Induction of antibodies and T cell responses by a recombinant influenza virus carrying an HIV-1 TatΔ51-59 protein in mice.

Recombinant influenza viruses hold promise as vectors for vaccines to prevent transmission of mucosal pathogens. In this study, we generated a recombinant WSN/TatΔ(51-59) virus in which Tat protein lacking residues 51 to 59 of the basic domain was inserted into the N-terminus of the hemagglutinin (HA) of A/WSN/33 virus. The TatΔ(51-59) insertion into the viral HA caused a 2-log reduction in viral titers in cell culture, compared with the parental A/WSN/33 virus, and severely affected virus replication in vivo. Nevertheless, Tat-specific antibodies and T cell responses were elicited upon a single intranasal immunization of BALB/c mice with WSN/TatΔ(51-59) virus. Moreover, Tat-specific immune responses were also detected following vaccine administration via the vaginal route. These data provide further evidence that moderately large HIV antigens can be delivered by chimeric HA constructs and elicit specific immune responses, thus increasing the options for the potential use of recombinant influenza viruses, and their derivatives, for prophylactic and therapeutic vaccines.

Vaccination for seasonal influenza in patients with cancer: recommendations of the Italian Society of Medical Oncology (AIOM).

BACKGROUND: Influenza virus causes annual epidemics in the winter-spring season with significant morbidity in the general population and important mortality in high-risk groups, including cancer patients. Opinions on the suitability of patients with malignancies not undergoing active treatment and in different phases of antineoplastic therapy, to receive influenza vaccination, vary considerably among oncologists, sometimes even within one center. METHODS: We reviewed available data, including recommendations by national health authorities, on impact of influenza in patients with cancer and their capacity to mount protective immunological responses to vaccination, thus allowing, on behalf of Italian Association of Medical Oncology, to make suitable recommendations for the prevention and treatment of seasonal influenza. RESULTS AND DISCUSSION: Patients with cancer often have disease- or treatment-related immunosuppression, and as a consequence, they may have a suboptimal serologic response to influenza vaccination. The protective effect of the different preparations of influenza vaccines in patients with cancer has not been widely investigated, especially in adult patients harboring solid tumors. The optimal timing for administration of influenza vaccines in patients receiving chemotherapy is also not clearly defined. However, since vaccination is the most effective method, along with antiviral drugs in selected patients, for preventing influenza infection, it has to be recommended for cancer patients. Implementing vaccination of close contacts of oncology patients would be an additional tool for enhancing protection in fragile patient populations.

Investigation of an imported case of Middle East Respiratory Syndrome Coronavirus (MERS-CoV) infection in Florence, Italy, May to June 2013.

On 31 May 2013, the first case of Middle East Respiratory Syndrome Coronavirus (MERS-CoV) infection in Italy was laboratory confirmed in a previously healthy adult man, who developed pneumonia with moderate respiratory distress after returning from a holiday in Jordan. Two secondary cases were identified through contact tracing, among family members and colleagues who had not previously travelled abroad. Both secondary cases developed mild illness. All three patients recovered fully.

[Primary Burkitt lymphoma of the heart--diagnosis and therapy]. / Primäres Burkitt-Lymphom des Herzens--Diagnose und Therapie.

An 85-year-old patient suffered from progressive deterioration (NYHA III) for several months. Cardiac disease was suspected. Echocardiography as well as a CT scan of the heart revealed a heart tumor to be the cause. Tumor staging was negative. After transvenous biopsy, the diagnosis of a Burkitt lymphoma could be established. Due to the advanced age of the patient, the intended surgical therapy was turned down and the patient was treated with 6 courses of a potentially therapeutic chemotherapy (CHOP scheme), which was well tolerated by the patient. The following CT scan showed a complete remission of the tumor. Six months after chemotherapy the patient is in NYHA stage I.

[Not Available]. / Primäres Burkitt-Lymphom des Herzens - Diagnose und Therapie.

An 85-year-old patient suffered from progressive deterioration (NYHA III) for several months. Cardiac disease was suspected. Echocardiography as well as a CT scan of the heart revealed a heart tumor to be the cause. Tumor staging was negative. After transvenous biopsy, the diagnosis of a Burkitt lymphoma could be established. Due to the advanced age of the patient, the intented surgical therapy was turned down and the patient was treated with 6 courses of a potentially therapeutic chemotherapy (CHOP scheme), which was well tolerated by the patient. The following CT scan showed a complete remission of the tumor. Six months after chemotherapy the patient is in NYHA stage I.

The role of antigen in the localization of naive, acutely activated, and memory CD8(+) T cells to the lung during influenza pneumonia.

The role of Ag in the recruitment and localization of naive, acutely activated, and memory CD8(+) T cells to the lung during influenza infection was explored using TCR-transgenic (Tg) mice. Naive, Thy1.2(+)CD8(+) OT-I TCR-Tg cells were primed and recruited to the lung after transfer into congenic Thy1.1(+) recipients challenged with a genetically engineered influenza virus (influenza A/WSN/33 (WSN)-OVA(I)) containing the K(b) restricted OVA(257-264) epitope (siinfekl) in the viral neuraminidase stalk. However, if the transferred animals were infected with a similar influenza virus that expressed an irrelevant K(b) epitope (WSN-PEPII), no TCR-Tg T cells were detectable in the lung, although they were easily visible in the lymphoid organs. Conversely, there were substantial numbers of OT-I cells found in the lungs of WSN-PEPII-infected mice when the animals had been previously, or were concurrently, infected with a recombinant vaccinia virus expressing OVA. Similar results were obtained with nontransgenic populations of memory CD8(+) T cells reactive to a murine gamma-herpesvirus-68 Ag. Interestingly, the primary host response to the immunodominant influenza nucleoprotein epitope was not affected by the presence of memory or recently activated OT-I T cells. Thus, although Ag is required to activate the T cells, the subsequent localization of T cells to the lung during a virus infection is a property of recently activated and memory T cells and is not necessarily driven by Ag in the lung.

Antitachycardia pacing for rapid VT during ICD charging: a method to prevent ICD shocks.

In patients with ICDs, rapid VTs are usually treated with shocks. It is unknown, if antitachycardia pacing (ATP) delivered once for rapid VT during capacitor charging can avoid painful shocks without increasing the risk of syncope. In patients in whom rapid monomorphic VT (cycle length 300-220 ms) could be reproducibly induced during predischarge ICD testing, the success of cardioversion (defibrillation threshold plus 10 J) and a single ATP attempt (burst with 8 or 16 stimuli) was compared using a randomized crossover study design. Consciousness of the patients was checked by the signal from a button constantly pushed by the patient. In 20 patients (ejection fraction 0.50 +/- 0.19) rapid VTs (253 +/- 26 ms) were reproducibly induced. A single burst successfully terminated 11 (55%) of 20 rapid VTs, 6 episodes could not be terminated with a single burst pacing and 3 VTs accelerated. Rapid VTs not terminated by ATP were significantly faster than those that could be terminated (246 vs 258 ms, P = 0.026). Cardioversion (19 +/- 3 J) terminated the VTs in all cases. No patient suffered syncope during rapid VTs. A single ATP may terminate rapid VT with cycle lengths < 300 ms in 55% of patients without increasing the risk of syncope. Therefore, in rapid VTs one attempt of ATP may be suitable as an additional therapy option during ICD capacitor charging to avoid painful shocks without compromise of safety. Thus, future ICDs should implement the option of ATP during charging of capacitors.

[Incidental findings in cardiology. Diagnosing today what can cause illness tomorrow]. / Zufallsbefunde in der Kardiologie. Heute diagnostizieren, was morgen krank macht.

Among the anatomical and functional findings in cardiology, congenitally corrected transposition of the major vessels (ventricular inversion), bicuspid aortic valve and prolapse of the mitral valve with simultaneous mitral insufficiency have at least a potential for causing future problems. In such cases, regular cardiological checks or a further diagnostic work-up is to be recommended. The assessment of cardiac sounds can usually be correctly interpreted on the basis of the history, physical examination and auscultation. Among the electrocardiological findings, complete left bundle-branch block and prolongation of the QT segment, mandate clarification of a structural heart condition. Furthermore, in the event of ventricular extrasystoles, right-ventricular cardiomyopathy needs to be excluded. Such isolated conduction disorders as left-anterior fascicular block or right bundle-branch block are of no prognostic significance.

Early alterations of the receptor-binding properties of H1, H2, and H3 avian influenza virus hemagglutinins after their introduction into mammals.

Interspecies transmission of influenza A viruses circulating in wild aquatic birds occasionally results in influenza outbreaks in mammals, including humans. To identify early changes in the receptor binding properties of the avian virus hemagglutinin (HA) after interspecies transmission and to determine the amino acid substitutions responsible for these alterations, we studied the HAs of the initial isolates from the human pandemics of 1957 (H2N2) and 1968 (H3N2), the European swine epizootic of 1979 (H1N1), and the seal epizootic of 1992 (H3N3), all of which were caused by the introduction of avian virus HAs into these species. The viruses were assayed for their ability to bind the synthetic sialylglycopolymers 3'SL-PAA and 6'SLN-PAA, which contained, respectively, 3'-sialyllactose (the receptor determinant preferentially recognized by avian influenza viruses) and 6'-sialyl(N-acetyllactosamine) (the receptor determinant for human viruses). Avian and seal viruses bound 6'SLN-PAA very weakly, whereas the earliest available human and swine epidemic viruses bound this polymer with a higher affinity. For the H2 and H3 strains, a single mutation, 226Q-->L, increased binding to 6'SLN-PAA, while among H1 swine viruses, the 190E-->D and 225G-->E mutations in the HA appeared important for the increased affinity of the viruses for 6'SLN-PAA. Amino acid substitutions at positions 190 and 225 with respect to the avian virus consensus sequence are also present in H1 human viruses, including those that circulated in 1918, suggesting that substitutions at these positions are important for the generation of H1 human pandemic strains. These results show that the receptor-binding specificity of the HA is altered early after the transmission of an avian virus to humans and pigs and, therefore, may be a prerequisite for the highly effective replication and spread which characterize epidemic strains.

Ventricular tachycardias above the initially programmed tachycardia detection interval in patients with implantable cardioverter-defibrillators: incidence, prediction and significance.

OBJECTIVES: This retrospective study was performed to provide data on ventricular tachycardias (VT) with a cycle length longer than the initially programmed tachycardia detection interval (TDI) in patients with implantable cardioverter defibrillators (ICDs). BACKGROUND: It has been clinical practice to program a safety margin of 30 to 60 ms between the slowest spontaneous or inducible VT and the TDI. METHODS: Baseline characteristics of 659 consecutive patients with ICDs were prospectively; follow-up information was retrospectively collected. RESULTS: During a mean follow-up of 31+/-23 months, 377 patients (57.2%) had at least one recurrent VT or ventricular fibrillation; 47 patients (7.1%) suffered 61 VTs above the TDI. The risk of a VT above the TDI ranged between 2.7% and 3.5% per year during the first four years after ICD implantation. The difference between the cycle length of the slowest VT before ICD implantation, spontaneous or induced, and the first VT above TDI was 108+/-58 ms. Fifty-four VTs (88.5%) above the TDI were associated with significant clinical symptoms (angina or palpitation 63.9%, heart failure 6.6% and syncope 8.2%). Six patients (9.8%) had to be resuscitated. Kaplan-Meyer analysis identified New York Heart Association class II or III (p = 0.021), ejection fraction < 0.40 (p = 0.027), spontaneous (p<0.001) or inducible (p<0.001) monomorphic VTs and the use of class III antiarrhythmic drugs (amiodarone, p<0.001; sotalol, p = 0.004) as risk predictors of VTs above the TDI. The risk of recurrent VTs above TDI was 11.8%, 12.5% and 26.6% during the first, second and third year after first VT above TDI, respectively. CONCLUSIONS: The risk of VTs above the TDI is significantly increased in some patients, and many VTs above TDI cause significant clinical symptoms. A larger safety margin between spontaneous or inducible VTs and the TDI seems to be necessary in selected patients. This is in conflict with an increased risk of inadequate episodes and demands highly specific and sensitive detection algorithms in these patients.

Balanced hemagglutinin and neuraminidase activities are critical for efficient replication of influenza A virus.

The SD0 mutant of influenza virus A/WSN/33 (WSN), characterized by a 24-amino-acid deletion in the neuraminidase (NA) stalk, does not grow in embryonated chicken eggs because of defective NA function. Continuous passage of SD0 in eggs yielded 10 independent clones that replicated efficiently. Characterization of these egg-adapted viruses showed that five of the viruses contained insertions in the NA gene from the PB1, PB2, or NP gene, in the region linking the transmembrane and catalytic head domains, demonstrating that recombination of influenza viral RNA segments occurs relatively frequently. The other five viruses did not contain insertions in this region but displayed decreased binding affinity toward sialylglycoconjugates, compared with the binding properties of the parental virus. Sequence analysis of one of the latter viruses revealed mutations in the hemagglutinin (HA) gene, at sites in close proximity to the sialic acid receptor-binding pocket. These mutations appear to compensate for reduced NA function due to stalk deletions. Thus, balanced HA-NA functions are necessary for efficient influenza virus replication.

Postexposure vaccination massively increases the prevalence of gamma-herpesvirus-specific CD8+ T cells but confers minimal survival advantage on CD4-deficient mice.

Mice that lack CD4(+) T cells remain clinically normal for more than 60 days after respiratory challenge with the murine gamma-herpesvirus 68 (gammaHV-68), then develop symptoms of a progressive wasting disease. The gammaHV-68-specific CD8(+) T cells that persist in these I-A(b-/-) mice are unable to prevent continued, but relatively low level, virus replication. Postexposure challenge with recombinant vaccinia viruses expressing gammaHV-68 lytic cycle epitopes massively increased the magnitude of the gammaHV-68-specific CD8(+) population detectable by staining with tetrameric complexes of MHC class I glycoprotein + peptide, or by interferon-gamma production subsequent to in vitro restimulation with peptide. The boosting effect was comparable for gammaHV-68-infected I-A(b-/-) and I-A(b+/+) mice within 7 days of challenge, and took more than 110 days to return to prevaccination levels in the I-A(b+/+) controls. Although the life-span of the I-A(b-/-) mice was significantly increased, there was no effect on long-term survival. A further boost with a recombinant influenza A virus failed to improve the situation. Onset of weight loss was associated with a decline in gammaHV-68-specific CD8(+) T cell numbers, though it is not clear whether this was a cause or an effect of the underlying pathology. Even very high levels of virus-specific CD8(+) T cells thus provide only transient protection against the uniformly lethal consequences of gammaHV-68 infection under conditions of CD4(+) T cell deficiency.

Is there a need for routine testing of ICD defibrillation capacity? Results from more than 1000 studies.

AIMS: Benefits and complications of postoperative implantable cardioverter-defibrillator tests are controversial matters. This study sought to assess the necessity of defibrillation function tests after implantation. METHODS AND RESULTS: We retrospectively analysed 1007 implantable cardioverter-defibrillator tests in 587 systems and 556 patients. Nine hundred and thirty implantable cardioverter-defibrillator tests (89.4%) were routinely performed. Seventy-one tests (7%) were performed after a change in the antiarrhythmic drug regimen and six tests (0.60%) because of a suspected dysfunction of the implantable cardioverter-defibrillator. During routine tests, four systems (0.4%) failed to defibrillate the patient. However, in all but one test, abnormalities of the system had been observed before the test. After the addition of antiarrhythmic drugs, two of 71 implantable cardioverter-defibrillator systems (2.8%) failed to defibrillate the patient. One of six systems tested due to a suspected dysfunction failed to defibrillate the patient. During 16 tests (1.6%), complications occurred. CONCLUSIONS: Our experience demonstrates that postoperative tests of the defibrillation function of implantable cardioverter-defibrillators rarely reveal dysfunctions. As testing is unpleasant for the patient and not free of complications, tests might be restricted to those patients in whom a dysfunction is suspected and to those patients in whom class I or class III antiarrhythmic drugs have been added to the antiarrhythmic drug regimen.

Contrast transcranial Doppler ultrasound in the detection of right-to-left shunts: comparison of different procedures and different contrast agents.

BACKGROUND AND PURPOSE: Cardiac right-to-left shunts can be identified by transesophageal echocardiography (TEE) and by transcranial Doppler ultrasound (TCD) with the use of different contrast agents and different provocation procedures. Currently, data on an appropriate time window for the appearance of contrast bubbles in the TCD recording after the injection of the contrast medium and the comparison of different provocation maneuvers to increase right-to-left shunting are insufficient. METHODS: Forty-six patients were investigated by both TEE and bilateral TCD of the middle cerebral artery. The following protocol with 6 injection modes was applied in a randomized way: (1) injection of 10 mL of agitated saline without Valsalva maneuver, (2) injection of 10 mL of agitated saline with Valsalva maneuver, (3) injection of 10 mL of a commercial galactose-based contrast agent (Echovist) without Valsalva maneuver, (4) injection of 10 mL of Echovist with Valsalva maneuver, (5) injection of 10 mL of Echovist with standardized Valsalva maneuver, and (6) injection of 10 mL of Echovist with coughing. RESULTS: In 20 patients, a right-to-left shunt was demonstrated by TEE and contrast TCD (shunt-positive). Sixteen patients were negative in both investigations, no patient was positive on TEE and negative on TCD, and 10 patients were only positive on at least 1 TCD investigation but negative during TEE. The amount of microbubbles detected in the various tests decreased in the following order: Echovist and Valsalva maneuver, Echovist with coughing, Echovist and standardized Valsalva maneuver, saline with Valsalva maneuver, Echovist, and saline. With a time window of 20 to 25 seconds for the bubbles to appear in the TCD recording and with a sequence of first Echovist and Valsalva maneuver and then Echovist with coughing, all shunts were reliably identified with a specificity of 65% compared with TEE as the traditional gold standard. The time of first microbubble appearance was not helpful to distinguish between shunts detected on TEE and other shunts. CONCLUSIONS: TCD performed twice with 2 provocation maneuvers using Echovist is a sensitive method to identify cardiac right-to-left shunts also identified by TEE.

A gamma-herpesvirus sneaks through a CD8(+) T cell response primed to a lytic-phase epitope.

To determine whether established CD8(+) T cell memory to an epitope prominent during the replicative phase of a gamma-herpesvirus infection protects against subsequent challenge, mice were primed with a recombinant vaccinia virus expressing the p56 peptide and then boosted by intranasal exposure to an influenza A virus incorporating p56 in the neuraminidase protein. Clonally expanded populations of functional, p56-specific CD8(+) T cells were present at high frequency in both the lung and the lymphoid tissue 1 month later, immediately before respiratory challenge with gammaHV-68. This prime-and-boost regime led to a massive reduction of productive gammaHV-68 infection in the respiratory tract and, initially, to much lower levels of latency in both the regional lymph nodes and the spleen. The CD8(+) T cell response to another epitope (p79) was diminished, there was less evidence of B cell activation, and the onset of the CD4(+) T cell-dependent splenomegaly was delayed. Within 3-4 weeks of the gammaHV-68 challenge, however, the extent of latent infection in the lymph nodes and spleen was equivalent, and both groups developed the prominent infectious mononucleosis-like syndrome that is characteristic of this infection. The reverse protocol (influenza then vaccinia) seemed to be slightly less effective. Even though immune CD8(+) T cells may be present at the time and site of virus challenge, establishing a high level of CD8(+) T cell memory to lytic-phase epitopes alone does not protect against the longer-term consequences of this gammaHV infection.

Amino acid residues contributing to the substrate specificity of the influenza A virus neuraminidase.

Influenza A viruses possess two glycoprotein spikes on the virion surface: hemagglutinin (HA), which binds to oligosaccharides containing terminal sialic acid, and neuraminidase (NA), which removes terminal sialic acid from oligosaccharides. Hence, the interplay between these receptor-binding and receptor-destroying functions assumes major importance in viral replication. In contrast to the well-characterized role of HA in host range restriction of influenza viruses, there is only limited information on the role of NA substrate specificity in viral replication among different animal species. We therefore investigated the substrate specificities of NA for linkages between N-acetyl sialic acid and galactose (NeuAcalpha2-3Gal and NeuAcalpha2-6Gal) and for different molecular species of sialic acids (N-acetyl and N-glycolyl sialic acids) in influenza A viruses isolated from human, avian, and pig hosts. Substrate specificity assays showed that all viruses had similar specificities for NeuAcalpha2-3Gal, while the activities for NeuAcalpha2-6Gal ranged from marginal, as represented by avian and early N2 human viruses, to high (although only one-third the activity for NeuAcalpha2-3Gal), as represented by swine and more recent N2 human viruses. Using site-specific mutagenesis, we identified in the earliest human virus with a detectable increase in NeuAcalpha2-6Gal specificity a change at position 275 (from isoleucine to valine) that enhanced the specificity for this substrate. Valine at position 275 was maintained in all later human viruses as well as swine viruses. A similar examination of N-glycolylneuraminic acid (NeuGc) specificity showed that avian viruses and most human viruses had low to moderate activity for this substrate, with the exception of most human viruses isolated between 1967 and 1969, whose NeuGc specificity was as high as that of swine viruses. The amino acid at position 431 was found to determine the level of NeuGc specificity of NA: lysine conferred high NeuGc specificity, while proline, glutamine, and glutamic acid were associated with lower NeuGc specificity. Both residues 275 and 431 lie close to the enzymatic active site but are not directly involved in the reaction mechanism. This finding suggests that the adaptation of NA to different substrates occurs by a mechanism of amino acid substitutions that subtly alter the conformation of NA in and around the active site to facilitate the binding of different species of sialic acid.

Efficacy and safety of the initial use of stability and onset criteria in implantable cardioverter defibrillators.

INTRODUCTION: Inappropriate therapies are the most frequent adverse event in patients with implantable cardioverter defibrillators (ICDs). Most ICDs offer a stability criterion to discriminate ventricular tachycardia (VT) from atrial fibrillation and an onset criterion to discriminate VT from sinus tachycardia. The efficacy and safety of these criteria, if used immediately after implantation, is unknown. METHODS AND RESULTS: In a case control study, 87 patients in whom stability and onset criteria had been activated immediately after ICD implantation were matched to 87 patients in whom these criteria had not been activated. The groups were matched for known predictors of inappropriate therapies. With stability and onset criteria off, 24 patients (28%) received inappropriate therapies due to atrial fibrillation (n = 14) or sinus tachycardia (n = 11); with stability and onset on, only 11 patients (13%) were treated by the ICD due to atrial fibrillation (n = 5) or sinus tachycardia (n = 7) (log rank: P = 0.029). Five patients suffered inappropriate therapies despite the fact that onset (n = 4) or stability (n = 1) criteria were not fulfilled once tachycardias continued for a prespecified duration. Only one patient experienced a failure to detect VT due to the onset criterion; none because of stability. CONCLUSION: The immediate use of stability and onset criteria after ICD implantation reduces inappropriate therapies due to atrial fibrillation and sinus tachycardia. Because of the potential for underdetection of VT, this approach should be limited to tachycardia rates hemodynamically tolerated by the patient.

Influence of host species on the evolution of the nonstructural (NS) gene of influenza A viruses.

The matrix (M) and nonstructural (NS) genes of influenza A viruses each encode two overlapping proteins. In the M gene, evolution of one protein affects that of the other. To determine whether or not this evolutionary influence operating between the two M proteins also occurs in the NS gene, we sequenced the NS genes of 36 influenza A viruses isolated from a broad spectrum of animal species (wild and domestic birds, horses, pigs, humans, and sea mammals) and analyzed them phylogenetically, together with other previously published sequences. These analyses enabled us to conclude the following host species-related points that are not found in the other influenza A virus genes and their gene products. (1) The evolution of the two overlapping proteins encoded by the NS gene are lineage-dependent, unlike the M gene where evolutionary constraints on the Ml protein affect the evolution of the M2 protein (Ito et al.. J. Virol. 65 (1991) 5491 5498). (2) The gull-specific lineage contained nonH13 gull viruses and the non-gull avian lineage contained H13 gull viruses, indicating that the gull-specific lineage does not link to the H13 HA subtype in the NS gene unlike findings with other genes. (3) The branching topology of the recent equine lineage (H7N7 viruses isolated after 1973 and H3N8) indicates recent introduction of the NS, M, and PB2 genes into horses from avian sources by genetic reassortment.

Molecular basis for the generation in pigs of influenza A viruses with pandemic potential.

Genetic and biologic observations suggest that pigs may serve as "mixing vessels" for the generation of human-avian influenza A virus reassortants, similar to those responsible for the 1957 and 1968 pandemics. Here we demonstrate a structural basis for this hypothesis. Cell surface receptors for both human and avian influenza viruses were identified in the pig trachea, providing a milieu conducive to viral replication and genetic reassortment. Surprisingly, with continued replication, some avian-like swine viruses acquired the ability to recognize human virus receptors, raising the possibility of their direct transmission to human populations. These findings help to explain the emergence of pandemic influenza viruses and support the need for continued surveillance of swine for viruses carrying avian virus genes.

The role of influenza A virus hemagglutinin residues 226 and 228 in receptor specificity and host range restriction.

Influenza A viruses can be isolated from a variety of animals, but their range of hosts is restricted. For example, human influenza viruses do not replicate in duck intestine, the major replication site of avian viruses in ducks. Although amino acids at positions 226 and 228 of hemagglutinin (HA) of the H3 subtype are known to be important for this host range restriction, the contributions of specific amino acids at these positions to restriction were not known. Here, we address this issue by generating HAs with site-specific mutations of a human virus that contain different amino acid residues at these positions. We also let ducks select replication-competent viruses from a replication-incompetent virus containing a human virus HA by inoculating animals with 10(10.5) 50% egg infectious dose of the latter virus and identified a mutation in the HA. Our results showed that the Ser-to-Gly mutation at position 228, in addition to the Leu-to-Gln mutation at position 226 of the HA of the H3 subtype, is critical for human virus HA to support virus replication in duck intestine.