Draft Genome Sequence of a Bordetella pertussis Strain with the Virulence-Associated Allelic Variant ptxP3, Isolated in Italy.

Despite a universal immunization program, pertussis has persisted and resurged, and is of particular concern for infants in terms of morbidity and mortality. Here, we report the genome sequence of a Bordetella pertussis strain with the virulence-associated allelic variant ptxP3, isolated from a 45-day-old infant.

Sporadic isolation of sabin-like polioviruses and high-level detection of non-polio enteroviruses during sewage surveillance in seven Italian cities, after several years of inactivated poliovirus vaccination.

Sewage surveillance in seven Italian cities between 2005 and 2008, after the introduction of inactivated poliovirus vaccination (IPV) in 2002, showed rare polioviruses, none that were wild-type or circulating vaccine-derived poliovirus (cVDPV), and many other enteroviruses among 1,392 samples analyzed. Two of five polioviruses (PV) detected were Sabin-like PV2 and three PV3, based on enzyme-linked immunosorbent assay (ELISA) and PCR results. Neurovirulence-related mutations were found in the 5'noncoding region (5'NCR) of all strains and, for a PV2, also in VP1 region 143 (Ile>Thr). Intertypic recombination in the 3D region was detected in a second PV2 (Sabin 2/Sabin 1) and a PV3 (Sabin 3/Sabin 2). The low mutation rate in VP1 for all PVs suggests limited interhuman virus passages, consistent with efficient polio immunization in Italy. Nonetheless, these findings highlight the risk of wild or Sabin poliovirus reintroduction from abroad. Non-polio enteroviruses (NPEVs) were detected, 448 of which were coxsackievirus B (CVB) and 294 of which were echoviruses (Echo). Fifty-six NPEVs failing serological typing were characterized by sequencing the VP1 region (nucleotides [nt] 2628 to 2976). A total of 448 CVB and 294 Echo strains were identified; among those strains, CVB2, CVB5, and Echo 11 predominated. Environmental CVB5 and CVB2 strains from this study showed high sequence identity with GenBank global strains. The high similarity between environmental NPEVs and clinical strains from the same areas of Italy and the same periods indicates that environmental strains reflect the viruses circulating in the population and highlights the potential risk of inefficient wastewater treatments. This study confirmed that sewage surveillance can be more sensitive than acute flaccid paralysis (AFP) surveillance in monitoring silent poliovirus circulation in the population as well as the suitability of molecular approaches to enterovirus typing.

Aseptic meningitis outbreak caused by echovirus 30 in two regions in Bulgaria, May-August 2012.

An aseptic meningitis outbreak emerged in two regions in Bulgaria in 2012 and echovirus 30 (E30) was established as the aetiological agent by cell culture isolation, serological test, and molecular-based techniques. A total of 157 patients with aseptic meningitis were investigated, of which 117 were confirmed as having E30-associated disease. Molecular analysis of 12 E30 isolates revealed 99-100% nucleotide and amino-acid identity between them and a close correlation with a Greek strain involved in an E30 outbreak in 2012. Children aged 5-14 years were mainly affected, which could reflect the absence of E30 epidemics in Bulgaria for a period of 11 years. The first case with E30 isolation (a 2-year-old patient from Plovdiv) was notified at the end of April 2012. This was most likely the index case, from which the spread of the virus started, causing sporadic cases first, which later led to an aseptic meningitis outbreak facilitated by person-to-person viral transmission.

[Evaluation of the Surveillance System of Acute Flaccid Paralysis in Puglia: 5 years of work]. / Valutazione del Sistema di Sorveglianza delle Paralisi Flaccide Acute in Puglia: cinque anni di attività.

Surveillance of acute flaccid paralysis (AFP) is the milestone to monitor the progress toward poliomyelitis eradication aim, fixed by WHA in 1988. Active AFP surveillance started in Apulia in 1997; this work evaluates five-year period activities. In this period, the total number of cases notified was 48, 7 of which were resident out of Apulia. Twenty-five were males and 23 females; the age ranged between 1 month and 15 years. Any collected serum specimens showed protective antibody levels against polioviruses. Polioviruses type 1 and type 2 Sabin-like were isolated from stool samples collected from two AFP patients. AFP surveillance targets improved in the years, with only exception, in 2001, of second serum specimen collected within 14 days because of children were discharged earlier form the hospitals. Apulia experience demonstrates the achievement of good levels of AFP surveillance targets. System sensitivity has been optimal in 2001 with a number of notified cases threefold the expected value and adequate specimen sampling (80%). Additional involved hospitals and availability of increased and dedicated human resources contributed to this outcome. The effort to achieve WHO targets for AFP surveillance needs to be maintained in next years until global certification of eradication will be declared.

Antigenic sites of poliovirus type 3 eliciting IgA monoclonal antibodies in orally immunized mice.

A panel of neutralizing IgA monoclonal antibodies was produced from mice orally inoculated with poliovirus type 3 Sabin and cholera toxin as adjuvant. Low levels of neutralizing antibodies were elicited in mice after several boosts, but only in the presence of cholera toxin. Characterization of IgA MAbs by neutralization-escape virus mutants showed that all but one neutralizing MAbs against type 3 poliovirus were directed to antigenic site N-AgIII, which was previously found by us to be the major target of mucosal immune response to Sabin 1 in the mouse. Our data indicate that residue 236 of VP3, not previously reported, is also involved in forming site N-AgIII in addition to formerly described VP3 (aa 58-59) and VP1 (aa 286-290) residues. Unlike poliovirus type 1 IgA MAbs, all IgA MAbs herein described neutralized the wild-type parental poliovirus.

Poliovirus Sabin type 1 neutralization epitopes recognized by immunoglobulin A monoclonal antibodies.

Immunity to poliomyelitis is largely dependent on humoral neutralizing antibodies, both after natural (wild virus or vaccine) infection and after inactivated poliovirus vaccine inoculation. Although the production of local secretory immunoglobulin A (IgA) antibody in the gut mucosa may play a major role in protection, most of information about the antigenic determinants involved in neutralization of polioviruses derives from studies conducted with humoral monoclonal antibodies (MAbs) generated from parenterally immunized mice. To investigate the specificity of the mucosal immune response to the virus, we have produced a library of IgA MAbs directed at Sabin type 1 poliovirus by oral immunization of mice with live virus in combination with cholera toxin. The epitopes recognized by 13 neutralizing MAbs were characterized by generating neutralization-escape virus mutants. Cross-neutralization analysis of viral mutants with MAbs allowed these epitopes to be divided into four groups of reactivity. To determine the epitope specificity of MAbs, virus variants were sequenced and the mutations responsible for resistance to the antibodies were located. Eight neutralizing MAbs were found to be directed at neutralization site N-AgIII in capsid protein VP3; four more MAbs recognized site N-AgII in VP1 or VP2. One IgA MAb selected a virus variant which presented a unique mutation at amino acid 138 in VP2, not previously described. This site appears to be partially related with site N-AgII and is located in a loop region facing the VP2 N-Ag-II loop around residue 164. Only 2 of 13 MAbs proved able to neutralize the wild-type Mahoney strain of poliovirus. The IgA antibodies studied were found to be produced in the dimeric form needed for recognition by the polyimmunoglobulin receptor mediating secretory antibody transport at the mucosal level.

Comparison of capillary blood versus venous blood samples in the assessment of immunity to measles.

Seroepidemiological investigations are essential for assessing the efficacy of measles vaccination programmes. However, when large-scale sampling is needed, a major difficulty is the problem of taking venous blood, especially in children. An alternative method is the collection of capillary blood samples spotted on filter papers. The eluted extract from these 'blood' spots can be used instead of serum samples for measles laboratory diagnosis or investigations. Measles antibody detection is readily carried out by ELISA on serum samples. The same technique can be used on eluates from capillary blood spots. Measles antibody titres determined on matched serum and blood spot samples from 27 children were compared. A strong correlation was found between the results obtained with the two methods of blood sampling.