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

Evaluation of the CareStart™ glucose-6-phosphate dehydrogenase (G6PD) rapid diagnostic test in the field settings and assessment of perceived risk from primaquine at the community level in Cambodia.

BACKGROUND:Primaquine is an approved radical cure treatment for Plasmodium vivax malaria but treatment can result in life-threatening hemolysis if given to a glucose-6-phosphate dehydrogenase deficient (G6PDd) patient. There is a need for reliable point-of-care G6PD diagnostic tests. OBJECTIVES:To evaluate the performance of the CareStart™ rapid diagnostic test (RDT) in the hands of healthcare workers (HCWs) and village malaria workers (VMWs) in field settings, and to better understand user perceptions about the risks and benefits of PQ treatment guided by RDT results. METHODS:This study enrolled 105 HCWs and VMWs, herein referred to as trainees, who tested 1,543 healthy adult male volunteers from 84 villages in Cambodia. The trainees were instructed on G6PD screening, primaquine case management, and completed pre and post-training questionnaires. Each trainee tested up to 16 volunteers in the field under observation by the study staff. RESULTS:Out of 1,542 evaluable G6PD volunteers, 251 (16.28%) had quantitative enzymatic activity less than 30% of an adjusted male median (8.30 U/g Hb). There was no significant difference in test sensitivity in detecting G6PDd between trainees (97.21%), expert study staff in the field (98.01%), and in a laboratory setting (95.62%) (p = 0.229); however, test specificity was different for trainees (96.62%), expert study staff in the field (98.14%), and experts in the laboratory (98.99%) (p < 0.001). Negative predictive values were not statistically different for trainees, expert staff, and laboratory testing: 99.44%, 99.61%, and 99.15%, respectively. Knowledge scores increased significantly post-training, with 98.7% willing to prescribe primaquine for P.vivax malaria, an improvement from 40.6% pre-training (p < 0.001). CONCLUSION:This study demonstrated ability of medical staff with different background to accurately use CareStart™ RDT to identify G6PDd in male patients, which may enable safer prescribing of primaquine; however, pharmacovigilance is required to address possible G6PDd misclassifications

MOESM1 of Ex vivo piperaquine resistance developed rapidly in Plasmodium falciparum isolates in northern Cambodia compared to Thailand

Additional file 1. Artemisinin and piperaquine susceptibility profiles do not suggest cross resistance

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

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

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

Individual and geometric mean peripheral blood CD4+ T-cell responses measured at baseline, two weeks after dose 1 and two weeks after dose 2, by study group: (A) against vaccine antigens in IIV, (B) against nucleoprotein H5N1, and (C) against nucleoprotein pH1N1 2009.

<p>Individual and geometric mean peripheral blood CD4+ T-cell responses measured at baseline, two weeks after dose 1 and two weeks after dose 2, by study group: (A) against vaccine antigens in IIV, (B) against nucleoprotein H5N1, and (C) against nucleoprotein pH1N1 2009.</p

Human Sentinel Surveillance of Influenza and Other Respiratory Viral Pathogens in Border Areas of Western Cambodia

<div><p>Little is known about circulation of influenza and other respiratory viruses in remote populations along the Thai-Cambodia border in western Cambodia. We screened 586 outpatients (median age 5, range 1–77) presenting with influenza-like-illness (ILI) at 4 sentinel sites in western Cambodia between May 2010 and December 2012. Real-time reverse transcriptase (rRT) PCR for influenza was performed on combined nasal and throat specimens followed by viral culture, antigenic analysis, antiviral susceptibility testing and full genome sequencing for phylogenetic analysis. ILI-specimens negative for influenza were cultured, followed by rRT-PCR for enterovirus and rhinovirus (EV/RV) and EV71. Influenza was found in 168 cases (29%) and occurred almost exclusively in the rainy season from June to November. Isolated influenza strains had close antigenic and phylogenetic relationships, matching vaccine and circulating strains found elsewhere in Cambodia. Influenza vaccination coverage was low (<20%). Western Cambodian H1N1(2009) isolate genomes were more closely related to 10 earlier Cambodia isolates (94.4% genome conservation) than to 13 Thai isolates (75.9% genome conservation), despite sharing the majority of the amino acid changes with the Thai references. Most genes showed signatures of purifying selection. Viral culture detected only adenovirus (5.7%) and parainfluenza virus (3.8%), while non-polio enteroviruses (10.3%) were detected among 164 culture-negative samples including coxsackievirus A4, A6, A8, A9, A12, B3, B4 and echovirus E6 and E9 using nested RT-PCR methods. A single specimen of EV71 was found. Despite proximity to Thailand, influenza epidemiology of these western Cambodian isolates followed patterns observed elsewhere in Cambodia, continuing to support current vaccine and treatment recommendations from the Cambodian National Influenza Center. Amino acid mutations at non-epitope sites, particularly hemagglutinin genes, require further investigation in light of an increasingly important role of permissive mutations in influenza virus evolution. Further research about the burden of adenovirus and non-polio enteroviruses as etiologic agents in acute respiratory infections in Cambodia is also needed.</p></div

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