‘Antigens of Influenza: Hemagglutinin (HA): Cause hemagglutination Neuraminidase (NA): causes reversal of hemagglutination called Elution CHAPTER 4.3 IN OOUICUS ae Ue MUL eC] (-) Properties: Orthomyxoviridae Paramyxoviridae Size 80-120 nm 100-800 nm Shape Spherical; Rarely flamentous | Pleomorphic Nucleic acid Negative sense ssRNA, Negative sense ssRNA ‘Segmented; eight pieces Unsegmented: single piece Genetic recombination | Seen Not seen ‘Antigenic variation | Seen Not seen Site for RNA Replication | Nucleus ‘Gytoplasm Important human Influenza virus Prainfluenza virus, Mumps virus pathogens Respiratory syncytial vis Measles virus, Metapneumovirus ‘Antigens: Both HA and NA spikes present, HA spike present in: Parainfluenza, Hemagglutinin (HA) and | So hemagglutination is reversible | Mumps, Measles, Neuraminidase (NA) | (Elution seen) NA spike present in: Parainfuenza, Mumps. Inclusion body: intracytoplasmic {only for measles, itis both intracytoplasmic and intranuclear) ORTHOMYXOVIRUSES Naar General Properties + Influenza virus is spherical and possesses helical symmetry + Viral RNA comprises of eight segments of negative sense single stranded RNA. * Site of RNA replication: In the nucleus (in contrast to cytoplasm. by most other RNA viruses) * Viral proteins: Influenza virus contains eight structural proteins (PB1, PB2, PA, NP, HA, NA, M1 and M2) and two nonstructural proteins (NS1 and NS2). Antigens and Typing Influenza possess two glycoprotein antigens inserted into the lipid envelope: HA (Hemagglutinin) and NA (Neuraminidase) + Hemagglutinin (HA): It binds to mucin or sialic acid receptors on RBCS, resulting in clumping of RBCs to cause hemagglutination. It also binds to the same receptors on host, cells, thus facilitating viral entry. Antibody to HA is protective in nature. + Neuraminidase (NA): It is present in fewer number. It is a sialidase enzyme that degrades the sialic acid receptors on RBCs; thus helps in: © It displaces HA from RBCs resulting in reversal of hemagglutination called elution. © It facilitates release of virus particles from infected cell surfaces. © NAhelps the virus to pass through the mucin layer in the respiratory tract. Scanned with CamScannerTyping + Three Genera: Based on Nucleoprotein (NP) and Matrix (M) proteins: © Influenza: A (MC cause of outbreak/epidemic, only cause of pandemic) © Influenza: B (rarely seen) and Influenza C (not circulating now) Influenza Typing: +" nluenza’A (MC cause of + Subtypes © Based on HA and NA antigens, Influenza A has distinct 16 H subtypes (F11 to H16) | _ ottbreatlepemic, only cause and 9 N subtypes (NI-N9). Influenza: B (rarely seen) © Influenza B and C viruses though have subtypes; but are not designated. Influenza: C (not circulating now). Antigenic Variation Influenza virus type A and to less extent type B undergo antigenic variation, which is of two types: antigenic shift and drift. Type C influenza virus is stable. ‘Occur in Type A Occur in type A, to less extent type B Results in Pandemic and major epidemic Results in periodic Epidemic and sporadic cases Due to genetic reassortment ‘Due fo point mutation Occurs every 10-20 years Occurs every 2-3 years Currently circulating strains: + Type A/HINI (2009 Pandemic flu strain) a + Type A/H3N2 (Seasonal flu strain) Sree ae mee + Type A/HBNI (Avian flu strains) e Type AHIN1 (2009 Pandemic + TypeB fu strain) ‘Type AH3N2 (Seasonal fu Clinical Features een ean ca + Incubation period: 18-72 hours strains) * Reservoir: Animals, birds and humans Type B. + MC manifestation: Mild flue like illness (URTI) + MC complication: © Bacterial pneumonia (S. aureus > pneumococci and H. influenzae) is more common than viral pneumonia © Reye's syndrome: Common with Type B (fatty liver following aspirin intake), Laboratory Diagnosis ‘Specimen collection: Nasopharyngeal swab (polyester or Dacron swabs). Transported in viral transport media, kept at 40°C up to 4 days, thereafter at 700°C. Isolation of virus: Embryonated eggs and primary monkey kidney cell lines. Egg inoculation © Amniotic cavity inoculation: It supports growth of Influenza A, B, C © Allantoic cavity inoculation: Supports growth of only Influenza A © Growth is detected by: Hemagglutination with Fowl and Guinea pig RBC © Type A: Agglutinates with Guinea pig RBC, Type C: Agglutinates with fowl RBC at 4°C; and Type B: Agglutinates with both. + Antigen detection from nasopharyngeal cells by direct IF test. Influenza Serology: + Neutralization test: It isthe + Antibody detection: Fourfold rise in the antibody titer is more significant: ‘most specific andthe best © Hemagglutination inhibition test (HAI) eae © Neutralization test It is the most specific and the best predictor of susceptibility to | {2mtuming and difical to infection, but is time-consuming and difficult to perform. perform © ELISA is more sensitive than other assays. +_ ELISA: Most sensitive tes. + Molecular methods: © Reverse transcriptase PCR (RT-PCR) is the most sensitive, 2 type specific. © Real time RT: PCR can be used to quantitate the viral load in the clinical sample. Scanned with CamScannerInactivated Influenza vaccines: + Whole virus (WV) vaccine + Subvirion (SV) vaccine + Surface antigen vaccines. Live nasal spray (Trivalent) contains: + Influenza A (H1N1) virus + Influenza A (H3N2) virus + Influenza B virus. Sialic acid receptors determines pathogenicity of Influenza: +" 2-6 receptors: Specific for human influenza + 2-3 receptors: Specific for avian infuenca + Both a 2-3.and o2-6 receptors: Found in pigs, hence pigs are the MC mixing vessels. ‘Avian flu strains infecting humans: + AIHSN1 (MC type) ‘+ AHTNT (Netherlands) ‘+ AIHN2 ( Hong Kong) + AJH7NS (caused an outbreak in China, 2013). Prepared in allantoic cavity of egg, contains HA antigen (15 g/dose). + 2doses, given by IM route, efficacy 70-90%, lasts for 6-12 months, + Single dose is administered by IM or SC route. Protectivity is about 50-80%, Immunity lasts for 6-12 months. + Indication: Annual inactivated influenza vaccination is recommended for high-risk ‘groups such as children, older age, underlying chronic lung, cardiac, renal, hepatic, and CNS conditions, low immunity (HIV ) and pregnancy. + Contraindication: People who have allergy to eggs or have history of hypersensiti previous dose. + Inactivated vaccines are of three types. All are efficacious. © Whole Virus (WV) vaccine: Contains intact, inactivated viruses © Subvirion (SV) vaccine: Contains purified virus disrupted with detergents. © Surface antigen vaccines contain purified HA and NA glycoproteins. ty to Live Nasal Spray (Trivalent) © It is trivalent: Contains influenza A (HIN1) virus, influenza A (H3N2) virus and influenza B virus. ‘+ Stimulates both local + systemic immunity, not given if immunity is low. + Cold-adapted strain is used. + Indication: Recommended to all healthy persons of 2-49 years age, but is not given to high risk groups. ‘Treatment ‘Matrix M2 inl Sialic Acid Receptors Determines Pathogenicity Influenza virus entry into the host cells is dependent on type of sialic acid receptors present cn the host cell surfaces; which are specific for HA antigens of influenza. * a 2-6 sialic acid receptors are specific for human influenza strains and are found abundantly on human upper respiratory tract epithelium but not on lower upper respiratory tract. This explains why most human flu strains cause mild upper respiratory tract infections but not pneumonia. * a 2-3 sialic acid receptors are specific for avian influenza strains and are found abundantly on bird's intestine. © Inhumans, they are present in very few numbers on lower upper respiratory tract. © Thisexplains why avian flu strains cannot easily infect humans and need close contact. tor (amantadine) and neuraminidase inhibitor (oseltamivir). However, once infected, they can infect lower respiratory tract and cause pneumonia. Why pigs are the most common mixing vessels? + Both a2-3and a 2-6 sialic acid receptors are found on the respiratory epithelium of pigs and swine flu strains have specificity for both the receptor types. * Hence pigs can be infected simultaneously by human, swine and avian strains, thus serving as a mixing vessel where reassortment takes place. Avian Flu Infection in Humans Birds are the primary reservoir for influenza viruses. It is believed that, to date, all human pandemic strains have originated by reassortment between avian and human influenza viruses and the mixing has occurred in pigs. + A/IBNI is the MC avian flu strain that has been endemic in the world. © Origin: It was first reported from Hong Kong in 1997 and has spread to various countries including India. Scanned with CamScannerMyxoviruses and Rubella © Transmission to man occurs only from birds, and requires close respiratory contact. Less morbidity: As there is no human-to-human transmission, morbidity is less © More mortality: Avian flu strains are highly virulent (due to PB1F2 protein) and mor- tality rate is > 60%. © Clinical feature: H5N1 avian flu strains are associated with higher rates of pneumo- nia (> 50%) and extrapulmonary manifestations such as diarrhea and CNS involve- ment. * Other avian flu strains that can cause human infections are: A/H7N7 (Netherlands), A/H9N2 ( Hong Kong) and A/H7N9 (caused an outbreak in China, 2013) + Laboratory diagnosis: By real time RT- PCR detecting specific HA and NA genes. + Treatment: Drug of choice is oseltamivir (Tamiflu). A/H1N1 2009 Flu It has caused the most recent pandemic of influenza, emerged in California in March 2009 and rapidly spread to the entire world including India, over the next few months. WHO declared the pandemic in 11th June 2009. INI situation in ina: Epidemiology f'Sinee 2009 "about 53,943 Origin: HIN1 2009 flu originated by genetic reassortment of four strains (I Human | cases and 3,315 deaths due strain + 2 swine strains + 1 avian strain) and the mixing has occurred in pigs. eR HINi ete pleas aoe + Though people commonly use the word ‘swine flu’ to describe HIN1 2008 flu, but this not the correct terminology ait isa ressotant of four strains an + Transmission It can be transmitted from human to human, which has accounted forts |+ However a threatening rapid spread. oi 3 + However, itis less virulent (as it lacks the P81F2 protein). Therefore in contrast toHSN1, | S820."\am aoas deaths op the HINT 2009 flu has caused more morbidity but less mortality. fo March 30th 2015) The * Situation in World: Currently, World is in the post pandemic period except in India and | worst hit states are Rajasthan, New Zealand where still local intense transmission is ongoing, Gujarat, | Maharashtra and ‘Madhya Pradesh. + Situation in India: (© Since 2009 about 53,943 cases and 3,315 deaths due to HIN1 were reported from India, out of which in 2013 alone nearly 708 cases with 132 deaths have occurred. © However, a threatening outbreak of HINI started again in 2015 affecting 33,761 people with 2035 deaths (up to March 30th 2015). The worst hit states are Rajasthan, Gujarat, Maharashtra and Madhya Pradesh. Clinical Feature + Uncomplicated influenza: Most of the cases are present with mild URTI and diarrhea. + Complicated/severe influenza can occur very rarely in high risk groups, characterized such as secon terial pneumonia, dehydration, CNS involvement, and_ | H181 Origin: by features such wary bacterial pne dehydration, CNS involvement, and | ATO Erg onetic multiorgan failure. eral tr arsre Human strain + 2 swine strains Laboratory Diagnosis ++ avian strain) and the mixing has occurred in pigs. Real time RT-PCR can detect and quantify the specific HA and NA genes. Treatment HINI flu is resistant to amantadine. Drug of choice is neuraminidase inhi + Oseltamivir (Tamiflu) tablet: 75 mg twice a day for 5 days + Zanamivir (10 mg, inhalational form) Prophylaxis + Vaccine: Both killed injectable and live nasal spray vaccines are available for A/H1N12009 flu. + Chemoprophylaxis: Oseltamivir-75 mg oncea day, duration depends on the clinical setting, Scanned with CamScannerMyxovirus Infections of Respiratory Tract Paramyxoviridae 1 Parainfluenza Viruses CHAPTER 66 Myxoviruses are a group of viruses that bind to mucin receptors on the surface of RBCs (myxo in Greek meaning ‘ mucin’); resulting in clumping of RBCs together to cause ‘hemagglutination. Most of the myxoviruses are respiratory pathogens. Myxoviruses are divided into two families—(1) ‘Onthomyxoviridae and (2) Paramyxoviridae. Both differ from ceach otherin variousaspects; the mostimportant difference is the presence of segmented RNA in Orthomyxoviridae family. Important human pathogens are: + Orthomysoviridae: Influenza viruses —cause upper respiratory tract infections (URTIs), rarely can cause pneumonia + Paramyxoviridae: It includes several viruses = Parainfluenza virus: Mainly cause laryngotracheo- bronchitis and other URTIs ‘= Mumps virus: Causes parotitis in children (salivary gland infection) and rarely complications such as meningitis ‘= Measles virus: Cause exanthematouslesions,can lead to rare but serious complications of CNS (Chapter 56 and74) 1 Respiratory syncytial virus: Causesacute bronchiolitis ininfants = Metapneumovirus: Causes URTIs = Zoonotic paramyxoviruses such as Nipah and Hendra viruses: Mainly cause encephalitis (Chapter 74). ORTHOMYXOVIRIDAE INFECTIONS Influenza viruses are the members of Orthomyxoviridae family. They are one of the major causes of morbidity and mortality and have been responsible for several epidemics ‘and pandemics of respiratory diseases in the last two centuries. INFLUENZA Influenza viruses consist of four genera—influenza A, B, CandD. 1 Respiratory Syncytial Vieus Morphology (Fig. 66.1) Influenza viruses are spherical in shape, measure about 80-120 nm in size. + Helical symmetry: Itcomprises ofahelical nucleocapsid, surrounded by an envelope ‘ Viral RNA comprises of multiple segments of negative sense single stranded RNA. Each segment codes for a specific vial protein having a specific function 1 Influenza A and B contain eight segments of RNA ‘= Influenza C and D contain seven segments of RNA. ‘The segment coding for neuraminidase is absent. + Site of replication: RNA replication occurs typically in the nucleus (in contrast to most other RNA viruses which replicate in the cytoplasm) + Viral proteins: Influenza virus contains eight structural proteins PB1, PB2, PA, NP, HA, NA, M1 and M2)and two non-structural proteins (NS1 and NS2) = PBI, PB2, and PA are the polymerase proteins responsible for RNA transcription and replication = Nucleoprotein (NP) is the major capsid protein, associated with viral RNA to forma ribonucleoprotein (RNP) or nucleocapsid with a helical symmetry Fig. 66.1: Influenza virus (schematic diagram). Scanned with CamScannerCHAPTER 66 @ Myxovirus Infections of Respiratory Tract 649 = Matrix proteins: M1 protein is the major viral protein. It forms a shell (protein layer) underneath the envelope. M2 proteins form ion channels in the envelope, help in transport of molecules = Non-structural proteins: NSI is an interferon antagonist and inhibits pre-mRNA splicing. NS2 helps in export of molecules across the nucleus = Hemagglutinin (HA) and neuraminidase (NA) are the glycoproteins inserted into the lipid envelope. Envelope: It consists ofa lipid envelope into which two types of glycoproteins are inserted. 1. Hemagglutinin (HA): It is triangular-shaped peplomer. It binds to mucin or sialic acid receptors on the respiratory epithelial cells, thus facilitating viral entry 2, Neuraminidase (NA): It is mushroom shaped, present fewer in number than HA. It is a sialidase enzyme that degrades the sialic acid receptors on the host cells 4+ Itfacilitates release of virus particles from infected cell surfaces during budding process by preventing self-aggregation of virions to the host cells NA also helps the virus to pass through the mucin layer in the respiratory tract to reach the target epithelial ces. Antigenic Subtypes and Nomenclature Based on RNPand M proteins, influenza viruses are divided, into four genera: A, B, Cand D. ‘Subtypes: Based on HA and NA antigens, % Influenza A has distinct 18 H subtypes (H1 to H18) and 11 N subtypes (NI-N11) = Most of the subtypes infect animals and birds, but occasionally undergo genetic changes and infect humans to cause major epidemics and pandem- ies ‘= Forexample, Six HA (H1, H2, H3, H5, H7 and H9) and two NA (NI and N2) subtypes have been recovered from humans. + Influenza B viruses are not classified into subtypes, but have diverged into lineages. Currently, circulating influenza type B viruses belong to either B/ Yamagata or B/Victoria lineage + Influenza Cvirusis detected less frequently and usually ‘causes mild infections, thus does not present public health importance + Influenza D virus primarily infects cattle and are not pathogenic to humans. ‘The standard nomenclature system for influenza virus: ‘Any influenza virus isolates should be designated based on the following information: Influenza virus type/ host (indicated only for non-human origin)/geographical origin/strain number/year of isolation/(HA NA subtype). For examples: Human strain: Influenza A/Hong Kong/03/1968 (H3N2) © Non-human strain: Influenza A/swine/lowa/15/ 1930 (HIN}), Antigenic Variation Antigenic variation is the unique property of influenza viruses, which is due to the result of antigenic changes occurring in HA and NA peplomers. Itis of two types ‘Antigenic Variations 1. Antigenic Drift Itisa minor change occurring due to point mutations in the HA/NA gene, resulting in alteration of amino acid sequence (of the antigenic sites on HA/NA, such that virus can partially ‘escape recognition by the host's immune system. The new variant must sustain two or more mutations to become cepidemiologically significant. @ Seen in both influenza virus type—A and 8 {Results in outbreaks and minor periodic epidemics Antigenic drift occurs more frequently, every 2-3 years. 2. Antigenic Shift Itis an abrupt, major drastic, discontinuous variation in the sequence of a viral surface protein (HA/NA), that occurs due to genetic reassortment between genomes of two ‘or more influenza viruses infecting the same host cells; re- sulting in a new virus strain, unrelated antigenically to the predecessor strains. Thus, antibodies developed against previous strain (due to infection or vaccination) become ineffective, Occurs only in influenza A virus @ Results in pandemics and major epidemics, e.g. HIN1 pandemics of 2009 Antigenic shift occurs less frequently every 10-20 years. Pathogenesis + Transmission: It is transmitted by (i) inhalation of respiratory droplets generated by coughing and sneezing, This mode can infectonly those people who are within I-meter distance, (ii) via contact with surfaces or fomites infected with respiratory droplets and then touching nose, eyes or mouth + Target cell entry: Viral HA attaches to specific sialic acid receptors on the respiratory mucosa that leads to viral entry Multiply locally: Virus replicates in the infected cells and infectious daughter virions spread to the adjacent of respiratory epithelial cells over several hours % Spread: Very rarely, virus spreads to the lower respiratory tract or spills over bloodstream to involve extra- pulmonary sites + Local damage: Influenza virus infection causes cellular destruction and desquamation of superficial mucosa of the respiratory tract, which may predispose to secondary bacterial infection. Scanned with CamScannerHost Immune Response Humoral immunity: It isthe predominant immunity that provides resistance against influenza infections. Immunity developed is both type and subtype-specific and long- lasting. Antibodies against HA and NA are protective in nature, and are subtype-specific Antibodies to HA prevent initiation of infection by inhibiting viral entry; whereas antibodies to NA decrease the severity ofthe disease and prevent the transmission of the virus to contacts ‘® Antibodies against other viral proteins are not protective ® Antibodies against the ribonucleoprotein are type- specificand are useful in typing viral isolates as influenza Aor Bor + All the three types of influenza viruses (i.e. A, Band C) are antigenically unrelated and there is no cross-pro- tection Immunity may be incomplete following influenza infection; reinfection can occur with the same virus. Components of both cell-mediated immunity (e.g. cytotoxic T cells) and innate immunity (NK cells, interferons) are also important in providing immunity against influenza infections. Clinical Manifestations Incubation Period Itis about 18-72 hours, which directly depends upon the inoculum size and the immune status of the host, Uncomplicated influenza (Flu Syndrome) Majority of the individuals are either asymptomatic or develop minor upper respiratory symptoms such as chills, headache, and dry cough, followed by high-grade fever, myalgia and anorexia. It isa self-limiting condition, indistinguishable from the infections caused by other upper respiratory tract pathogens. ‘Complications ® Pneumonia: Secondary bacterial pneumonia is the ‘most common complication to occurin patients infected ‘with influenza virus. Common agents re staphylococci, pneumococci and Haemophilus influenzae. Primary influenza pneumonia is rare but leads to more severe complication Other respiratory tract complications include worsening of chronic obstructive pulmonary disease, exacerbation of chronic bronchitis, bronchiolitis, otitis media, parotitis and asthma Extrapulmonary complications: Myositis, rhabdo- myolysis, myocarditis, encephalitis, post-influenza Guillian-Barre syndrome + Reye's syndrome: It is fatty degeneration of liver with acute encephalopathy occurring in children and SECTIONS © Respiratory Tract Infections adolescents (2 to 16 years of age) following aspirin or salicylate intake. Though the cause is unknown, this condition is often seen following influenza B, varicella- zoster and rarely influenza A viral infections. Epidemiology Influenza viruses cause seasonal flu epidemics worldwide almost every year, however they differ widely in severity and the extent of spread % Incidence: It isestimated that annually about3-5 million cases of severe illness and 3-6 lakhs of deaths occur due to seasonal flu epidemics worldwide and is associated with significant economic impact % Global pandemics of novel influenza A subtypes occur every 10-40 years, which can cause much higher mortality than seasonal flu % Seasonality: Influenza outbreaks are common during winters. The most common seasonal flu strain varies, from season to season and from place to place (e.g. H3N2 in Puducherry in 2018) Annual attack rate: About 5-10% in adults and 20-30% in children are infected annually + Epidemiological pattern: It depends upon the nature of antigenic variation that occurs in the influenza types (as described earlier) Risk Factors Following risk factors are important determinants for patients ‘going for complications following influenza. Age: Child of age < 2 years or age 2 65 years 2 Chronic diseases: Chronic pulmonary, cardiac, renal, hematologic, metabolic, neurological, and neuro- developmental disorders 2 Immunosuppression (including HIV/AIDS, use of long- term corticosteroids, post-transplant patients), diabetes mellitus 2 Other risk factors: Extreme obesity, residing in nursing home, Americanindians and Alaska Natives, and pregnancy. People with these risk factors are the first priority group for receiving influenza vaccination. History of Influenza Outbreaks ‘Till now several influenza pandemics and major epidemics have occurred worldwide (Table 66.1), ‘ Seroarcheology: The outbreaks that occurred prior to influenza isolation (influenza isolated firstin 1933 using ferrets) were detected later by retrospective serologic survey of individuals alive during those years ‘The severe most pandemic (Spanish flu) recorded so far was the swine flu strain HINI in 1918-1919, where >50 million people died, mostly due to secondary bacterial pneumonia. ‘his strain was not a reassortant, but believed to be derived entirely from an avian strain that had adapted to human conditions and pigs acted asa mixing vessel Scanned with CamScannerCHAPTER 66 @ Myxovirus Infections of Respiratory Tract 1889-1890 HANS ‘Severe pandemic 1900-1903 -H3NB ‘Moderate epidemic 1918-1919 HINT (HswNt) Severe pandemic (Spanish fu) 1933-1935 -HINI+(HONT) Mild epidemic 1946-1947 HINT ‘Mild epidemic 1957-1958 H2N2(Asianflu) Severe pandemic 1968-1969 -H3N2 (Hong Kong lu) Moderate pandemic 1977-1978" HINT (Russian lu) Mild pandemic 2009-2010 HIN1 pdmo9 Pandemic “Hemaggutnins formerly designated as How and HO are now casiied as variants of “From 1978 upto 2008-2009, viuses of theHIN1 and HBN2 subtypes chcuated ‘ether in aerating years or concurrent. This was followed by series of several epidemics and pandemics as mentioned in Table 66.1 Sialic Acid Receptors Sialic acid receptors found on the host cell surfaces are specific for HA antigens of influenza virus, which in turn determines the different host specificities of influenza virus. Alpha-2-6 sialic acid receptors are specific for human influenza strains and are found abundantly on human upper respiratory tract epithelium, but not on lower respiratory tract. This explains why most human flu strains cause mild upper respiratory tract infections but not pneumonia Alpha-2-3 sialic acid receptors are specific for avian influenza strains and are found abundantly on bird's Intestinal epithelium > Inhumans, they are present in very few numbers on ‘upper respiratory tract, and also on some epithelial cells inthe lower tract This explains why avian flu strains cannot easily infect humans and need close contact. However, once Infected, they can infect the lower respiratory tract and ‘cause pneumonia, Why pigs are the most common mixing vessels? 1 Botha-2-3and a-2-6 salicacid receptors are found on the same respiratory epithelial cells of pigs and swine flu strains have specificity for both the receptor types @-Hence pigs can be infected simultaneously by human, swine and avian strains thus serving as a mixing vessel @_ Reassortment between the segments of various strains can take place inside the same swine cell. Avian Flu Birds are the primary reservoir for influenza viruses. ® All influenza subtypes (18H types and 11N types) are found in birds and some of the subtypes can be transmitted to mammals (e.g. H1, H2, H3, H5, H7 and H9 to humans; H1 and H3 to swine; and H3 and H7 to horses) + Usually the avian flu strains are highly virulent as they possess PBIF2 protein, which targets host mitochondria ‘and induces apoptosis. Avian Flu Infection in Birds + Bird flu strains are highly lethal to chickens and turkeys (but avirulent to ducks) and are the major cause of ‘economic loss in poultry causing severe mortality in chickens % Unlike in mammals, avian flu multiplies in intestinal tracts of birds and shed through feces into water (avian flu isa water-borne disease in birds) ‘© ‘The influenza viruses do not undergo antigenic variation in birds, because of the short life span of birds. Avian Flu Infection in Humans It is believed that, to date, all human pandemic strains have originated by reassortment between avian and hhuman influenza viruses and the mixing has occurred in pigs. A/HISN1 is the most common avian flu strain that has been endemic in the world for the past 15 years. Origin: Itwas first reported from Hong Kong in 1997 and has spread to various countries including India within few years Transmission to man occurs only from birds, and requires close respiratory contact + Less morbidity, but more mortality: As there is no human to human transmission, morbidity is less. However, the avian flu strains are highly virulent (due to presence of PBIF2 protein) and mortality rate is >60% Globally only 861 cases have been reported between 2003 102019, out of which about 455 people succumbed death (52%) Clinical features: H5N1 avian flu strains are associated with higher rates of pneumonia (250%) and. extrapulmonary manifestations such as diarthea and CNS involvement. Other avian flu strains infecting humans are: 4 A/H7N7(Netherlands) % A/H9N2 (Hong Kong) A/H7N9 (caused an outbreak in China, 2013). Laboratory Diagnosis (Avian Flu) Avian flu strains can be identified by real time reverse transcriptase PCR detecting specific HA and NA genes. Influenza A (H1N1) pdm09 It has caused the most recent pandemic of influenza, emerged in California in March 2009 and then rapidly spread to the entire world including India over the next few months.WHO declared the pandemic on 11 June 2008. Scanned with CamScanner652 SECTION 8 © Respiratory Tract Infections + Origin: HN1 2009 flu originated by geneticreassortment of four strains (1 human strain +2swine strains + 1 avian strain) and the mixing had occurred in pigs (Fig, 66.2) + Though people commonly use the word ‘swine flu’ to describe HINI 2009 flu, but this is not the correct terminology as itis a reassortant of four strains Transmission: It can be transmitted from human to human, which has accounted for its rapid spread + However, itisless virulent (asitlacks the PBI F2 protein) % Therefore in contrast to HSNI, the H1N1 2009 flu has caused more morbidity but less mortality. Clinical Features + Uncomplicated influenza: Most of the cases present with mild upper respiratory tract illness and diarrhea + Complicated/severe influenza can occur very rarely in high-risk groups, is characterized by features such as secondary bacterial pneumonia, dehydration, CNS involvement, and multiorgan failure. Reassortment in A ‘a swine host Pig a Lp» Mixing vessel = Novel human Human fu vius endemic vine Fig. 66.2: Evolution of pandemic influenza virus. rr eri ‘Mild fever plus cough/sore throat with or without bodyache, headache, diarthea and vomiting Category A plus any one: |. High-grade fever and severe sore twoat or li. Presence of any of the risk factors as described earlier in the highlight bor, under epidemiology) Category C Severe acute respiratory syndrome (SARI) Category 8 plus any one: 1, Breathlessness, chest pain fallin blood pressure, sputum ‘mixed with blood, buish discoloration of nails {i Children with influenza-like illness who had a severe disease as manifested by the red flag signs (inability to feed ‘well, convulsions, dificultyin breathing, etc) li, Worsening of underlying chronic conditions Categorization of Seasonal Influenza A/H1N1 “Ministry of Health and Family Welfare, Government of India has published the guideline on categorization of seasonal influenza A/HIN1 cases (Table 66.2). While screening the patients with influenza ike illness, this guideline helps in taking decision on performing laboratory test, initiating antiviral treatment and putting the patient on home isolation or hospitalization. Epidemiological Surveillance for Influenza Influenza surveillance is routinely carried out globally and also at national level. This helps to monitor the changes the circulating influenza strain and. serves as a global alert mechanism for the emergence of pandemic influenza viruses. + GISRS: Influenza surveillance has been conducted globally through Global Influenza Surveillance and Response System (GISRS) under World Health Organization (WHO) + IDSP (Integrated Disease Surveillance Program) under NCDC (National Center for Disease Control) conducts Influenza (HIN1) surveillance in India + India (2019): According to IDSP Report 2019—28,798 cases of HINI were reported with 1,218 deaths. Rajasthan reported the maximum cases, followed by Gujarat; whereas Maharashtra reported the maximum deaths. The mortality rate is around 4-5% every year 4 World: According to WHO, 3-6 lakh deaths occur due to seasonal flu annually (4.0-8.8 per 1 lakh population). Global pandemics of novel influenza A subtypes occur every 10-40 years, which can cause much higher ‘mortality than seasonal flu. during screening for home isolation, testing, tre Sone Laboratory testing for HIN1—not required Treatment—only symptomatic, antiviral drugs not required Isolation confine patients at home, avoid contact with public and high-risk members in the family Laboratory testing for HIN!—not required Treatment—symptomatic treatment required. Antiviral drug (oseltamivir) may be required [solation—confine patients at home, avoid contact with ‘public and high-risk members in the family Laboratory testing for HIN! —required Immediate hospitaization—required Teatment-—start antiviral drug osetamiv?)immedtatly Without waiting for laboratory rest Iolation—all components droplet precaution tobe followed (fer prevention of influenza’ section) * Rea me reverse transcriptase PCRs recommended to detect and quantly the speiic HA and NA genes of HIN + Ozetamivi anv) tablet or Zanamvr inhalation frm). Scanned with CamScannerCHAPTER 66 © Myxovirus Infections of Respiratory Tract Cnr: Influenza Specimen: Nasopharyngeal swab, kept at 4°C Isolation of virus: > Inoculation in embryonated eggs and primary monkey kidney celines > Growth is detected by hemadsorption, hemagglutination test 1 Viral antigens detection by direct F test |B Molecular methods: Simultaneously detects seasonal flu strains such as A/HINI, A/H3N2, Influenza B > RTPCR:detects viral RNA > Real ime-RT PCR: quantifies viral RNA 1 Antibody detection by hemagglutination inhibition test, neutralization test and ELISA. Laboratory Diagnosis Specimen Collection + Ideal specimens are nasopharyngeal swab or lavage fluid, nasal aspirate or to a less extent throat swab + Swabs with a synthetic tip (e.g. polyester or Dacron swabs) are best for specimen collection (Fig. 66.3). Cotton or alginate swabs are unsatisfactory + Transport: Swabs are immediately put inside the viral transport media, kept at 4°C during transport up to 4 days, thereafter at -707 Isolation of Virus Embryonated eggs (amniotic cavity) and primary monkey kidney cell lines have been the methods of choice for the isolation of influenza viruses in the past. The viral growth in cell line was detected by hemadsorption or hemagglutination test. Because of technical difficulty, isolation is not routinely performed for diagnostics purpose. Direct Immunofluorescence Test Viral antigens coated onto epithelial cells can be directly detected in nasal aspirates by using fluorescent tagged antibodies. This is rapid, but less sensitive than viral isolation, Molecular Methods Molecular methods have revolutionized the diagnosis of influenza. RT-PCR (reverse transcriptase polymerase chain reaction): It is highly sensitive, specific and rapid Fig, 66.3: Viral transport medium and swab, Source Department of Microbiology, MER, Puduchery (wth permis). 2 ere bye AM a 8 a: Qiaa ss : one 2a 28} Sanco [i - nt a on me” “oem Figs 66.48 to D: Real time RT-PCR showing the result amplification curves) of specimen(s) tested positive fr Influenza type: A. A/H.N,; B. A/H.N,:. Type 8; D, Negative (refer Table 66.3). Source Department of Microbiology, JPMER, Puduchery (th permission). sonal influenza types. Table 66.3: Real time RT-PCR forse Specimens positive with Pee en) Parent ores Influenza A matricgene) + erie es = HINT (HA gene) a a = = HBN2 (HA gene) - as Be z Influenza B(HAgene) =~ : z 5 RNP(ibonuckeoprtcin” + = tt 5 *RNP(bonvcleoprotel sued astral contol real ne RT-PCR i not detected then the et considered abd (cumaround time of <1 day). Itcan also detect the specific type and subtype of influenza virus + Real-time RT-PCR: It is currently the gold standard method for influenza diagnosis. It is quantitative, has higher sensitivity and specificity than RT-PCR with turnaround time of 2-3 hours. It simultaneously detects the three common seasonal fu strains (A/HAN1, A/H3N2 and type B). The result is expressed as the emission of fluorescence during the cycles as described in Figure 66.4 and Table 66.3 % BioFire FilmArray Respiratory Panel (RP) tests simultaneously 20 respiratory pathogens, including Influenza A, Influenza A/H1, Influenza A/H3, Influenza A/H1-2009 and Influenza B. Antibody Detection (Serology) Various assays are available to detect subtype specific serum antibodies by using specific influenza antigens. It Scanned with CamScanneris mainly useful for sero-epidemiology purpose, not for clinical diagnosis; as antibodies may be present in normal individuals. The tests available are: ELISA, neutralization test, and previously used HAI (hemagglutination inhibition) test. UU sfuensze Specific antiviral therapy is available for influenza virus infection. Start therapy within 48 hours of onset of symptoms for influenza. & Neuraminidase inhibitors (such as zanamivir, oseltamivir {and peramivir) can be administered for influenza A and influenza B infections > Its the drug of choice for A/HIN1 2009 flu, A/HSN1 avian flu and influenza-8 » Dosage: ¢ Oseltamivir (Tamiflu 75 mg tablets) ¢ Zanamivir (10 mg, inhalational form). > Schedule: For treatment—given twice a day for 5 days ‘¢ For chemoprophylaxis—given once daily. Duration depends on the clinical setting; usually 7 days. @ Matrix protein M2 inhibitor such as amantadine and rimantadine can be given for some strains of influenza A infection. However, strains of A/HIN1 2009 flu and A/HSNI avian flu and influenza B virus have developed resistance. Prevention General Preventive Measures Measures of droplet precaution (Chapter 21) should be followed: * Strict hand hygiene + Isolation room: Patients should be kept in room or cohorting to be followed Containment of coughs and sneezes = Respiratory hygiene and cough etiquette = Use of personal protective equipment (PPE) such as gloves, 3-ply masks, gown and googles for a HCW. Patient should wear a mask. % Work restriction: CDC recommends that people with influenza-like illness remain at home until at least 24 hours after they are free of fever <100°F) without the use of fever-reducing medications. Vaccine Prophylaxis Vaccine Strains Based on WHO recommendations, influenza vaccines are prepared every year. % Strains to be included in the vaccine depend upon the strains isolated in the previous influenza seasons and strains that are anticipated to circulate in the upcoming © Formulations: Influenza vaccines are available in cocktail of either three strains or four strains = Trivalent form: Comprises of three strains: A/H1N1, A/H3N2 and influenza B strain = Quadrivalent form: Comprise of four strains: A/ HINI, A/H3N2 and two lineages of influenza B strain. SECTION 8 © Respiratory Tract Infections ‘ Composition: The lineages of A/HIN1, A/H3N2 and influenza B strains included in vaccine change every year = The lineages included in vaccine are different for northern and southern hemispheres = WHO constantly reviews the vaccine composition and decides on which lineages to be included in vaccine every yeat. ® Types: Both injectable (inactivated) and nasal spray (live attenuated) vaccines are available. Injectable Vaccines Injectable vaccines are the most widely used vaccines in immunization programs. Types: There are three types of injectable vaccines 1. Inactivated influenza vaccine (IIV), e.g. Fluzone: It is prepared by growing the vaccine strains in embryonated chick eggs; then harvested, purified, inactivated and then standardized to contain 15 4g, of HA/dose. 2. Cell culture-based inactivated influenza vaccine (ccl1V3); e.g, Flucelvax: Same as IIV, but prepared in cell lines such as Madin-Darby Canine Kidney (MDCK) cell line. 3. Recombinant influenza vaccine (RIV), €.g. FluBlok: Contains recombinant influenza HA antigens in trivalent/quadrivalent formulations, RIV does not contain any egg protein, ® Schedule: Single dose administered by intramuscular (IM) route; except for 6 months-8 years of age (2 doses are required >4 weeks apart) Timing of vaccination: Optimally before onset of influenza season, i.e. by end of October % Efficacy: The vaccine efficacy varies from 25-67% (25% for H3N2, 42% against type B and 67% against HIN1). ‘The efficacy is lower if vaccine virus does not match to currently circulating viruses in the locality. Immunity lasts for 6-12 months Side effects: Mild reactions can occur in 5% of cases such as redness at injection site, fever and aches. Serious side effects such as allergic reactions can occur very rarely + Indications: Routine annual influenza vaccination is recommended for all persons aged >6 months who do not have contraindications. If not feasible, then high- risk groups should be given first priority for vaccination (high-risk group individuals have been listed in highlight box, under epidemiology earlier) Contraindication: //V should not be administered to people who have history of severe allergic reaction to previous dose of vaccine. In patients with egg allergy, vaccine can be given, but under supervision © Travelers: If traveling to an area of increased influenza activity; can consider vaccination, preferably 22 weeks before departure. Scanned with CamScannerCHAPTER 66 © Myxovirus Infections of Respiratory Tract Live Attenuated Influenza Vaccine (LAIV) This vaccine is generated by reassortment between currently circulating strains of influenza A and B virus with a cold-adapted attenuated master strain which is adapted to grow at 25-33°C. + Such ive attenuated strains can growin upper respiratory tract (at 33°C) but notin lower respiratory tract (at 37°C); therefore they may cause mild flu like symptoms but never infect lower respiratory tract, hence never cause serious adverse effects + Itisa trivalent vaccine, administered by intranasal spray Indication: It can be given to all healthy persons of 2-49 yearsage (except in pregnancy), but isnot given to high- risk groups. Chemoprophylaxis Antiviral drugs are not recommended for routine seasonal or pre-exposure prophylaxis. tis recommended only for post-exposure and during outbreak situations in hospitals. % Indications: Following exposure to an influenza case, it is recommended to the following groups: (i) if not vaccinated or vaccinated recently (<2 weeks), (ii) HIV infected people Duration: = Non-outbreak exposure e.g, in community): Itshould bbe started as soon as possible following exposure (within 48 hours) and continued for 7 days = During outbreaks in hospitals (for elderly persons, children and health care workers): Duration for a minimum of 2 weeks, and to be continued up to 1 ‘week after the last known case was identified, Antiviral drugs recommended are: ‘© Oseltamivir is the drug of choice. Itis given as 75 mg orally, once a day for 7 days ‘= Zanamivir: 10 mg (two 5-mg inhalations) once daily for 7 days. + Efficacy: The efficacy of chemoprophylaxis is about 70-90% in preventing influenza. Pea a Paramyxoviridae contains a group of viruses, which are transmitted via the respiratory route following which: % ‘They may cause localized respiratory infection inchildren (eg respiratory syncytial virus, metapneumoviruses and parainfluenza viruses) or; ‘They may disseminate throughout the body to cause highly contagious diseases of childhood such asmumps (parotid enlargement) and measles (Chapter 56).. Paramyxoviruses resemble orthomyxoviruses in morpho- logy, but they differ by the following properties (Fig. 66.5). 4 ‘Theyarelarger(100-300nm)insize and more pleomorphic % Possess linear non-segmented RNA (compared to segmented RNA in influenza virus) Fig. 66.5: Schematic diagram of paramyxoviruses (measles virus). Contain six structural proteins (compared to 8 in influenza virus) ¢ HA and NA antigens: The paramyxoviruses vary from each other in expression of HA and NA antigens = Parainfluenza and mumps possess both HA and NA antigens (similar to influenza virus) ‘= Measles virus possess HA, butlack NA = RSV and metapneumoviruses lack both HA and NA. ‘Note: There are few zoonotic paramyxoviruses stich as Nipah and Hendra viruses, which can occasionally infect humans (Chapter 74), PARAINFLUENZA Human parainfluenza viruses are one of the major causes of lower respiratory tract disease in young children. They have five serotypes: % ‘Types | and 3 belong to the genus Respirovirus % ‘Types 2, 4 aand 4b belong to the genus Rubulavirus. Clinical Manifestations + Transmission is by respiratory route (by direct salivary contact or by large-droplet aerosols) + The incubation period appears to be 5-6 days + Virus multiplies locally and causes various respiratory ‘manifestations such as: = Mild common cold syndrome like rhinitis and pharyngitis are the most common presentation, seen ‘with all serotypes ‘= Croup (laryngotracheobronchitis): + Occurs in 2-3% of cases + Typically seen with type Land 2 4+ Involves children (between 6to 18 months of age). = Pneumonia or bronchioliti + Occurs very rarely ‘+ Seen especially with serotype 3 4+ Involves infants below 6 months of age. = Otitis media: Itis the most common complication of parainfluenza virus infection. + Reinfections are common, but less severe. There is no cross protection between the serotypes. Scanned with CamScannerKAPLAN) MEDICAL 339 Microbiology ORTHOMYXOVIRIDAE Family Characteristics ‘+ Negative-sense ssRNA + Enveloped + Segmented (8 segments) + Helical Figure II-4-28. Orthomyxovirus Viruses of Medical Importance + Influenza A * Influenza B Key Vignette Clues Influenza Virus Influenza Distinguishing Features Patient with headache, malaise, fever, uel + Envelope conta myalgia, cough two glycoproteins, Hand N Used to serotype virus Reservoir + Influenza A (birds, pigs, humans) + Influenza B (humans only) Transmission + Direct contact + Respiratory + 1997 HSNI strain jumped directly from birds to humans + 2009 HIN1 strain—quadruple reassortment virus (North American swine, avian, human; Asian and European swine) Pathogenesis + Antigenic drift Influenza A and B ~ Slight changes in antigenicity due to mutations in H and/or N = Causes epidemics Scanned with CamScanner340 KAPLAN) MEDICAL a aa aaa Chapter 4 « Medically Relevant Vi + Antigenic shift = Influenza A only Rare genetic reassortment ~ Coinfection of cells with two different strains of influ and H3N2); reassortment of segments of genome za. A (HSNI ~ Production of a new agent to which population has no immunity = Responsible for pandemics Disease: influenza + Headache and malaise + Fever, chills, myalgias, anorexia + Bronchiolitis, croup, otitis media, vomiting (younger children) + Pneumonia/secondary bacterial infections + Can lead to Rey syndrome or Guillain-Barré syndrome Diagnosis + Rapid tests (serology) + Clinical symptoms plus season Treatment + Amantadine/rimantadine (current isolates are commonly resistant) = Inhibit viral uncoating Administer orally + Zanamivir/oseltamivir ~ Neuraminidase inhibitors = Zanamivir is inhaled = Oseltamivir is given orally Prevention + Killed vaccine ~ ‘Two strains of influenza A (H3N2, HINI, for example) and one strain of influenza B are incorporated into the vaccine + Live, attenuated vaccine = Intranasal administration = Similar composition ~ No longer recommended Scanned with CamScanner