Auto-and cross-correlation analysis of the electrical activity of muscles

The paper describes apparatus which has been built to measure the auto- and cross-correlation functions. An application of the apparatus, an investigation into the effect on the auto- and cross-correlograms of fatigue, is reported.     

Pseudomyxoma peritonei presenting with inguinal hernia

Pseudomyxoma peritonei (PMP) is rare being characterized by intraperitoneal accumulation of mucinous ascites produced by neoplastic cells, mostly originating from a perforated appendiceal adenoma. The clinical signs of the disease are variable, and preoperative diagnosis is often difficult. We describe the clinical case of a 67-year-old patient referred to our unit one month after a left inguinal hernia repair, presenting clinical signs compliant with PMP. Surgical cytoreduction, peritonectomy, appendectomy, and greater omentectomy with perioperative intraperitoneal chemotherapy were performed. The patient was disease free for a 15 month period when he died apparently due to a cardiac event. We advocate that in all cases of gelatinous fluid in a hernia sac PMP must be suspected, thus histological investigation is mandatory as well as abdominal computed tomography (CT) in order to confirm the diagnosis.     

Comparative activities of oseltamivir and A-322278 in immunocompetent and immunocompromised murine models of influenza virus infection

We developed an immunocompromised murine model of influenza virus infection and demonstrated comparable efficacy of oral oseltamivir and A-322278 (both given at dosages of 10 mg/kg/day) in reducing viral replication, decreasing weight loss, and prolonging survival. Once the treatment was discontinued, severe combined immunodeficient (SCID) mice had progressive viral replication and clinical decline. Drug-resistant variants were detected in 4 (29%) of 14 and 2 (13%) of 15 mice (both BALB/c and SCID) treated with oseltamivir or A-322278, respectively; no resistant variants were detected in placebo-treated mice. Amino acid substitutions in the hemagglutinin receptor-binding site at aa 137 or 225 were detected in cloned resistant isolates. A substitution in the neuraminidase (NA) active site (Arg292Lys) was detected in the cloned virus recovered from an oseltamivir-treated mouse. This model would be useful for elucidation of the molecular mechanisms of resistance to NA inhibitors and for testing of anti-influenza therapy options that might prevent the emergence of resistant variants.     

Design of a polymer drug for serological diagnosis of hepatitis C

A new immunobiological polymer drug has been designed for the serological identification of hepatitis C. The drug is able to reveal specific antibodies in the sera of patients with hepatitis C, meets the current requirements of diagnostic test systems, and shows a high sensitivity and specificity. It is based on polyacroleinic microspheres; the concentrated cell culture biomass of hepatitis C virus (HCV), which contains an adequate set of viral antigens, is used as sensitin. A new diagnosticum is proposed to be used during primary (screening) laboratory studies based on the serological detection of total antibodies to HCV antigens in the volume agglomeration test. The latter is both one of the alternative methods during serological studies and an additional procedure when a set of diagnostic techniques is used.     

Role of galectin-3 in acetaminophen-induced hepatotoxicity and inflammatory mediator production

Galectin-3 (Gal-3) is a β-galactoside-binding lectin implicated in the regulation of macrophage activation and inflammatory mediator production. In the present studies, we analyzed the role of Gal-3 in liver inflammation and injury induced by acetaminophen (APAP). Treatment of wild-type (WT) mice with APAP (300 mg/kg, ip) resulted in centrilobular hepatic necrosis and increases in serum transaminases. This was associated with increased hepatic expression of Gal-3 messenger RNA and protein. Immunohistochemical analysis showed that Gal-3 was predominantly expressed by mononuclear cells infiltrating into necrotic areas. APAP-induced hepatotoxicity was reduced in Gal-3-deficient mice. This was most pronounced at 48-72 h post-APAP and correlated with decreases in APAP-induced expression of 24p3, a marker of inflammation and oxidative stress. These effects were not due to alterations in APAP metabolism or hepatic glutathione levels. The proinflammatory proteins, inducible nitric oxide synthase (iNOS), interleukin (IL)-1β, macrophage inflammatory protein (MIP)-2, matrix metalloproteinase (MMP)-9, and MIP-3α, as well as the Gal-3 receptor (CD98), were upregulated in livers of WT mice after APAP intoxication. Loss of Gal-3 resulted in a significant reduction in expression of iNOS, MMP-9, MIP-3α, and CD98, with no effects on IL-1β. Whereas APAP-induced increases in MIP-2 were augmented at 6 h in Gal-3(-/-) mice when compared with WT mice, at 48 and 72 h, they were suppressed. Tumor necrosis factor receptor-1 (TNFR1) was also upregulated after APAP, a response dependent on Gal-3. Moreover, exaggerated APAP hepatotoxicity in mice lacking TNFR1 was associated with increased Gal-3 expression. These data demonstrate that Gal-3 is important in promoting inflammation and injury in the liver following APAP intoxication.     

Neuraminidase inhibitor susceptibility testing in human influenza viruses: a laboratory surveillance perspective

Neuraminidase inhibitors (NAIs) are vital in managing seasonal and pandemic influenza infections. NAI susceptibilities of virus isolates (n = 5540) collected during the 2008-2009 influenza season were assessed in the chemiluminescent neuraminidase inhibition (NI) assay. Box-and-whisker plot analyses of log-transformed IC(50)s were performed for each virus type/subtype and NAI to identify outliers which were characterized based on a statistical cutoff of IC(50) >3 interquartile ranges (IQR) from the 75(th) percentile. Among 1533 seasonal H1N1 viruses tested, 1431 (93.3%) were outliers for oseltamivir; they all harbored the H275Y mutation in the neuraminidase (NA) and were reported as oseltamivir-resistant. Only 15 (0.7%) of pandemic 2009 H1N1 viruses tested (n = 2259) were resistant to oseltamivir. All influenza A(H3N2) (n = 834) and B (n = 914) viruses were sensitive to oseltamivir, except for one A(H3N2) and one B virus, with D151V and D197E (D198E in N2 numbering) mutations in the NA, respectively. All viruses tested were sensitive to zanamivir, except for six seasonal A(H1N1) and several A(H3N2) outliers (n = 22) which exhibited cell culture induced mutations at residue D151 of the NA. A subset of viruses (n = 1058) tested for peramivir were sensitive to the drug, with exception of H275Y variants that exhibited reduced susceptibility to this NAI. This study summarizes baseline susceptibility patterns of seasonal and pandemic influenza viruses, and seeks to contribute towards criteria for defining NAI resistance.     

In Vitro Antiviral Activity of Favipiravir (T-705) against Drug-Resistant Influenza and 2009 A(H1N1) Viruses

Favipiravir (T-705) has previously been shown to have a potent antiviral effect against influenza virus and some other RNA viruses in both cell culture and in animal models. Currently, favipiravir is undergoing clinical evaluation for the treatment of influenza A and B virus infections. In this study, favipiravir was evaluated in vitro for its ability to inhibit the replication of a representative panel of seasonal influenza viruses, the 2009 A(H1N1) strains, and animal viruses with pandemic (pdm) potential (swine triple reassortants, H2N2, H4N2, avian H7N2, and avian H5N1), including viruses which are resistant to the currently licensed anti-influenza drugs. All viruses were tested in a plaque reduction assay with MDCK cells, and a subset was also tested in both yield reduction and focus inhibition (FI) assays. For the majority of viruses tested, favipiravir significantly inhibited plaque formation at 3.2 μM (0.5 μg/ml) (50% effective concentrations [EC₅₀s] of 0.19 to 22.48 μM and 0.03 to 3.53 μg/ml), and for all viruses, with the exception of a single dually resistant 2009 A(H1N1) virus, complete inhibition of plaque formation was seen at 3.2 μM (0.5 μg/ml). Due to the 2009 pandemic and increased drug resistance in circulating seasonal influenza viruses, there is an urgent need for new drugs which target influenza. This study demonstrates that favipiravir inhibits in vitro replication of a wide range of influenza viruses, including those resistant to currently available drugs.In the United States alone, seasonal influenza is responsible annually for infecting between 5 and 20% of the American population, resulting in more than 200,000 hospitalizations and 36,000 deaths (8). Globally, seasonal influenza causes between 250,000 and 500,000 deaths every year (60). Influenza is not only a disease of great medical importance but also of economic importance. Despite available vaccines, a recent study predicted that in the United States influenza results in direct medical costs of the order of $10.4 billion each year, with the total economic burden for the United States being projected at $87.1 billion each year (44). It is widely accepted that vaccination remains the most effective approach for the prevention of viral infections (48). Although there is a safe and effective annual trivalent influenza vaccine, a large proportion of the global population does not receive the yearly influenza vaccine. This can be due to a variety of reasons, including the lack of access to adequate health care, unavailability of vaccine supply, allergies, and adverse reactions. During the 2009 pandemic (pdm), in addition to the vaccination and epidemiological control measures being exerted by health care officials, antivirals targeting influenza offer an essential tool in treating infected patients, in addition to protecting those at high risk of infection, such as the young, elderly, and health care workers.Currently, there are two classes of anti-influenza drugs licensed in the United States for use in the treatment and management of influenza infections in humans: M2 ion channel blockers (also known as adamantanes) and neuraminidase (NA) inhibitors (NAIs) (30). Influenza antivirals are highly effective in the treatment of influenza infections if used promptly following the onset of symptoms or following exposure (45, 46). Both the M2 blockers amantadine and rimantadine are taken by the patient orally (45). However, of the two available NAIs, only oseltamivir is available as an oral formulation (zanamivir has to be inhaled [14, 53]), although other routes of administration have been investigated (31). The use of the M2 blockers amantadine and rimantadine is limited due to the rapid emergence of transmissible drug-resistant mutant viruses and the fact that they offer protection only against influenza A virus infections (32). The high prevalence of adamantane resistance in seasonal A(H3N2) viruses and oseltamivir resistance in seasonal A(H1N1) viruses is reflected in the CDC recommendations for the use of influenza antivirals (6).The majority of adamantane-resistant A(H3N2) and A(H1N1) viruses circulating globally in recent years share the same mutation, S31N, in the M2 protein (20), although other resistance-conferring mutations have been detected also (including A30T, L26F, and V27A) (20, 49). The globally spread oseltamivir-resistant seasonal A(H1N1) viruses share the same mutation, H275Y (H274Y in N2 subtype amino acid numbering), in the drug-targeted enzyme neuraminidase, although other mutations are known to cause reduced susceptibility in vitro (19, 47, 50).Seasonal A(H1N1) viruses resistant to both the adamantanes and the NAI oseltamivir have previously been reported, without an apparent link to treatment (12, 50). Currently, zanamivir is the only drug effective against both adamantane-resistant and/or oseltamivir-resistant influenza viruses, but due to the fact that it has to be inhaled, it is less suitable for use with several high-risk groups, including the severely ill (41), infants (33), and the elderly (22). Furthermore, zanamivir may decrease pulmonary function, so it is not recommended for the treatment of infections in individuals with chronic underlying lung and heart disease conditions (23).Since 1997, there have been several outbreaks of highly pathogenic avian influenza A(H5N1) infections in poultry, with a substantial number of infections occurring in humans (1). The overall case fatality of A(H5N1) infections in humans is over 60% and, unlike seasonal influenza, is most deadly in the young and healthy (ages 10 to 19 years) (59). Oseltamivir is the medication of choice for treating individuals infected with A(H5N1) (17). However, resistance in A(H5N1) viruses has been detected following the treatment of patients with oseltamivir (18, 38). In addition, naturally occurring reduced susceptibility to oseltamivir (35, 40) and possibly to zanamivir (29) has been documented for circulating A(H5N1) viruses, including novel mutations in the NA (29, 35). Adamantane resistance is widely spread among A(H5N1) viruses that carry mutations at amino acid residues 26, 27, and 31 in the M2 protein (13, 35) and among swine viruses circulating in Eurasia (27).In April 2009, a novel reassortant A(H1N1) virus was first identified as circulating in humans in both Mexico and the United States (7, 9). Since April, the virus has continued to transmit among humans, and on 11 June 2009 the World Health Organization classified the outbreak as the first influenza pandemic of the 21st century (58). The 2009 A(H1N1) pandemic viruses consist of a unique combination of gene segments, including those of the North American (triple reassortants) and Eurasian swine lineages (27, 54). The 2009 A(H1N1) pandemic viruses are resistant to the adamantanes and sensitive to the NAIs (3, 16). Yet, concerns exist about the possibility of acquisition of resistance to the NAI oseltamivir, since the majority of A(H1N1) viruses which have been circulating predominantly worldwide during the 2008-2009 influenza season are oseltamivir resistant due to the resistance-conferring H275Y mutation in the NA. Such an acquisition of resistance by the 2009 A(H1N1) pandemic viruses would be a major setback and would further limit the already sparse therapeutic options (15, 57). There have been laboratory-confirmed cases of oseltamivir-resistant 2009 A(H1N1) pandemic viruses (each carrying the H275Y resistance-conferring mutation in the NA) in the United States (5).Collectively, these recent findings emphasize not only the need for new effective antivirals to control and treat influenza infections but also the need to identify new molecular targets (47).One such compound which is currently being investigated and undergoing clinical trials for the treatment of influenza infections is favipiravir (T-705), a pyrazine derivative (2, 26, 31). Favipiravir targets the RNA-dependent RNA polymerase (RdRp), a component of influenza virus different from that of currently licensed influenza antivirals (24, 25). It was shown that favipiravir can inhibit the viral replication of influenza type A, B, and C viruses (24, 25, 55). Favipiravir reduces influenza virus replication by selectively inhibiting the viral RdRp, since it does not affect the synthesis of host cellular DNA and RNA (25). Favipiravir has also shown great potential to act as a broad-spectrum antiviral against many RNA viruses, as reviewed by Furuta and coworkers (26).The purpose of this study was to evaluate the ability, in vitro, of favipiravir to inhibit the viral replication of contemporary influenza viruses as well as viruses with pandemic potential, including viruses resistant to the currently available and licensed anti-influenza drugs. In this report we demonstrate that favipiravir is a potent inhibitor of seasonal influenza A and B virus replication, including that of drug-resistant and drug-sensitive viruses. In addition, favipiravir was shown to effectively inhibit influenza A viruses of other antigenic subtypes, including A(H2N2), viruses of avian origin [A(H4N2), A(H7N2), and A(H5N1)], and viruses of swine origin [A(H1N1) and A(H1N2)], as well as the 2009 A(H1N1) pandemic viruses.