Rotavirus Vaccine Virus Shedding, Viremia and Clearance in Infants with Severe Combined Immune Deficiency

Abstract

We report for the first time Rotarix vaccine-acquired rotavirus infections with viremia in 2 infants vaccinated before being diagnosed with severe combined immune deficiency. Monitoring the first infant revealed that persistent rotavirus infection resolved after complete immune reconstitution was achieved by gene therapy.

Two live attenuated rotavirus vaccines are recommended by the World Health Organization for the routine immunization of infants¹ and are progressively being introduced in universal mass vaccination programs: the pentavalent human-bovine reassortant vaccine RotaTeq^® (RV5; Merck and Co, Whitehouse Station, NJ) and the single-strain human G1P[8] vaccine Rotarix^® (RV1; GlaxoSmithKline Biologicals, Rixensart, Belgium).

These vaccines are contraindicated in infants diagnosed with severe combined immune deficiency (SCID).² In the absence of neonatal screening for SCID, rotavirus vaccines may be administered before the diagnosis is made, resulting in severe and persistent gastroenteritis with prolonged vaccine viral shedding as previously reported for the RV5 vaccine.^3–7

We report 2 cases of infants subsequently diagnosed with SCID who developed severe and persistent rotavirus diarrhea with failure to thrive after receiving the second dose of RV1 vaccine. The presence of viremia and the vaccine origin of the rotavirus infections were demonstrated. The rotavirus infection was monitored during immune reconstitution until virus clearance in 1 of the infants.

CASE REPORTS

Patient 1 was a full-term male infant born without complications. He received the second dose of RV1 vaccine at 3 months of age, after which he developed chronic diarrhea with failure to thrive. Rotavirus was found in stool specimens with no concomitant enteric viruses. At 5 months of age, he developed respiratory distress due to Pneumocystis jiroveci infection. Agammaglobulinemia and profound T lymphopenia (first CD3+ cells count: 7%, 259 cells/mm³. Normal CD3+ count: 2500–5600 cells/mm³) led to a diagnosis of SCID. A mutation was found in the common gamma chain of the interleukin-2 receptor (X-linked SCID). In the absence of an HLA-matched donor, the child was included in a gene therapy trial (ClinicalTrials.gov Identifier: NCT01410019) at the age of 6 months. Progressive immune reconstitution was observed, with >1500 CD3+ cells/mm³ 2 months after a second infusion of transduced CD34+ cells. Diarrhea lasted until the age of 13 months but with a gradual decrease, and parenteral nutrition was stopped at 10 months of age.

Patient 2, a full-term female infant, presented failure to thrive at 2 months of age. She received the second dose of RV1 vaccine at 3 months of age, and she was hospitalized at 4 months of age for chronic diarrhea, feeding difficulties, failure to thrive and BCG-related axillary adenopathy with fistulization. Rotavirus was found in stool specimens with no concomitant enteric viruses. Agammaglobulinemia and the quasi-absence of T cells (first CD3+ cells count: 3%, 15 cells/mm³) led to the diagnosis of SCID. Adenosine deaminase (ADA) deficiency was established in red cells and genetically confirmed. In the absence of an HLA-matched donor, the child received pegylated-ADA replacement therapy⁸ and she is awaiting her inclusion in a gene therapy trial (pending at the time of this report). A progressive increase in lymphocyte counts was observed with 336 CD3+ cells/mm³ 3 months after the beginning of enzymotherapy. Although reduced under parenteral nutrition, diarrhea persisted.

METHODS

To characterize and monitor these rotavirus infections, we investigated seven stool samples (from D-16 through D+278 from the beginning of gene therapy) and three blood samples (D+31, D+123 and D+278) for the first infant, and three stool samples (D-3, D+43 and D+78 from the beginning of the pegylated-ADA treatment) and two blood samples (D+1 and D+42) for the second infant.

Rotavirus was detected and viral load was assessed by real time reverse transcription-polymerase chain reaction targeting the Viral Protein (VP) 2 coding gene.⁹ Rotavirus strains were characterized by sequencing the partial or complete genes coding for the VP7, VP4, VP6 and nonstructural protein (NSP) 4 as previously described.¹⁰ To determine the origin of the rotavirus strains detected, the same genes were sequenced for the RV1 vaccine strain (lot number: AROLA490AB) and for a contemporary G1P[8] epidemic strain detected in the stools of another infant at the same hospital.

RESULTS

Rotavirus was detected in the initial stool and blood samples of both infants. Sequence analyses of the rotavirus strains detected in the initial stool samples revealed very high nucleotide and amino-acid identities for VP7 (>99.8% and >99.3%, respectively), VP4 (99.8% and 99.3%, respectively), VP6 (100% identical) and NSP4 (99.9% and 99.4%, respectively) with the RV1 vaccine strain. These identities were considerably higher than those observed with the contemporary G1P[8] epidemic strain (see Tables, Supplemental Digital Content 1–4, https://links.lww.com/INF/C20, https://links.lww.com/INF/C21, https://links.lww.com/INF/C22, and https://links.lww.com/INF/C23, which show sequence identities).

Only the VP7 and NSP4 coding genes were successfully sequenced in the initial blood sample of the first infant. The sequence analysis of the VP7 coding gene revealed very high nucleotide and amino-acid identities with the strain detected in the stools of this infant (99.8% and 99.6%, respectively) and with the vaccine strain (99.6% and 99.3%, respectively). The NSP4 sequence of the blood strain was identical to that in the stool sample, thus showing very high identities with the vaccine strain. For both genes, identities with the contemporary G1P[8] epidemic strain were lower (see Tables, Supplemental Digital Content 1 and 4, https://links.lww.com/INF/C20, and https://links.lww.com/INF/C23, which show sequence identities). No rotavirus gene sequence was obtained for the initial blood sample of the second infant.

The strains found in the infants, the RV1 vaccine strain and the contemporary epidemic strain belonged to the same genotypes (G1, P[8], I1 and E1 for VP7, VP4, VP6 and NSP4, respectively). Nevertheless, the identity rates observed strongly indicated that the strains found in the infants were vaccine-derived.The sequences of these strains were not totally identical to the vaccine sequences possibly because of genetic drift since the administration of the vaccine or because of the in vivo selection of a minor variant already present in the vaccine.

The vaccine origin of the strains detected in the subsequent stool samples of both infants was also confirmed by the sequence analyses of at least 2 genes (data not shown). Of note, rotavirus was not detected in subsequent blood samples.

Vaccine rotavirus was still detected in the stools of the first infant at D+181, but the viral load decreased from 8.10 × 10¹² copies/g of feces at the time of gene therapy to 2.23 × 10⁵ copies/g 6 months later, with the progressive decline corresponding to progressive reconstitution of the immune system (Fig. 1). Indeed, the total blood CD3+, CD4+ and CD8+ cell counts were extremely low and the fecal viral load was very high before gene therapy, whereas the viral load decreased when the number of T-cells increased after the gene therapy boost performed at D+63. No immune reconstitution syndrome, and notably no clinical sign of intussusception, was observed.

FIGURE 1: Rotavirus viral load in stools (black squares) and blood (open stars), and total blood CD3+ (black triangle), CD4+ (open circle) and CD8+ (times symbol) cell counts of the first infant according to time. The day of the first infusion of gene therapy is considered day zero (D0).

The decrease in the viral load was also associated with diminished clinical signs of gastroenteritis, highlighted by a continuous decrease in the number of loose stools per day (from initially 5 to 7/d to 2 at D+181, data not shown). Soon after the last detection of rotavirus in stools, the infant became nondiarrheic and the virus was undetectable at D+278.

In the blood, rotavirus was only detected at D+31 with a load of 6.77 × 10⁴ copies/mL.

In the second infant, the rotavirus load in the stools was initially very high (9.18 × 10¹¹ copies/g at D-3) and relatively stable 43 days after the beginning of the pegylated-ADA treatment (3.14 × 10¹² copies/g) but was lower at D+78 (4.2 × 10⁷ copies/g) at the beginning of an enzymotherapy-induced immune recovery. Viremia was observed only at D+1 with a viral load of 6.79 × 10³ copies/mL (data not shown). Of note, the low viral load in blood samples may explain the difficulties encountered for the amplification of rotavirus genes.

DISCUSSION

Although it has already been described for RV5 vaccine,^3–7 this is the first report clearly demonstrating vaccine-acquired rotavirus infections in 2 SCID infants vaccinated with RV1 before being diagnosed with immune deficiency. One previous study reported rotavirus infection subsequent to RV1 vaccine administration in 1 SCID infant⁶ but the strain responsible for this infection was unspecified, leading only to a presumption that it was RV1-derived. The virus was initially shed in both infants with viral loads (around 10¹² copies/g) higher than those reported after the second dose of RV1 vaccine in the general population (around 10⁷ copies/g),¹¹ suggesting that this attenuated vaccine strain has high replication ability in a context of immunodeficiency. Monitoring of the first infant revealed that the fecal shedding decreased continuously during immune reconstitution following gene therapy, until total clearance after complete immune reconstitution. Preliminary data for the second infant confirmed that the fecal shedding decreased only after induction of the immune reconstitution. For the first infant, the vaccine shedding lasted more than 6 months, which is within the range of the shedding time reported for RV5 vaccine-derived strains in infants treated for SCID (ie, 5–12 months)^3,4,7 and substantially longer than 15 days reported after the second dose of RV1 vaccine in healthy children.¹¹

Viremia is common during natural rotavirus infection,¹² but this report gives the first evidence of viremia induced by RV1 vaccine, and confirms that attenuated rotavirus strains could spread outside the intestine of infants with primary immune deficiency as previously reported for RV5 vaccine.⁴ Whether the low viremia and the initiated immune reconstitution can explain the disappearance of the virus in the blood earlier than in the stools needs to be investigated further.

Although this report does not dispute the benefit of rotavirus vaccines in the general population, it raises concerns about the safety of these vaccines in severely immunocompromised patients and fuels potential interest for neonatal screening of SCID.