Human Papillomavirus Antibody Reference Reagents for Use in Postvaccination Surveillance Serology

Suitably controlled serosurveillance surveys are essential for evaluating human papillomavirus (HPV) immunization programs. A panel of plasma samples from 18-year-old females was assembled, the majority of the samples being from recipients of the bivalent HPV vaccine. Antibody specificities were evaluated by three independent laboratories, and 3 pools that displayed no antibodies to any HPV type tested or intermediate or high levels of antibody to HPV16, HPV18, HPV31, and HPV45 were created. These pools will be useful as control reagents for HPV serology

No Evidence of XMRV or MuLV Sequences in Prostate Cancer, Diffuse Large B-Cell Lymphoma, or the UK Blood Donor Population

Xenotropic murine leukaemia virus-related virus (XMRV) is a recently described retrovirus which has been claimed to infect humans and cause associated pathology. Initially identified in the US in patients with prostate cancer and subsequently in patients with chronic fatigue syndrome, doubt now exists that XMRV is a human pathogen. We studied the prevalence of genetic sequences of XMRV and related MuLV sequences in human prostate cancer, from B cell lymphoma patients and from UK blood donors. Nucleic acid was extracted from fresh prostate tissue biopsies, formalin-fixed paraffin-embedded (FFPE) prostate tissue and FFPE B-cell lymphoma. The presence of XMRV-specific LTR or MuLV generic gag-like sequences was investigated by nested PCR. To control for mouse DNA contamination, a PCR that detected intracisternal A-type particle (IAP) sequences was included. In addition, DNA and RNA were extracted from whole blood taken from UK blood donors and screened for XMRV sequences by real-time PCR. XMRV or MuLV-like sequences were not amplified from tissue samples. Occasionally MuLV gag and XMRV-LTR sequences were amplified from Indian prostate cancer samples, but were always detected in conjunction with contaminating murine genomic DNA. We found no evidence of XMRV or MuLV infection in the UK blood donors

Details of sequences obtained.

<p>WB = Whole Blood.</p

Details of PCRs performed.

<p>400 nM concentrations of primers and 200 nM probes were used in all the TaqMan assays.</p

Maximum likelihood tree of gag sequences.

<p>The MLV RAxML tree for gag sequences lying between XMRV primers gagIF and gagIR was developed from the endogenous MLV sequences identified by Jern et al <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0019953#pone.0019953-Jern1" target="_blank">[19]</a>, sequences derived from CFS patients by Lo et al <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0019953#pone.0019953-Lo1" target="_blank">[5]</a> (shown in red and blue) and the sequences amplified in this study (in green). In addition to sequences from the RNA and DNA, “1F_131010” and “1F” are included. 1F_131010 was derived from the amplification of cDNA synthesized from RNA extracted from the tissue culture fluid supernatant of an XMRV culture, kindly provided by Professor M McClure. 1F was derived from the amplification of Balb/c mouse DNA (Sigma). Bootstrap values >50 are indicated.</p

Agarose gel analysis of R2 from experiment 1.

<p><i>gag</i> nested PCR for 80 cDNAs derived from blood donors and controls. Lanes 1A–1H ten fold dilution series of XMRV TCS (Lane 1A equivalent to 5.5×10⁶ to 5.5×10⁻¹ cDNA molecules/PCR – determined by Poisson distribution). Lanes 2A–11H 80 cDNAs derived from blood donor whole blood. Lanes 12A–12H ten fold dilution series of Balb/c DNA 10⁵ pg to 0.10 pg, negative.</p

Alignment between cfs1, MLV006, Pmv15 and Pmv22.

<p>The nine nucleotide differences between cfs1 and the other viral sequences are highlighted.</p