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JAIDS Journal of Acquired Immune Deficiency Syndromes Publish Ahead of Print

DOI: 10.1097/QAI.0000000000001003

Performance Evaluation of the new HIV-1 quantification assay, Xpert HIV-1 Viral Load,
on a wide panel of HIV-1 variants

Marie Gueudin1,2$, Adeline Baron1$, Elodie Alessandri-Gradt1,2, Vronique Leme1, Thomas

Mourez1,2, Manuel Etienne2,3, Jean Christophe Plantier1,2*

$ : equal contribution

1- Laboratoire de Virologie Associ au Centre National de Rfrence du VIH, Hpital Charles
Nicolle, CHU de Rouen, Rouen, France;
2- GRAM, Equipe dAccueil 2656, Facult de Mdecine-Pharmacie, Institut de Recherche et
dInnovation en Biomdecine, Universit de Rouen, Rouen, France;
3- COREVIH Haute-Normandie, Hpital Charles Nicolle, CHU de Rouen, Rouen, France

*Corresponding author:
Correspondence to Pr JC PLANTIER
Laboratoire de Virologie, Institut de Biologie Clinique, Hpital Charles Nicolle, CHU de
Rouen, 1 rue de Germont, 76031 Rouen, France.

Tel: +33 2 32 88 14 62; fax: +33 2 32 88 04 30;

e-mail: jc.plantier@chu-rouen.fr

Running title: HIV-1 VL Xpert evaluation

Conflicts of Interest/Sources of Funding: The authors have no conflicts of interest to disclose.


The work was supported by Cepheid, the Institut de Veille Sanitaire (InVS) and the CHU de

Presented in part as a poster presentation at the 18th Annual Meeting of European Society for
Clinical Virology, 9th12th September 2015, Edinburg, Scotland.

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4 Objective

5 To evaluate the quantification performance of the new Cepheid GeneXpert HIV-1 Viral Load

6 assay, on a wide panel of HIV-1 variants.

7 Methods

8 Clinical performance was evaluated relative to the Abbott RealTime HIV-1 assay on 285 HIV-1

9 seropositive samples selected to cover the assays quantification range (40 cp/mL to 10,000,000

10 copies/mL), and included RNA undetectable or detected seropositive samples. The panel

11 comprised 120 subtype B, 150 non-B and 15 non-typable clinical samples; serial dilutions of 18

12 viral supernatants representative of the divergent viruses of HIV-1 groups N, O and P were also
13 tested.

14 Results

15 Based on samples selected according to the Abbott assay viral loads (VL), the Cepheid assay

16 detected or quantified 222/285 (78%) samples and the Abbott assay 240/285 (84%). Xpert

17 yielded VLs for 162 (76%) of the 213 quantifiable samples with Abbott. This difference

18 corresponded to 51 samples with VL >40 cp/mL by the Abbott assay (all below 200 cp/mL) but

19 detected (n=40) or undetectable (n=11) by the Cepheid assay. VL of samples quantifiable by both

20 assays (n=162) showed very strong correlation, with a Spearmans correlation coefficient of

21 0.985 and a Bland-Altmans mean of differences of -0.01. Performance for quantification of the

22 non-M samples showed very good correlation, with significantly higher values with Cepheid for

23 the group N and 2 group O samples.



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26 Conclusion

27 Our study showed that the Xpert HIV-1 VL assay offered very good performance for detection

28 and quantification of the current HIV-1 genetic diversity; differences reported at the threshold

29 could be an issue and requires further evaluations. The practicability of this new assay makes it

30 suitable for low income countries, where it could facilitate and improve follow-up of patients, as

31 well as for high income regions.

Key words: HIV viral load, GeneXpert, genetic diversity


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34 Plasma viral load (VL) is a key element of the virological follow-up for patients infected with

35 HIV to measure the intensity of viral replication in untreated patients, as well as for assessing

36 virological response to antiretroviral drugs. Assessment of HIV-1 VL has been performed for

37 many years by different techniques, initially the measurement of p24 antigen, and currently using

38 the measurement of the RT enzymatic activity [1] or more largely by molecular techniques such

39 as nucleic acid sequence based amplification (NASBA), transcription mediated amplification

40 (TMA) or real-time reverse transcriptase polymerase chain reaction (RT-PCR) [2-6].

41 These different technologies use relatively impressive high capacity platforms, combining a
42 separate system of extraction and amplification of the viral genome. Automation improved the

43 reproducibility of results, facilitated technical work components, avoided contamination and

44 allowed savings due to processing of batched sets of samples. These platforms are advantageous

45 for large laboratories but may be inadequate or difficult to implement in smaller laboratories or in

46 field conditions in low income countries, due to required technical expertise, cost, and

47 maintenance of the instruments. In addition, traditional batched samples systems are not able to

48 perform HIV VL in real time or for emergency cases.

49 The GeneXpert (Cepheid, Sunnyvale, CA) technology is based on cartridges used in individual

50 modules that perform the extraction and quantitative RT-PCR in a unique instrument system [7].

51 This technology has been developed for different markers such as mycobacterium tuberculosis,

52 hepatitis C virus (HCV), methicillin-resistant Staphylococcus aureus (MRSA), for

53 oncohaematology and others applications. A new assay, Xpert HIV-1 Viral Load (Xpert HIV-1

54 VL), has been developed for use on the Cepheid GeneXpert Instrument System. Due to its

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55 simplicity and ease of use, this new HIV-1 VL assay could be an alternative to the current large

56 analytical platforms, for laboratories that do not have sufficient numbers of samples to perform

57 VL testing on large instruments in a cost-effective manner. It also could be interesting for larger

58 laboratories wishing to conduct their high volume viral load tests in real time, and for emergency

59 cases if necessary.

60 To be successful, a new test for quantifying HIV-1 must have analytical performance and a

61 threshold of sensitivity comparable to what is currently available (20 to 40 copies/ml plasma

62 depending on the assay). Due to the intrinsic genetic properties of HIV-1, an assay must also be

63 able to detect and quantify the wide genetic diversity of the pandemic HIV-1 group M viruses.

64 Moreover, quantifying divergent viruses of groups N, O and P, as is the case with the Abbott

65 RealTime HIV-1 and Roche CAP/CTM Cobas Taqman HIV-1 assays [8], would be an additional
66 desired feature, and would be essential for the effective monitoring of the patients infected with

67 these more rare groups of HIV-1.

68 The aim of this study was to conduct a performance evaluation of the Xpert HIV-1 VL assay

69 relative to the Abbott RealTime HIV-1 assay on a wide panel of HIV-1 variants representative of

70 the current genetic diversity of HIV-1, as well as its practicability.


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72 Cepheid Xpert assay

73 The Xpert HIV-1 VL assay is performed on the Cepheid GeneXpert instrument systems. These

74 systems automate and integrate sample preparation, nucleic acid amplification, and detection of

75 the target sequence (3-LTR region) in samples using real-time reverse transcription PCR (RT-

76 PCR). The systems require the use of single-use disposable GeneXpert cartridges that include

77 reagents as well as High (IQSH) and Low (IQSL) internal controls and a Probe Check Control

78 (PCC). The IQSH and IQSL are present to control for adequate processing of the target sample

79 and to monitor the presence of inhibitors in the RT and PCR reactions as well as for

80 quantification of HIV-1. The PCC verifies reagent rehydration, PCR tube filling in the cartridge,
81 probe integrity, and dye stability. External controls used during the study consisted of one HIV-1

82 negative, one low HIV-1 positive containing the 8E5 LAV strain in EDTA based normal human

83 plasma matrix (expected range 2.95-4.15 Log copies/mL), and one high HIV-1 positive control

84 (expected range 5.48-6.44 Log copies/mL). The negative control was made of HIV-1 negative

85 human plasma matrix. Controls were tested with the Xpert HIV-1 VL on each day that specimens

86 were tested prior to analyzing any study specimens. The run time for one sample is 90 minutes.

87 The input volume of plasma (ACD-A or EDTA) is 1 mL. The range of quantification in human

88 plasma is from 40 to 10,000,000 copies/mL (1.6 Log cp/mL to 7 Log cp/mL), and the limit of

89 detection (LOD) is determined to be 40 cp/mL for HIV-1 subtype B in EDTA and ACD plasma.

90 Results of INVALID (e.g., failure of IQSH and/or IQSL), ERROR (test failure due to Probe

91 Check: FAIL*; all or one of the probe check results fail or insufficient volume) or NO RESULT

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92 (insufficient data collected, e.g. a test in progress was stopped) are defined as indeterminate

93 results.

94 Abbott Assay

95 For comparison, all specimens were tested using the Abbott RealTime HIV-1 Assay, as

96 recommended by the manufacturer. This technique is based on an automated-system of extraction

97 of the nucleic acid (m2000 SP) and an amplification system (m2000RT) using real-time reverse

98 transcription PCR (RT-PCR); the detection is performed in the Integrase region and the linear

99 range is 40 cp/mL to 10,000,000 cp/mL (1.6 Log to 7 Log) and the LOD is 40 copies/mL with the

100 1.0 mL sample volume procedure.

101 Samples for clinical performance

102 The specificity was evaluated by testing 20 plasma (EDTA) specimens from HIV-1 negative

103 blood donors.


104 The clinical performance was assessed on 295 HIV-1 seropositive samples, collected during

105 routine viral load measurements for 285 patients managed at the Charles Nicolle Hospital,

106 Rouen, France, and requiring no additional blood samples. They were obtained from 224 subjects

107 on antiretroviral therapy (ART) and 59 untreated patients. No ART information was available for

108 two patients. The samples were initially quantified with the Abbott assay used routinely in the

109 laboratory, and selected to cover the quantification range (1.6 Log n=71 2 Log; 2 Log < n=

110 50 3 Log; 3 Log < n= 32 4 Log; 4 Log < n= 33 5 Log; n = 32 5 Log) and included RNA

111 undetectable (n= 47) or detected (n= 30) samples; 176 plasma (EDTA) were selected from

112 registered collection of samples frozen and stored at 80C and 119 plasma samples were

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113 selected prospectively and tested fresh. Fresh samples were stored at 2-8C and tested

114 simultaneously by both techniques within 5 days of collection and separation, and frozen samples

115 were tested simultaneously within the same freeze/thaw cycle.

116 The genetic diversity of the panel was determined by grouping and subtyping using molecular

117 characterization, i.e. by sequencing the Pol region (Protease and Reverse Transcriptase) on an

118 automated capillary CEQ 8000 DNA sequencer (Beckman Fullerton, CA), followed by

119 phylogenetic analyses as previously described [9]. Their genetic distribution was as follows:

120 subtypes A=15, B=120, C=9, D=6, F=6, G=6, H=6, J=1, K=1, CRF02_AG=43, other CRFs=36,

121 Unique Recombinant Forms (URFs)=21, non-typable (NT)=15 (these latter corresponded to

122 samples without genetic material, i.e. undetectable or very low viral load making the molecular
123 analysis impossible). In addition, 18 viral supernatants from cell cultures representative of non-

124 pandemic HIV-1 non-M groups, i.e. group O (n=15), group N (n=2) and group P (n=1) were

125 tested; they were obtained from strains characterized, isolated and propagated in the laboratory,

126 from samples genetically characterized as HIV-1 non-M groups. The supernatants were diluted

127 with human plasma HIV negative, to obtain reference viral loads using the Abbott assay, between

128 3 to 4 Log cp/mL, 4 to 5 Log cp/mL, and 5 to 6 Log cp/mL.



130 Statistical Analysis

131 Samples with HIV-1 RNA quantitative results within the linear range of both the Xpert HIV-1

132 VL and the Abbott RealTime HIV-1 Assay were compared using MedCalc Statistical Software

133 version 15.11.4 (MedCalc Software bvba, Ostend, Belgium; https://www.medcalc.org; 2015). A

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134 Passing-Bablok regression was calculated and a Bland-Altman graph was generated. A maximum

135 difference less than 0.5 Log between the two assays was considered as acceptable.



138 Specificity

139 None of the 20 negative samples were found positive thus the specificity of Xpert HIV-1 VL was

140 100% (95% CI: 83.9-100.0).


142 Clinical concordance

143 Xpert HIV-1 VL tests were successful on the first attempt for 96.6% (285/295) of the samples.
144 The 10 indeterminate cases reported included 3 ERROR, 6 INVALID, and 1 NO RESULT;

145 corresponding VL obtained with the Abbott assay was undetectable (n=2), detected at < 40

146 copies/mL but not quantifiable (n=3), and from 1.7 to 5.3 Log cp/mL (n=5). None of these

147 indeterminate cases was retested due to insufficient sample volume.


148 The results of the 285 specimens with valid results by both VL assays are presented in table 1.

149 They are classified as follows: HIV-1 undetectable; HIV-1 detected at < 40 copies/mL but not

150 quantifiable, and HIV quantifiable at 40 copies/mL. In both techniques, the HIV-1 VL was

151 undetectable for 38 samples, detected for 13 plasmas and quantifiable for 162 plasmas, leading to

152 overall concordance of 74.7% (213/285).

153 Taking into account that samples were initially selected on the Abbott VL, the Cepheid assay

154 detected or quantified HIV RNA in 222/285 (78%) samples, and the Abbott assay detected or

155 quantified HIV RNA in 240/285 (84%) samples. Xpert yielded VLs for 162 (76%) of the 213

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156 quantifiable samples selected with the Abbott assay; this difference corresponded to 51 samples

157 that were quantifiable with the Abbott assay but only detected or undetectable with the Cepheid

158 assay.

159 Regarding the genetic diversity of the 222 samples detected or quantified by Cepheid, 84 (38%),

160 135 (61%) and 3 (1%) were subtype B, non-B and NT respectively; among the 240 samples

161 detected or quantified by Abbott, 94 (39%), 139 (58%), and 7 (3%) were of subtype B, non-B

162 and NT respectively. Regarding the 162 samples quantified by Cepheid, 58 (36%) and 104 (64%)

163 were subtype B and non-B respectively; among the 213 samples quantified by Abbott, 84 (39%),

164 124 (58%), and 5 (3%) were subtype B, non-B and NT respectively. No difference was thus

165 observed in terms of distribution of subtype B vs subtype non-B clinical samples among data
166 obtained by both techniques.

167 Among the 51 samples not quantifiable with the Cepheid assay, forty were detected with Cepheid

168 (i.e. detected at < 40 copies/mL but not quantifiable). These samples had a low VL with the

169 Abbott assay, with a range of 40 cp/mL (1.6 Log) to 170 cp/mL (2.23 Log), a median of 56

170 cp/mL (1.75), and an average of 61 cp/mL (1.76 Log). There were 11 samples quantifiable with

171 the Abbott assay that were undetectable with Cepheid Xpert HIV-1 VL. These samples also had a

172 low VL with a range of 41 cp/mL (1.61 Log) to 77 cp/mL (1.89 Log), a median of 55 cp/mL

173 (1.74 Log) and an average of 54 cp/mL (1.73 Log). Regarding the genetic diversity of these 51

174 different samples, 26 were subtype B, 10 were CRF02_AG, 5 were URFs, 5 were NT, and there

175 was 1 for D, F1, CRF01, CRF06, CRF11, each.

176 There were 14 samples detected with Abbott that were undetectable with the Cepheid kit. Of

177 these 7 were subtype B, 2 were CRF02, 2 were NT and there was 1 for A, CRF06, CRF11 each.


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178 Conversely there were 7 samples detected with Cepheid that were undetectable with Abbott. Of

179 these samples 3 were subtype B, 2 was subtype A, 1 was CRF02 and 1 was an URF.


181 Quantitative analysis was performed for 162 clinical samples with quantifiable results by both

182 systems (104 subtype non-B and 58 B). These samples had VLs ranging from 40 cp/mL (1.6

183 Log) to 6,800,000 cp/mL (6.83 Log). Correlation between the Cepheid Xpert HIV-1 VL and the

184 Abbott RealTime HIV-1 Assay was performed using a Passing-Bablok regression on the 162

185 samples. Figure 1 shows very good correlation between both assays with a slope for the

186 regression line of 1.0355 [1.0114 to 1.0597] and an intercept of -0.1184 [-0.2151 to -0.03756],

187 showing a slightly higher quantification in the lowest VL for Abbott and in the highest VL for
188 Cepheid, with a close quantification among the middle ranges of VL.

189 Using a Bland-Altman analysis (Figure 2), we observed an extremely close quantification of all

190 samples, since the mean of differences was -0.01. Values were close along the VL level, with

191 slightly higher values for the Abbott assay in the lowest levels (between 1.6 to 2.5 Log, Figures 1

192 and 2). No dispersion or heterogeneous distribution was observed, with SD of +0.4 and -0.4,

193 values less than 0.5 considered as the maximum difference between two techniques. Only two of

194 the 162 (1.2%) samples had a difference of quantification of more than 0.5 Log. Among them 1

195 (CRF02_AG) had a difference of -0.53 (in the 2-2.5 Log range) in favor of Abbott. The other

196 sample (subtype B) had a difference of + 0.78 (in the 4-4.5 Log range) in favor of Cepheid.

197 Performance for detection and quantification of the HIV-1 groups non-M samples (N, O, and P)

198 was analyzed for the 3 dilutions of the 18 culture supernatants (Figure 3). Results showed a

199 median with Cepheid and Abbott of 2.80 and 2.74 for the 2-3 Log dilution, of 3.80 and 3.69 for


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200 the 3-4 Log dilution and of 4.81 and 4.74 for the 4-5 Log dilution, respectively. Globally

201 these values were linear along the dilutions for the same assay and very close between the assays

202 (slightly higher for Xpert). For 4 supernatants, we observed significantly higher values with

203 Cepheid than with Abbott. Of those found, 2 were group N samples (difference of +0.87 Log and

204 +0.74 Log each at the 3-4 Log dilutions; similar differences were found for the 2 other dilutions)

205 and 2 of the 15 group O samples (difference of +0.87 Log and +1.83 Log at the 3-4 Log

206 dilutions; similar differences were found for the 2 other dilutions). No difference (regardless of

207 dilutions) was observed for the unique group P sample.


209 The aim of this study was to evaluate the performance of the new HIV-1 viral load test

210 from Cepheid. Given the important genetic diversity of HIV-1, we wanted to evaluate

211 performance on a large panel of samples representative of the current HIV-1 diversity. We thus

212 tested 295 clinical samples including 120 subtype B and 150 subtype non-B of HIV-1 group M,

213 as well as 18 culture supernatants derived from the diversity of the rare variants of groups N, O

214 and P.

215 This wide panel of specimens tested demonstrated that the performance of Xpert HIV-1

216 VL assay was globally close to that of the Abbott HIV-1 RealTime assay. Comparison of the

217 quantifiable samples (i.e. > 40 cp/mL) by both techniques showed a remarkable correlation

218 between the two systems regardless of the VL level since the mean difference was almost zero,

219 what we did not observe in previous comparisons of viral load assays [4, 6]. Contrary to data

220 reported recently by Mor et al., i.e. many samples presenting a mean difference of more than 0.5


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221 in favor of Xpert [10], only two samples in the current study were discrepant over 0.5 Log, with

222 one very close to this limit at 0.53 Log. As Mor et al. did not describe the subtype characteristics

223 of the samples involved in these differences, it is not possible to assess the impact of genetic

224 diversity from their samples; in particular, involvement of a particular subtype that will not be

225 well represented in our panel. Distribution of VL values is also highly correlated with very little

226 scattering of points in a variance of +/-0.4 Log. This very strong correlation is even more

227 interesting given that the technologies and regions of amplification are different between the two

228 assays. For the rarest divergent variants group N, O and P, circulating in Central Africa and in the

229 countries associated with this region, the results showed a good correlation between the two

230 assays with a more significant quantification for some samples with the Xpert HIV-1 VL kit. In

231 particular, an underquantification of group N samples by the Abbott test was observed,
232 confirming data of a previous evaluation [4]. The most important differences observed for some

233 samples of group O showed that these variants remain problematic, and highlight the need to

234 determine what is the best tool before follow-up [11].


235 Concerning the concordance of assay results near the limit of quantification, data showed

236 that 11 quantifiable samples above the threshold of 40 cp/mL with the Abbott technique, were

237 undetectable with the Cepheid assay. These data seem concordant with those reported by Mor et

238 al., but rigorous comparison is difficult, since the authors did not distinguish undetectable from

239 detected samples below 40 cp/mL and did not indicate mean or median values for these samples

240 [10]. As all of these samples in our work correspond to very low values (less than 80 cp/mL)

241 below the clinical VL cut-off of 1,000 cp/mL defined by WHO [12], this assay could improve the

242 patient monitoring in low income settings [13]. Although considering a definition of the

243 virological failure at 50 cp/mL [14, 15], these undetectable samples could be wrongly considered

244 to be a virological success following ART ; this relative sensitivity could be an issue for


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245 monitoring especially using integrase inhibitors, since virological escape was observed at low

246 viral load [16]. Low values (below 170 cp/mL) were also found for the 40 samples quantifiable

247 with the Abbott technique but detected with Cepheid. Analysis of genetic diversity of these

248 samples showed that these results were not related to a particular subtype, since it involved

249 samples of subtype B (n=26) as well as non-B subtypes (n=20) or non-typable (n=5).

250 It is important to note that a low indeterminate rate of approximately 3% was observed

251 with the Cepheid assay. This rate, rarely shown in studies, should be considered, because it is

252 important for questions of cost and time for results; a low indeterminate rate is an indirect

253 indicator of quality and robustness of the test. Unfortunately insufficient samples were available

254 to allow for assessment of whether invalid results were sample specific or incidental.

255 The use of this assay and its unique technology in a cartridge has proved extremely simple
256 compared to the Abbott platform. The volume requirement of 1 mL of plasma to add to the

257 single-use disposable cartridge greatly simplifies the technical task and can be performed by any

258 staff member from reference laboratories to point of care sites. This is an extremely positive and

259 favorable element for implementation in small laboratories or in difficult field conditions. In

260 addition, the achievement of extraction and amplification in a single cartridge within a relatively

261 short time period (90 minutes) is also extremely important to be able to respond in emergency or

262 on demand situations, as compared to the longer and heavier workflow of the series with the

263 Abbott platform.


265 Other positive elements of this technology are the absence of mechanical requirements

266 (i.e. for extraction), and the ease of transport (for a module of 4 cartridges), allowing the

267 technology to be implemented anywhere, especially in low income countries. The GeneXpert can

268 also be used in larger laboratories for high-volume samples with up to 80 modules or even for


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269 emergency use since the technology is not burdened by the series process. The sample addition to

270 the cartridge remains manual compared to the automated Abbott platform. This can be a

271 disadvantage but if the test is carried out in 'real time', this manual step is equivalent to a stage of

272 manual decantation of the blood samples for freezing prior to testing on the platform.


274 In conclusion, this study, using a wide panel in terms of number of samples, genetic

275 diversity and VL ranges, allowed us to evaluate this new assay in contexts representative of

276 diverse clinical situations and genetic background. Our data showed that the Xpert HIV-1 Viral

277 Load assay offered very good performance for detection and quantification of the current HIV-1

278 genetic diversity. The samples quantifiable by the Abbott assay that were below 40 cp/mL by the

279 Cepheid technique are an issue and requires further studies with non-selected samples near the
280 limit of quantification. It is also necessary now to compare this new test to other assays, such as

281 the Roche Cobas Taqman HIV-1, also widely used, to confirm its global performances. In

282 addition, the practicability of this new assay makes it a useful and interesting tool both for high

283 income and in low income countries, where it could facilitate and improve follow-up of patients.


We thank Cepheid, the Institut de Veille Sanitaire and the CHU de Rouen for financial support.

We also thank the patients for the COREVIH Haute-Normandie and all the staff of the

laboratory, particularly Fabienne De Oliveira and Marie Leoz, for characterizing the collection



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284 1. Labbett W, Garcia-Diaz A, Fox Z, Clewley GS, Fernandez T, Johnson M, et al.

285 Comparative evaluation of the ExaVir Load version 3 reverse transcriptase assay for
286 measurement of human immunodeficiency virus type 1 plasma load. J Clin Microbiol
287 2009,47:3266-3270.
288 2. Bourlet T, Signori-Schmuck A, Roche L, Icard V, Saoudin H, Trabaud MA, et al. HIV-1
289 load comparison using four commercial real-time assays. J Clin Microbiol 2011,49:292-

290 297.
291 3. Garcia-Diaz A, Labbett W, Clewley GS, Guerrero-Ramos A, Geretti AM. Comparative
292 evaluation of the Artus HIV-1 QS-RGQ assay and the Abbott RealTime HIV-1 assay for
293 the quantification of HIV-1 RNA in plasma. J Clin Virol 2013,57:66-69.

294 4. Mourez T, Delaugerre C, Vray M, Lemee V, Simon F, Plantier JC. Comparison of the
295 bioMerieux NucliSENS EasyQ HIV-1 v2.0-HIV-1 RNA quantification assay versus
296 Abbott RealTime HIV-1 and Roche Cobas TaqMan HIV-1 v2.0 on current epidemic
297 HIV-1 variants. J Clin Virol 2015,71:76-81.
298 5. Hopkins M, Hau S, Tiernan C, Papadimitropoulos A, Chawla A, Beloukas A, et al.
299 Comparative performance of the new Aptima HIV-1 Quant Dx assay with three
300 commercial PCR-based HIV-1 RNA quantitation assays. J Clin Virol 2015,69:56-62.
301 6. Sire JM, Vray M, Merzouk M, Plantier JC, Pavie J, Maylin S, et al. Comparative RNA
302 quantification of HIV-1 group M and non-M with the Roche Cobas AmpliPrep/Cobas
303 TaqMan HIV-1 v2.0 and Abbott Real-Time HIV-1 PCR assays. J Acquir Immune Defic
304 Syndr 2011,56:239-243.
305 7. Raja S, Ching J, Xi L, Hughes SJ, Chang R, Wong W, et al. Technology for automated,
306 rapid, and quantitative PCR or reverse transcription-PCR clinical testing. Clin Chem
307 2005,51:882-890.
308 8. Mourez T, Simon F, Plantier JC. Non-M variants of human immunodeficiency virus type

309 1. Clin Microbiol Rev 2013,26:448-461.

310 9. Plantier JC, Dachraoui R, Lemee V, Gueudin M, Borsa-Lebas F, Caron F, et al. HIV-1
311 resistance genotyping on dried serum spots. AIDS 2005,19:391-397.
312 10. Mor O, Gozlan Y, Wax M, Mileguir F, Rakovsky A, Noy B, et al. Evaluation of the

313 RealTime HIV-1, Xpert HIV-1, and Aptima HIV-1 Quant Dx Assays in Comparison to
314 the NucliSens EasyQ HIV-1 v2.0 Assay for Quantification of HIV-1 Viral Load. J Clin
315 Microbiol 2015,53:3458-3465.
316 11. Gueudin M, Leoz M, Lemee V, De Oliveira F, Vessiere A, Kfutwah A, et al. A new real-

317 time quantitative PCR for diagnosis and monitoring of HIV-1 group O infection. J Clin
318 Microbiol 2012,50:831-836.
319 12. Organization WH. Consolidated guidelines on the use of antiretroviral drugs for treating
320 and preventing HIV infection : Recommendations for a Public health Approach. 2013.
321 13. Essajee S, Kumarasamy N. Commentary: The monitoring of adults and children on
322 antiretroviral therapy in the 2013 WHO consolidated ARV guidelines. AIDS 2014,28
323 Suppl 2:S147-149.
324 14. European AIDS Clinical Society (EACS) guidelines Version 6.1. 2012.
325 15. ANRS. Prise en charge mdicale des personnes vivant avec le VIH. Recommandations du
326 groupe dexperts. Sous la direction du Pr Morlat; 2013.


Copyright 2016 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited.
327 16. Marcelin AG, Delaugerre C, Beaudoux C, Descamps D, Morand-Joubert L, Amiel C, et
328 al. A cohort study of treatment-experienced HIV-1-infected patients treated with
329 raltegravir: factors associated with virological response and mutations selected at failure.
330 Int J Antimicrob Agents 2013,42:42-47.


332 Figure 1. Scatter diagram and the regression line according to Passing & Bablok on the 162

333 samples quantifiable by both the Cepheid and Abbott assays. In addition to the regression line
334 (solid line), the confidence interval for the regression line (dashed lines) is displayed. Samples

335 are differentiated between subtype B ( , n=58), subtype CRF02_AG ( , n=19), and other non-B
336 ( , n=85).
337 Figure 2. Degree of agreement in Log cp/mL for the 162 samples quantifiable by both the
338 Cepheid and Abbott assays. In the graphical method of Bland and Altman, the differences
339 between the two techniques are plotted against the averages of the two techniques. Samples are
340 differentiated between subtype B ( , n=58), subtype CRF02_AG ( , n=19), and other non-B ( ,
341 n=85).
342 Figure 3. HIV-1 VL results obtained for the 3 dilutions of supernatants of groups N, O and P. In
343 the Box-and-Whisker plot, the central box represents the values from the lower to upper quartile
344 (25 to 75 percentile) and the middle line represents the median. For each dilution, groups are

345 differentiated between group O ( , n=15), group N ( , n=2) and group P ( , n=1).


Copyright 2016 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited.
Table 1. Qualitative results for Xpert HIV-1 VL and Abbott RealTime HIV-1 assays

Abbott HIV-1 RealTime

undetectable detected quantifiable total

undetectable 38 14 11 63

Cepheid detected 7 13 40 60
Xpert HIV-1
VL quantifiable 0 0 162 162

total 45 27 213 285

Copyright 2016 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited.

Copyright 2016 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited.

Copyright 2016 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited.

Copyright 2016 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited.