Evaluation of in vitro cross-reactivity to avian H5N1 and pandemic H1N1 2009 influenza following prime boost regimens of seasonal influenza vaccination in healthy human subjects: A randomised trial

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High rates of apoptosis in human Mycobacterium ulcerans culture-positive Buruli ulcer skin lesions

Buruli ulcer, a disease caused by Mycobacterium ulcerans, causes ulcerative skin disease likely generated by a toxin that mediates apoptosis. We analyzed paraffin-embedded sections of surgically excised Buruli ulcer lesions (two ulcers and one edematous plaque) and adjacent non-lesional skin samples (n = 9) for apoptosis by an indirect immunofluorescent terminal deoxynucleotide transferase-mediated dUTP-digoxigenin nick end labeling (TUNEL) assay. All samples were stained for acid-fast bacilli (AFB) and cultured for mycobacteria, and most were analyzed with an M. ulcerans-specific diagnostic polymerase chain reaction (PCR). TUNEL (+) bodies were numerous in both ulcers and the plaque, and sparse or absent in adjacent non-lesional skin. The AFB tissue stains and cultures for M. ulcerans were positive only in the three lesions. The result of the PCR for M. ulcerans was positive in all three lesions and in four of six non-lesional tissue samples; three contained sparse TUNEL (+) bodies. An abundance of TUNEL (+) bodies in the three AFB stain (+), culture (+), and PCR (+) Buruli ulcer lesional samples, but not in nearby AFB stain (-), culture (-), and PCR (+) non-lesional skin samples, strengthen the evidence that apoptosis is an important tissue destruction mechanism in human lesions closely associated with viable M. ulcerans

Evaluation of in vitro cross-reactivity to avian H5N1 and pandemic H1N1 2009 influenza following prime boost regimens of seasonal influenza vaccination in healthy human subjects: a randomised trial.

Recent studies have demonstrated that inactivated seasonal influenza vaccines (IIV) may elicit production of heterosubtypic antibodies, which can neutralize avian H5N1 virus in a small proportion of subjects. We hypothesized that prime boost regimens of live and inactivated trivalent seasonal influenza vaccines (LAIV and IIV) would enhance production of heterosubtypic immunity and provide evidence of cross-protection against other influenza viruses.In an open-label study, 26 adult volunteers were randomized to receive one of four vaccine regimens containing two doses of 2009-10 seasonal influenza vaccines administered 8 (±1) weeks apart: 2 doses of LAIV; 2 doses of IIV; LAIV then IIV; IIV then LAIV. Humoral immunity assays for avian H5N1, 2009 pandemic H1N1 (pH1N1), and seasonal vaccine strains were performed on blood collected pre-vaccine and 2 and 4 weeks later. The percentage of cytokine-producing T-cells was compared with baseline 14 days after each dose.Subjects receiving IIV had prompt serological responses to vaccine strains. Two subjects receiving heterologous prime boost regimens had enhanced haemagglutination inhibition (HI) and neutralization (NT) titres against pH1N1, and one subject against avian H5N1; all three had pre-existing cross-reactive antibodies detected at baseline. Significantly elevated titres to H5N1 and pH1N1 by neuraminidase inhibition (NI) assay were observed following LAIV-IIV administration. Both vaccines elicited cross-reactive CD4+ T-cell responses to nucleoprotein of avian H5N1 and pH1N1. All regimens were safe and well tolerated.Neither homologous nor heterologous prime boost immunization enhanced serum HI and NT titres to 2009 pH1N1 or avian H5N1 compared to single dose vaccine. However heterologous prime-boost vaccination did lead to in vitro evidence of cross-reactivity by NI; the significance of this finding is unclear. These data support the strategy of administering single dose trivalent seasonal influenza vaccine at the outset of an influenza pandemic while a specific vaccine is being developed.ClinicalTrials.gov NCT01044095

Baseline characteristics in 26 healthy subjects, n (%).

<p>LAIV = live attenuated influenza vaccine; IIV = inactivated influenza vaccination; * as assessed by haemagglutination inhibition, neutralization and neuraminidase inhibition assays.</p

Individual and geometric mean serum haemagglutination inhibition assay, H5pp assay and serum neuraminidase inhibition assay results against avian H5N1 virus in 26 healthy human volunteers measured at baseline and two weeks following each dose of prime boost seasonal influenza vaccination (2 doses administered 8 weeks apart).

<p>Individual and geometric mean serum haemagglutination inhibition assay, H5pp assay and serum neuraminidase inhibition assay results against avian H5N1 virus in 26 healthy human volunteers measured at baseline and two weeks following each dose of prime boost seasonal influenza vaccination (2 doses administered 8 weeks apart).</p

CONSORT (2010) flow diagram.

<p>CONSORT (2010) flow diagram.</p

Individual and geometric mean serum haemagglutination inhibition assay, microneutralization assay and serum neuraminidase inhibition assay results against pandemic H1N1 2009 virus in 26 healthy human volunteers measured at baseline and two weeks following each dose of prime boost seasonal influenza vaccination (2 doses administered 8 weeks apart).

<p>Individual and geometric mean serum haemagglutination inhibition assay, microneutralization assay and serum neuraminidase inhibition assay results against pandemic H1N1 2009 virus in 26 healthy human volunteers measured at baseline and two weeks following each dose of prime boost seasonal influenza vaccination (2 doses administered 8 weeks apart).</p

Adverse events reported during 28 days following Doses 1 and 2 of trivalent seasonal influenza vaccine: number of adverse events (number of vaccine-attributable adverse events).

<p>LAIV = live attenuated influenza vaccine; IIV = inactivated influenza vaccination; AE = adverse event.</p