on R&R; (4) consumption of alcoholic drinks before sexual intercourse; (5) involvement in casual relationships since his divorce 9 years ago; and (6) reported medical visit for syphilis 3 years ago and gonorrhea 25 years ago. He denied (1) having donated blood during his current deployment and (2) drinking alcoholic beverages or engaging in sexual intercourse while deployed. He reported having a prior deployment to Iraq. DISCUSSION The successful identification of AHI in a deployed soldier can be attributed to several factors: (1) the enhanced U.S. Army testing algorithm enabled successful diagnosis of AHI from a single specimen drawn at the initial visit for medical care in an austere setting; (2) the soldier s awareness of his risk for HIV from sexual activities during R&R, trust in, and access to, the Military Health System prompted him to seek care for his symptoms and request an HIV test; (3) health care providers were knowledgeable about resources for HIV testing in a combat region and protecting the soldier s confidentiality; (4) sufficient infrastructure at the CSH to ensure immediate acquisition, transport, and maintenance of specimen integrity from Afghanistan to the U.S.-based laboratory. This case identified areas of improvement for turnaround time from sample acquisition to report of result from CENTCOM AOR from 27 days to less than 2 weeks. Before December 2009, a soldier with an indeterminate antibody response would have been retested within 1 to 3 months. The enhanced U.S. Army HIV diagnostic algorithm included transition from a second generation whole viral lysate/recombinant EIA, Genetic Systems rlav EIA (BioRad Laboratories) to a third generation recombinant EIA, Genetic Systems HIV-1/HIV-2 Plus O EIA that detects both IgM and IgG HIV antibody (BioRad Laboratories), thus reducing the serological diagnostic window period from to days postinfection. 15 In this case, the index AHI specimen was strongly EIA repeat reactive (S/CO = >12.0; Table I) with an indeterminate, albeit suspicious, HIV-1 WB that before December 2009 would have been reported as HIV infection status Indeterminate. The Multispot rapid test result for this specimen was nonreactive, which is not surprising given that the diagnostic window period for this IgG antibody test may be as long as 4 to 6 weeks. A second MILITARY MEDICINE, Vol. 177, May

4 specimen, acquired 34 days later and approximately 52 days post a reported high-risk exposure, was reactive; thus providing a cautionary note for those utilizing HIV rapid tests for clinical indication and suspicion of AHI. This algorithm also incorporates the Aptima HIV-1 Qualitative RNA Assay, a 2006 FDA-licensed assay for diagnosis of acute and primary HIV-1 infection in the absence of HIV-1 antibody, which enables detection of 10 copies/ml of RNA (sensitivity 79%; S. Peel, unpublished data) within a few days of infection. 16 In this case, index specimen results of repeat reactive EIA and reactive qualitative RNA test led to early diagnosis of AHI, which were confirmed by the second independent specimen with a positive HIV-1 WB, reactive rapid test, and HIV-1 viral load of 360,088 copies/ml. Together, the alterations to the diagnostic algorithm ensure resolution of an indeterminate infection status with a single sample. Predeployment and R&R periods are likely times of risk for HIV acquisition. Forty-two percent of incident HIV cases detected among soldiers returning from deployment to Iraq and Afghanistan were acquired in the predeployment period and 27% in the R&R period. 17 Seronegative samples of 7 of the 20 (35%) predeployment-acquired HIV cases tested HIV RNA NAT positive. Timely identification of incident HIV cases in the predeployment period translates into preventing infected personnel from getting potentially harmful live vaccines and from exposure to the stressors of a combat environment, preventing secondary transmission through parenteral exposure in mass casualty settings or through sexual transmission, and preventing significant psychological distress for the service member, who otherwise has to be immediately evacuated following HIV notification. In addition, the identification of AHI cases prevents potential disruptions of military operations. AHI diagnosis is often missed in patients presenting for medical care. In a prospective study of AHI patients enrolled from primary and urgent care clinics, and emergency departments, only 17% of patients who sought medical care for AHI were diagnosed with HIV at their first visit. 18 Health care providers in theater should consider HIV in their differential diagnosis among Military personnel presenting with flu-like symptoms and having risk factors for HIV. HIV risk assessment should include a thorough evaluation of history of recent sexual, drug, or alcohol use risk behaviors, a history of sexually transmitted infections or mental disorders, and other risk factors for HIV. 19 CONCLUSION This report stresses the importance of the convergence of many factors for the detection of AHI. A combination of patient risk awareness, epidemiological data, provider knowledge, health care resources, and use of a highly sensitive NAT detection method enabled timely medical management of an individual in a resource-limited setting, prevented further virus transmission via blood transfusion, as well as parenteral or sexual exposures. ACKNOWLEDGMENT The Defense Health Program 8 funds for clinical diagnostics. REFERENCES 1. Fiebig EW, Wright DJ, Rawal BD, et al: Dynamics of HIV viremia and antibody seroconversion in plasma donors: implications for diagnosis and staging of primary HIV infection. AIDS 2003; 17(13): Pilcher CD, Joaki G, Hoffman IF, et al: Amplified transmission of HIV-1: comparison of HIV-1 concentrations in semen and blood during acute and chronic infection. AIDS 2007; 21(13): Schacker T, Collier AC, Hughes J, Shea T, Corey L: Clinical and epidemiologic features of primary HIV infection. 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