Comparison of the Pathology Caused by H1N1, H5N1, and H3N2 Influenza Viruses
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Archives of Medical Research 40 (2009) 655e661
REVIEW ARTICLE
Comparison of the Pathology Caused by H1N1, H5N1,
and H3N2 Influenza Viruses
Jeannette Guarnera and Reynaldo Falcón-Escobedob
a
Department of Pathology and Laboratory Medicine, Emory University Hospital, Atlanta, Georgia
b
Department of Pathology, Hospital Central Dr. Ignacio Morones Prieto, San Luis Potosi, Mexico
Received for publication September 8, 2009; accepted September 21, 2009 (ARCMED-D-09-00424).
The spectrum of morbidity and mortality of H1N1, H5N1, and H3N2 influenza A viruses
relates to the pathology they produce. In this review, we describe and compare the
pathology of these viruses in human cases and animal models. The 1918 H1N1, the novel
2009 H1N1 pandemic virus, and H5N1 show inflammation, congestion, and epithelial
necrosis of the larger airways (trachea, bronchi and bronchioles) with extension into
the alveoli causing diffuse alveolar damage. Seasonal influenza A viruses (H3N2 and
H1N1) have primarily caused inflammation, congestion and epithelial necrosis of the
larger airways with lesser extension of the inflammatory process to alveoli. Localization
of the inflammation and cellular damage relate to the presence of virus in different cell
types. Infections with 1918 H1N1, the novel 2009 H1N1 pandemic virus, and H5N1 show
virus in mucosal epithelial cells of the airways (from the nasopharynx to the bronchioles),
alveolar macrophages, and pneumocytes, whereas infections with seasonal influenza
viruses show viral antigens primarily in mucosal epithelial cells of the larger airways.
The increased morbidity that has been encountered with the 2009 H1N1 virus is related
to infection of cells in the upper and lower airways. The 2009 H1N1 virus shows similar
pathology to that encountered with other highly virulent influenza A viruses such as the
1918 H1N1 and H5N1 viruses. Ó 2009 IMSS. Published by Elsevier Inc.
Key Words: Pathology, Influenza A virus, Novel H1N1, avian H5N1, Seasonal H3N2, Human cases,
Animal models.
Introduction to recognition that the virus causing disease was of swine
origin (H1N1) (12,13). Reconstruction and successful
Preparedness for an influenza pandemic has included
animal infections with the 1918 H1N1 virus have added
surveillance for all influenza viruses but attention has been
to the spectrum of clinicopathological outcomes that the
primarily focused on transmission of avian H5N1 virus to
different influenza A viruses can cause (14,17). Before
humans and further transmission of H5N1 virus among
the 2009e2010 season begins, a comparison of the
humans. Attention to the H5N1 virus started in 1997 when
pathology between the recent 2009 H1N1 pandemic virus
humans became infected during a poultry outbreak of this
(18,19) and the other influenza A viruses may help to better
highly pathogenic avian influenza virus in Hong Kong
prepare healthcare professionals to understand the
(1e3). Animal studies and studies of autopsies of patients
morbidity and mortality that may be encountered.
infected with the different viruses (avian H5N1, swine
The signs and symptoms of H1N1, H5N1 and H3N2
H1N1, and seasonal H3N2) have revealed that the cells in
influenza A viruses is primarily in the respiratory tract
the respiratory tract these viruses infect are different result-
and the pathological damage encountered mirrors the clin-
ing in different pathologic pictures and morbidity and
ical presentation with the most frequent pathology observed
mortality rates (4e11). In addition, further molecular
in the airways and lungs. Clinical symptoms of myocarditis
studies of autopsy material from the 1918 pandemic lead
and encephalopathy have been described less frequently in
Address reprint requests to: Jeannette Guarner, MD, Emory University
patients with influenza A infections, and pathology in the
Hospital, 1634 Clifton Rd, Room C179A, Atlanta, GA, 30322; Phone 404- heart has been observed in some of these cases. It should
712-2631; E-mail: jguarne@emory.edu be noted that the pathological descriptions in humans come
0188-4409/09 $esee front matter. Copyright Ó 2009 IMSS. Published by Elsevier Inc.
doi: 10.1016/j.arcmed.2009.10.001
656 Guarner and Falcón-Escobedo/ Archives of Medical Research 40 (2009) 655e661
primarily from autopsies; thus, our knowledge relates to the The1918 H1N1 pandemic virus is recorded in history as
most severe cases with the poorest outcomes (20). Study of having caused the highest number of excess deaths (20).
the pathology of confirmed cases is important to be able to Multiple studies of the pathology were published at the
better understand the differences in presenting signs and time and newer studies have reviewed the archived
symptoms, duration of the disease, complications, and pathology material to ascertain the role of viral pneumonia
mortality that these viruses can produce. In addition, and bacterial superinfections (17,21e24). From the patho-
knowing the pathology these viruses cause will enable logical standpoint, H1N1 infections during the 1918
pathologists to perform presumptive diagnosis and seek pandemic can be divided into those patients with viral
other diagnostic techniques (PCR, viral culture) that will pneumonia and those with superimposed bacterial pneu-
allow confirmation of the virus involved for specific cases. monia. Those patients who died early during the infection
This paper will describe and compare the pathology showed viral pneumonia with focal necrosis of the alveolar
encountered in autopsies and animal models infected with walls associated with capillary thrombosis, prominent
H1N1, H5N1 and H3N2 influenza viruses. hyaline membrane formation, and intraalveolar edema. In
the trachea and major bronchi, autopsies showed necrotic
mucosa with edema and inflammation in the submucosa.
Pathology of H1N1
The loss of epithelium was more pronounced in the bron-
Detailed descriptions of the pathology of cases infected chioles where the epithelial layer is thinner. In comparison,
with the pandemic 2009 H1N1 virus have not yet appeared those patients with superimposed bacterial pneumonias
in the literature. The descriptions presented here are based showed abundant intraalveolar neutrophilic infiltrate asso-
on the authors’ observations of four cases and a published ciated with edema, fibrin and necrosis in a predominant
report of one case (18). The lung pathology of deceased lobular distribution. It is estimated that up to 90% of
patients infected with the pandemic 2009 H1N1 have patients who died during the 1918 H1N1 pandemic had
shown primarily diffuse alveolar damage with extensive bacterial superinfections with Streptococcus pneumoniae,
hyaline membrane formation and intraalveolar edema S. hemolyticus, and mixed pathogens being the most
(Figure 1A). The vessels in alveolar walls show thrombosis, frequently isolated bacteria (17). Myocarditis was reported
which has lead to necrosis of the alveolar walls (Figure 1B) in patients with the 1918 H1N1 infections (22).
and varying degrees of mixed inflammatory infiltrate. Several animal models have been used to study the
Alveolar epithelial cells and pneumocytes show reactive pathology of H1N1 infections and the results vary depend-
changes, and their nuclei appear enlarged. Desquamated ing on the host animal and virus used for the infection
macrophages and pneumocytes are observed within the (5,8,10,14e16,25). In guinea pigs and pigs, low pathogenic
alveolar spaces. Prominent proliferation of fibroblasts is seasonal H1N1 virus (A/Puerto Rico/8/34 and A/swine/
observed in patients with longer duration of disease. Intra- Thailand/HF6/05) showed viral antigens in the mucosal
alveolar hemorrhage and erythrophagocytosis have also epithelium associated with epithelial cell damage and could
been observed. Immunohistochemical assays have demon- be seen from the upper airways to the bronchioles (5,10).
strated viral antigens in pneumocytes type I and II and Damage of the epithelium resulted in increased mucous
intraalveolar macrophages (personal communication, production and mild inflammatory infiltrate. Inflammation
Dr. Sherif Zaki, Centers for Disease Control and Preven- surrounded bronchioles but did not spill into the lung inter-
tion, Atlanta, GA, 2009). Bacterial pneumonias, defined stitium. In ferrets, the seasonal H1N1 (A/Netherlands/26/
as accumulation of neutrophils in the alveoli, have been 2007) showed multifocal necrotizing rhinitis with abundant
observed in 30% of cases and may be either community viral antigens in the nasal turbinates and focal consolidation
acquired or nosocomial (ventilator-associated) (18,19). of the lungs without evidence of viral antigens in the lower
The trachea and major bronchi of deceased patients in- respiratory tract (25). When low pathogenic seasonal H1N1
fected with the pandemic 2009 H1N1 have shown necrotic virus (A/Puerto Rico/8/34) was used to infect mice, the
desquamation of the mucosa and edema and mixed inflam- lungs showed complete consolidation due to bronchitis
matory infiltrate in the submucosa (Figure 1C). The glands and interstitial pneumonia, the thymus was reduced in size
in these airways show loss of goblet cells and necrosis of by 30% due to depletion of cortical lymphocytes, and the
surrounding submucosa (Figure 1D). Viral antigens have heart showed inflammatory foci (8). Thus, for the low path-
been noted in the epithelial cells and mucous glands of ogenic seasonal H1N1 viruses the extent of damage de-
trachea, bronchi, and bronchioles (personal communica- pended on the infected host.
tion, Dr. Sherif Zaki, Centers for Disease Control and Animal models using ferrets and pandemic 2009 H1N1
Prevention, Atlanta, GA, 2009). PCR demonstrating virus have shown more extensive consolidation of the lungs than
in tissues from the respiratory tract and lung has shown that encountered with seasonal H1N1 virus (25). Histopath-
better sensitivity and specificity than immunohistochem- ologically, ferrets show multifocal mild to moderate necro-
istry. Myocardial infarction has been documented in several tizing rhinitis, tracheitis, bronchitis, and bronchiolitis, and
patients infected with the 2009 H1N1 virus (18). viral antigens are observed from the nasal turbinates to
Comparison of Pathology of Influenza A Viruses 657
Figure 1. Histopathological characteristics of fatal cases with confirmed pandemic 2009 H1N1 virus infection showing diffuse alveolar damage in the early
exudative phase characterized by hyaline membranes (A,B), intraalveolar edema (A), and alveolar wall necrosis (B) accompanied by inflammatory infiltrate
(A). The trachea and bronchi showed denuded mucosa (C,D) with inflammation (C) and necrosis (D) of the submucosa. Hematoxylin and eosin stains; orig-
inal magnifications: 10 (A), 20 (B,D), and 5 (C). Color version of this figure available online at www.arcmedres.com.
the bronchioles. The pandemic 2009 A (H1N1) influenza in bronchiolar epithelium and alveolar macrophages. These
viruses replicate to higher titers in lung tissue and can be animals have also shown increased expression of cytokines
recovered from the upper and lower airways as well as and chemokines.
the intestinal tract (26).
Infection of mice or monkeys with highly pathogenic re-
Pathology of H5N1
constructed 1918 H1N1 viruses (1918 HA/NA:WSN or
1918 HA/NA:Tx/91) produced large areas of pneumonia Even though the mortality from H5N1 infection is |60%,
that microscopically showed necrotizing bronchitis and there have been !15 autopsies reported in the literature
inflammation of alveolar walls by neutrophils and macro- (2e4,27e29). The lung pathology of deceased patients in-
phages (14e16). Necrotic debris was noted in the lumen fected with the H5N1 virus has shown diffuse alveolar
of bronchioles and alveoli. Viral antigens were localized damage in the lungs. Depending on the duration of illness,658 Guarner and Falcón-Escobedo/ Archives of Medical Research 40 (2009) 655e661 diffuse alveolar damage may be in the exudative early shown intraalveolar hemorrhage (Figure 2B), whereas phase and show hyaline membrane formation, congestion others have shown hemophagocytosis. Hemophagocytosis of vasculature, and lymphocytes infiltrating the interstitium has been noted not only in the lung but also in bone or in the later fibrous proliferative phase where fibroblast marrow, lymph nodes (Figure 2C), spleen, and liver. Other proliferation is added to the picture (Figure 2A) (3). Inflam- organs with pathology have included the spleen with deple- mation around bronchioles, alveolar interstitium, and in the tion and apoptosis of lymphocytes, kidneys with acute pleura has been present. Cells involved in the pathology tubular necrosis, and liver with necrosis, fatty change, acti- include presence of T-lymphocytes in the alveolar septae, vation of Kupffer cells, and cholestasis. Two cases have macrophages in the alveolar lumen, and hyperplasia and shown demyelinated areas in the brain with necrosis and desquamation of type II pneumocytes. Several cases have reactive histiocytes. Several techniques have demonstrated Figure 2. Histopathological characteristics of fatal cases with confirmed H5N1 virus infection showing diffuse alveolar damage in the later fibrous prolif- erative phase characterized by fibroblast proliferation (A) and deposition of collagen (A,B). Intraalveolar hemorrhage is seen in some cases (B) as well as hemophagocytosis (C). Hematoxylin and eosin stains; original magnifications: 10 (A), 5 (B), and 63 (C). Color version of this figure available online at www.arcmedres.com.
Comparison of Pathology of Influenza A Viruses 659 Figure 3. Histopathological characteristics of fatal cases with confirmed seasonal H3N2 virus infection showing mucosal desquamation and disorganization and submucosal congestion, hemorrhage, and inflammation in the trachea (A). The lungs show intraalveolar hemorrhage and edema (B). The heart shows focal areas of mononuclear inflammatory infiltrate (C). Hematoxylin and eosin stains; original magnifications: 10 (A), 5 (B), and 40 (C). Color version of this figure available online at www.arcmedres.com. virus that can replicate in the lung, and immunohistochem- The animals that have been used include mice, ferrets, ical assays have localized the viral antigens in ciliated and and primates. nonciliated epithelial cells and in type II pneumocytes. Macroscopically, all animals showed various degrees of Viral sequences and antigens have been found in the consolidation and discoloration in the lungs and hemor- brain, intestines, and placenta as well as lymphocytes and rhages in adipose tissue surrounding liver, intestines, macrophages. kidneys, and bladder (11,30). Macroscopic changes Several animal models have been used to study the occurred earlier in animals infected with the highly virulent pathology of H5N1 infections and the results vary depend- viruses compared to those infected with low-virulence ing on the pathogenicity of the virus used for the infection viruses. Microscopically, the lungs showed inflammation (11,14,15,30). In general, those viruses that have been and edema surrounding bronchioles and spilling into the recovered from patients with fatal infections (i.e., A/Hong alveolar septae (11,15,30). Histopathology and flow cytom- Kong/483/97) tend to produce more severe disease in the etry have shown that the inflammatory infiltrate consisted different animal models than those recovered from patients of neutrophils and macrophages (14). The extent of inflam- with milder disease (i.e., A/HongKong/486/97) (11,30). mation spilling into the alveoli appeared earlier (about
660 Guarner and Falcón-Escobedo/ Archives of Medical Research 40 (2009) 655e661
5e9 days postinfection) and was more pronounced in those epithelium and inflammation of the mucosa of the airways
animals infected with highly virulent virus compared to (nasal, paranasal, sinuses, trachea, bronchus, and bronchi-
those infected with low-virulence virus. Similarly, the pres- oles) starting at 1 day postinfection. Changes in the respira-
ence of epithelial necrosis has been described for those tory mucosa are accompanied by the presence of viral
animals infected with highly virulent virus. At about 3 days antigens that decline sharply 3 days postinfection. Macro-
postinoculation and before there are inflammatory infil- scopically the lungs appear consolidated, and microscopi-
trates, viral antigens can be detected in type II pneumocytes cally there is neutrophilic and mononuclear inflammation
(15). The brains of animals infected with the highly virulent in the bronchioles and alveolar intestitium. Viral antigens
viruses showed glial nodules, perivascular cuffing by were detected in the epithelial cells and macrophages.
lymphocytes and neutrophils, neuronophagia, and inflam- Outside the lungs, viral antigens have been found in the heart
mation in the choroid plexus more frequently and exten- and thymus. In pigs, lesions produced by seasonal H1N1
sively than those animals infected with low-virulence were more extensive than those produced by infections with
viruses (11,30). Viral antigens have been detected in the H3N2 virus (10).
neurons.
Comparison of the Pathological Features Caused by
Pathology of H3N2 Influenza A Viruses
Publications of the pathology observed in patients infected The pathology caused by these viruses in humans or animal
with seasonal or interpandemic influenza have included models depends on the virulence of the infecting agent and
51 pediatric patients with fatal disease and 11 adults (six with host response. All the viruses infect the respiratory epithe-
fatal disease and five who survived the infection) (6,7,31,32). lium from the nasal passages to bronchioles; however, high-
The pathological features described for H3N2 have a bias virulence viruses (1918 H1N1, H5N1, and probably 2009
towards severe and complicated infections that have required H1N1) also tend to infect pneumocytes and intraalveolar
open lung biopsies or were fatal, and an autopsy was per- macrophages. The localization and amount of the inflam-
formed to understand the cause of death. The pathological matory reaction will depend on the infected cells. Thus,
changes observed in surviving patients have ranged from low-virulence viruses (seasonal H3N2 and H1N1) cause
acute lung injury including patchy fibrinous alveolar primarily inflammation, congestion and epithelial necrosis
exudates, hyaline membrane formation, interstitial edema, of the larger airways (trachea, bronchi and bronchioles),
and necrosis of bronchiolar mucosa to reparative changes whereas high-virulence viruses have equal inflammation
such as proliferation of type II pneumocytes, interstitial in these localizations with additional, more extensive areas
chronic inflammatory infiltrates, and organization of of inflammation in the alveoli. In susceptible individuals,
airspaces and interstitium (31,32). inflammation of the alveolar walls will result in diffuse
In autopsies of patients infected with the H3N2 virus, the alveolar damage. It needs to be remembered that influenza
most frequent and sometimes the only pathology observed A viruses do not cause inclusions or visible cytopathic
has been mononuclear inflammation and congestion of the effects in vivo; however, infected cells show reactive
trachea, major bronchi and bronchioles commonly accom- changes with enlargement of the nuclei. In addition, the
panied by necrosis of the epithelium and submucosal pathological picture will change to intraalveolar neutro-
hemorrhage (Figure 3A) (6,7). Viral antigens may only be philic infiltrate if bacterial superinfection is present. The
present in the bronchial epithelium. Changes in the lower frequency and type of bacterial superinfections that we
respiratory tract include mononuclear inflammation in the can expect during the 2009e2010 season will be different
alveolar septa, hyaline membrane formation and intraalveo- from those encountered during the 1918 H1N1 pandemic.
lar hemorrhage (Figure 3B). Viral antigens are rarely found The use of antibiotics, supportive measures (ventilators),
in the alveoli. In 50% of the cases there were focal areas of and vaccinations for S. pneumoniae and Haemophilus influ-
pneumonia with Staphylococcus aureus and group A strep- enzae will probably reduce the frequency of these superin-
tococci being the most frequently identified bacterial super- fections but will bring community and nosocomial bacterial
infections. Lymph nodes showed hemophagocytosis in half infections that will have acquired antibiotic resistance.
the cases. In the heart, patchy areas of mononuclear inflam-
mation and necrosis (Figure 3C) were observed in 30% of
cases.
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