[Seroepidemiology and molecular epidemiology of enterovirus type 71 in the world and the Russian Federation].

A review of recent publications on epidemiology and seroepidemiology of enterovirus type 71 in various regions of the world and authors' own results of study of seroepidemiology and molecular epidemiology of EV71 in Russia are presented.

[Encephalomyelitis caused by enterovirus type 71 in children].

Enterovirus type 71 (EV71) is a causative agent of large outbreaks of hand, foot, and mouth disease (HFMD) in Europe (Bulgaria, 1975; Hungary, 1978) and South-East Asia (Malaysia, 1977; Taiwan, 1998; Singapore, 2000-2007; People's Republic of China, 2007-2009). HFMD afflicted children less than 10 years of age and resulted in recovery within 3-7 days. In a small percentage of infants (aged 6 months to 3 years), HFMD was accompanied by acute neurological complications, such as serous meningitis, poliomyelitis-like syndrome (extremity pareses and muscle paralyses); brain stem encephalitis (myoclonic jerks, tremor, lethargy, swallowing and speech disorders, cardiopulmonary failure, pulmonary edema, shock, coma, death). X-ray study revealed pulmonary hemorrhages and edema. Mortality rates were as high as 82-94% in severe cases. Incapacitating motor, respiratory, and psychoemotional disorders persisted in some surviving children. Pathomorphologically, patients with central nervous system disease and cardiopulmonary failure were found to have acute inflammation of the grey matter of the brain stem (medulla oblongata, pons) and spinal cord. Inflammatory changes in the lung and myocardial tissues were negligible or absent. Fatal pulmonary edema was neurogenic in origin and resulted from damage to the respiratory and vasomotor centers of the brain stem.

[Enterovirus 71: epidemiology and diagnostics].

In 1970s enterovirus type 71 (EV71) caused several epidemics of poliomyelitis-like disease with severe neurologic sequelae. In the last 20 years EV71 was the cause of series of outbreaks and epidemics of foot-and-mouth disease-like conditions with neurologic sequelae in countries of South-East Asia. During the last epidemic of EV71 infection, which occurred in China in 2008, more than 60,000 cases was registered, 38 of which were lethal. Some aspects of epidemiology and laboratory diagnostics of disease caused by EV71 are considered in this review.

[Acute enterovirus uveitis in infants].

Enterovirus uveitis (EU) is a new infant eye disease that was first detected and identified in Russia in 1980-1981. Three subtypes of human echoviruses (EV19K, EV11A, and EV11/B) caused 5 nosocomial outbreaks of EU in different Siberian cities and towns in 1980-1989, by affecting more than 750 children mainly below one year of age. Sporadic and focal EU cases (more than 200) were also retrospectively diagnosed in other regions of Russia and in different countries of the former Soviet Union. There were following clinical manifestations: common symptoms of the infection; acute uveitis (rapid focal iridic destruction, pupillary deformities, formation of membranes in the anterior chamber of the eye); and in 15-30% of cases severe complications, cataract, glaucoma, vision impairments. Uveitis strains EV19 and EV11 caused significant uveitis in primates after inoculation into the anterior chamber of the eye, as well as sepsis-like fatal disease with liver necrosis after venous infection. The uveitis strains are phylogenetically and pathogenetically close for primates to strains EV19 and EV11 isolated from young children with sepsis-like disease. The contents of this review have been published in the Reviews in Medical Virology, 2004, vol. 14, p. 241-254.

Recombination in uveitis-causing enterovirus strains.

The complete nucleotide sequences of three human echovirus (EV) 11 strains and one EV19 strain, all of which caused outbreaks of enterovirus uveitis (EU), a new infant disease first identified in 1980 in Siberia, were determined. One EV11 strain which caused an outbreak of sepsis-like disease in Hungary was also sequenced. All four EV11 strains were mosaic recombinants of the prototype EV11 strain Gregory, with their non-structural coding regions and 5' NTRs being more similar to other prototype enteroviruses (EV1, EV9). However, this finding is probably a feature of all circulating enterovirus strains and may not be related to their altered virulence. A full genome sequence comparison of the three subtypes of EU-causing strains excludes the role of recent recombination in their emergence, and points to their independent emergence.

Molecular epidemiology of enteroviruses causing uveitis and multisystem hemorrhagic disease of infants.

We studied molecular epidemiology of highly virulent echovirus 11 and 19 strains that were isolated during five outbreaks of enterovirus uveitis (EU) in Siberia in 1980-1989, and three outbreaks of multisystem hemorrhagic disease of infants (MHD) in 1988-1991. Three genome regions, 5'NTR, VP1-2A junction, and a fragment of 3D polymerase, were analyzed. Phylogenetic grouping in the VP1-2A region correlated with serotyping results. All studied EV11 and EV19 strains, including the prototype EV11 and EV19, formed a major phylogenetic group in VP1-2A region. Within that group, several EV11 isolates from EU and MHD outbreaks formed a distinct cluster in VP1-2A and 5' NTR genome regions, designated EV11/B. All strains of this cluster possessed high virulence for monkeys compared with the prototype echoviruses. Subgrouping within this cluster correlated with year of virus isolation, not with the disease the viruses caused in infants (EU or MHD).

Phylogenetic and serological characterization of echovirus 11 and echovirus 19 strains causing uveitis.

The strains of echovirus 19 (EV 19) and echovirus 11 (EV11), isolated from infants with similar clinical symptoms of acute enterovirus uveitis (EU) in Russia (Siberia) in 1980-1989, were investigated phylogenetically (nucleotide sequence of a 300 nt fragment in 5' NTR and VP4 junction) and serologically. The result confirmed that viruses belong to the Enterovirus genus, with 58-80% nt sequence homology with previously sequenced enteroviruses, and showed the genetical identity between the strains isolated during each of five outbreaks of the EU. The results also demonstrated that isolates from the last three outbreaks of EU belong to the same phylogenetic group despite the remarkable spatial and temporal distance between the outbreaks. The results confirm the role of these echoviruses in the etiology of the EU. Based on phylogenetic and serological comparisons the studied strains were divided into three distinct groups: group I, EV19/K (Krasnoyarsk, 1980-1981), group II, EV11/A (Krasnoyarsk, 1982), group III, EV11/B (Krasnoyarsk, 1986; Omsk, 1987-1988; Irkutsk, 1989). Minor details of the epidemiology of the outbreaks were also revealed.

[Enteroviral persistence in eye infections in monkeys (postuveitis syndrome)]. / Enterovirusnaia persistentnaia infektsiia glaza u obez'ian (postuveitnyi sindrom).

Follow-up (5.5 years) of monkeys with a history of acute enterovirus uveitis (AEU) after intraocular infection with ophthalmotropic ECHO 11 and ECHO 19 strains showed progressive changes in the infected eye; destruction of the iris, formation of films in the anterior chamber of the eye, deformation of the pupil, and signs of cataract and glaucoma. A short (1-30 days) stage of active reproduction of infective virus (up to 10(10) TCD50/g tissue in the first days postinfection) was followed by the second very long stage of limited specific multiplication of virus in ocular tissues. The virus was not detected during the second stage, but its components (virusspecific antigen and crystalloid accumulations of virions) were found in ophthalmic and conjunctival tissues; moreover, high levels of neutralizing antibodies were detected in the blood. The results indicate AEU transformed into a stubborn persistent infection of the eye in monkeys. The findings of this follow-up and published reports about many-year follow-up of children with AEU permit us to classify a post-uveitis disease as a special nosological form of a persistent enterovirus infection and denote it as enterovirus post-uveitis syndrome. The main signs of this syndrome are a history of AEU, progressive destructive changes in the anterior segment of the eye, presence of virus components in ocular tissues in remote periods after infection, and high antibody level in the blood.

[Persistence of ophthalmotropic enteroviruses in ocular tissues after enterovirus uveitis]. / Persistentsiia oftal'motropnykh 'enterovirusov v tkaniakh glaza posle perenesennogo 'eksperimental'nogo 'enterovirusnogo uveita.

Thirteen monkeys were infected with ophthalmotropic strains of ECHO11 and ECHO19 in the anterior chamber of one eye. Echovirus antigens were detected by direct immunofluorescence in the membranes of the infected and paired eyes in various periods after inoculation (from 3 days to 5 years 8 months). Virus antigen was detected both in the infected and intact eyes. During the acute period of uveitis the antigen was detected mainly in the anterior segment of the infected eye, whereas in remote period it was found mainly in the posterior segment. Long persistence of the virus antigen in various ocular membranes may explain many of the late complications observed in children with enterovirus uveitis.

[Clinical and experimental parallels in children with enterovirus uveitis]. / Kliniko-éksperimental'nye paralleli pri énterovirusnykh uveitakh u detei.

Clinical features of acute and remote periods of experimental enterovirus uveitis were studied. The disease was induced in 22 monkeys by inoculation of ECHO 11 and ECHO 19 strains in the anterior chamber. The animals were followed up for 3 months to 5 years 8 months. The main clinical manifestations of the acute and remote periods of experimental disease were similar to its clinical presentation in children. The developed experimental model can be used to study the pathogenesis of complications of enterovirus uveitis.

[Superacute lethal liver necrosis in monkeys infected with highly pathogenic variants of enteroviruses (ECHO 11 and ECHO 19 viruses)]. / Sverkhostryi letal'nyi nekroz pecheni u obez'ian, infitsirovannykh vysokopatogennymi variantami énterovirusov (virusy ECHO 11 i ECHO 19).

The authors studied the phenomenon of superhigh virulence for monkeys of viruses ECHO 11 (E11) and ECHO 19 (E19) isolated in 1981-1991 in Russia and Tadzhikistan from children with acute enterovirus uveitis or severe enterovirus infection. In 21 morphologically examined animals after coma, macrofocal or total necrosis of hepatocytes in weak inflammation was seen as early as experiment day 1-6. Hepatic lesion interpreted as hepatosis underlies acute hepatic failure with lethal outcome. The most intensive reproduction of viruses E11 and E19 occurred in the liver. Changes in the kidneys, lungs, spleen, adrenals, CNS detected in many animals aggravated the disease, but were not the primary cause of death. The disease in monkey was similar by the main criteria to superacute lethal diseases (acute hepatic failure against massive hepatic necrosis, hemorrhagic syndrome) registered in outbreaks of E11 and E19-virus infection in children in the USA, Great Britain, Israel, Russia and Ukraine. Monkeys can be used as man-adequate model for study of enterovirus superacute lethal necrosis of the liver.

[Detection of poliomyelitis viral strains in natural isolates and identification of them by polymerase chain reaction]. / Obnaruzhenie shtammov virusa poliomielita v prirodnykh izoliatakh i ikh identifikatsiia s pomoshch'iu polimeraznoi tsepnoi reaktsii.

One hundred and fifty nucleotide-long VP1/2A junction regions were sequenced in the RNAs of 19 strains isolated in 1990-1991 from patients with paralytic poliomyelitis in different regions of the former USSR. On the basis of the alignments of these sequenced RNAs, four pairs of 19-25 base-long oligodeoxynucleotide PCR primers were designed capable of detecting polio RNAs in isolated strains and of discriminating between polio genotypes. PCR with 520 polio virus strains isolated from patients, normal subjects, and environmental objects showed 428 of these strains to be related to Sabin's vaccine strains, whereas the rest were referred to A (30), T (24), and G (1) genotypes of serotype 1 and to C-genotype (37) of serotype 3. The designed primers were highly specific and did not cross-react between themselves and with primers specific for Sabin's vaccine strains in PCR.

[An outbreak of poliomyelitis in Tajikistan in 1991 caused by the poliomyelitis virus type 1 in concomitant echovirus 19 infection]. / Vspyshka poliomielita v Tadzhikistane v 1991 g., vyzvannaia virusom poliomielita tipa 1 pri soputstvuiushchei infektsii virusom ECHO 19.

In 1991 in Tajikistan 36 cases of acute poliomyelitis were recorded (spinal form 27, bulbospinal 3, pontospinal 6 cases) in children ranging in ages from 6 months to 6 years. The outcome of the disease was fatal in 4 cases and with crude residual effects in 32 children. Out of 23 children examined, poliomyelitis virus type I was isolated from 13, antibodies to poliovirus type I in titres 1:16 to 1:1024 were detected in 21 children, ECHO 19 virus was isolated from 5 children and antibodies to this virus in titres 1:16 to 1:8192 were demonstrated in 13 children. The fresh isolates of poliomyelitis virus type I had rct+, DS+ and N+ markers. The strains of ECHO 19 virus were highly pathogenic for monkeys causing deaths of the animals within 3-4 days. It is assumed that the main causative agent of the Tajikistan outbreak of poliomyelitis was a virulent "wild" variant of poliovirus type I. The preceding or concomitant infection with ECHO 19 virus (considering the data from the literature and the demonstrated high virulence of this agent for monkeys) could exert an aggravating effect on the course of poliomyelitis or, in some cases, be an independent cause of the disease and, possibly, death of the children.

[Uveitis, caused by Echo-11 virus]. / Uveit, vyzvannyi virusom Echo-11.

Analysis of clinical manifestations of a total enteroviral infection with ECHO-11 virus-induced uveitis in 58 infants aged 2 weeks to 1.5 years has shown that uveitis develops on days 1-10 of the disease and is associated with a weak injection, endothelial edema, hyperemia and edema of the iris rapidly eventuating in destruction of the pigment lamina and fenestration. Posterior synechiae, pupil deformations, grave uveitis with hypotonia of 4-10 mm Hg are rapidly developing. A two-year follow-up has shown marked hemo- and hydrodynamic disorders, retarded growth of the involved eye, development of grave complications in 60 percent of patients. The most characteristic late complications were iridal fenestration, pseudocoloboma-type pupil deformation, formation of prelental films, often pigmented. The condition is associated with active production of virus-neutralizing antibodies in titers of 1:4 to 1:16384. No relationship between the clinical course and antibody titers was revealed.

[The CNS lesion in monkeys spinally infected with the vaccinal poliomyelitis virus]. / Porazhenie TsNS obez'ian pri spinal'nom zarazhenii vaktsinnym virusom poliomielita.

Specific features of the CNS lesions in monkeys inoculated with a vaccine poliovirus strain were studied in comparison with those developing after inoculation with virulent strains. The lesions in the lumbar cord and other regions of the brain and spinal cord after intraspinal inoculation of the vaccine virus are described at length. Certain signs reflecting the specific nature of the lesions were observed. The results of histopathological and immunofluorescent studies were shown to be important for differentiation of specific and nonspecific lesions.

[Role of enterovirus infection in the etiology of uveitis in children]. / Rol' énterovirusnoi infektsii v proiskhozhdenii uveitov u detei.

A total of 224 cases with endogenic uveitis in infants are analyzed. The results of complex clinical and immunological analysis and serologic findings bring the authors to a conclusion that enteroviral uveitis makes up 42 percent of all uveitis in children aged under 3 and that this condition is prevalent in this country. Typical late complications, long persistence of virus-neutralizing antibodies in high titers (up to 8-10 years after convalescence) permit a retrospective etiological diagnosis.

[The geographical spread of enterovirus uveitis in the USSR]. / Geograficheskoe rasprostranenie énteroviusnogo uveita v SSSR.

Enteroviral uveitis is accompanied by intensive and long-term production of specific neutralizing antibodies. The antibody titer to the causative virus ECHO 19/K (Siberia, Krasnoyarsk, 1980-1981) was 1:1000-1:65,000 in 80% of children 5 to 6 1/2 years after the illness. In retrospective examinations of blood sera from children with the history of uveitis using a neutralization test with ophthalmotropic strains of ECHO 19 and ECHO 11 viruses, the ECHO-virus etiology of this disease was first established in 80 patients living in the European part of the country (Moscow, Leningrad, Volgograd, Krasnodar, Donetsk, etc) and in the Caucasus (Yerevan, Tbilisi, Baku). The epidemic process in uveitis is associated with the emergence and alternation of antigenic ECHO virus variants. Both in the Asian and European USSR the causative agent of uveitis in 1979-1983 was ECHO 19/K, in 1980-1987 ECHO 11/A, and in 1986-1989 ECHO 11/B virus. In 1979-1989, altogether over 800 cases of uveitis in children caused by ophthalmotropic variants of ECHO 19 and ECHO 11 viruses were detected in the Soviet Union.

[Outbreak of enteroviral uveitis in children in Omsk in 1987-1988]. / Vspyshka énterovirusnogo uveita u detei v Omske v 1987-1988 gg.

From May, 1987, to July, 1988, 56 cases of acute enterovirus infection complicated by uveitis were reported in Omsk, Western Siberia, USSR. Infants aged from 15 days to 19 months were involved. The infection was nosocomial, the peak of the incidence occurring in January 1988. Severe and moderately severe cases of uveitis (43 out of 56 infants) were prevalent running a course with destruction of the iris, formation of posterior synechias, deformation of the ciliary body, lenticular opacity, appearance of prelental films, retardation of the development of the affected eye, hemorrhages in the fundus of the eye. The causative agent of uveitis outbreak was ECHO-11 virus: 40 strains of ECHO-11 virus were isolated from 27 out of 33 examined infants with uveitis; in 32 out of 33 infants a high titre of antibodies (1:256-1:16384) to the endemic ECHO-11 virus was demonstrated. Uveitis was reproduced in monkeys infected with Omsk strains of ECHO-11 virus. The uveitis outbreak in Omsk is similar in its etiology, epidemiology, and clinical pattern to three previous outbreaks of enterovirus uveitis observed in Krasnoyarsk (central Siberia) observed in 1980-1986.

[The determination of free hemoglobin in blood plasma using the hemiglobincyanide method]. / Opredelenie svobodnogo gemoglobina plazmy krovi gemiglobintsianidnym metodom.

The authors have proved the possibility of using the hemiglobincyanide method for measurements of blood plasma free hemoglobin: free hemoglobin was measured in autoblood reinfused from the operation field by various methods (45-benzidine method, 3 parallel analyses in every study, and 45-hemiglobincyanide method, modified by increasing the amount of the tested material from 0.02 to 2.0 ml, also 3 parallel analyses per sample); a correction coefficient equal to 71.7 is suggested, and correction of the data according to a formula.