India reported its first case of H1N1 in July 2009 in Pune and since then, the number of reported cases and deaths exploded in India. Since very little data is available about histopathological findings in patients of H1N1 fatal cases in India, a retrospective chart analysis of necropsy findings of 15 cases of 2009 H1N1 fatal cases was performed. Common clinical features were fever, cough , and breathlessness followed by sore throat and rhinorrhea. Common lung findings were mononuclear cell infiltration, thick alveolar septae, intraalveolar hemorrhage . The other findings were congested pulmonary blood vessels, pulmonary edema, cytomegaly, fibrin accumulation and formation of eosinophilic membrane. These findings are suggestive of diffuse alveolar damage ( DAD) and DAD with hemorrhage. All patients who underwent necropsy had radiographic findings suggestive of unilobar or multilobar pneumonia. This clinical finding can be correlated pathologically in these patients as all of them had either polymorphonuclear or mononuclear infiltrate. Furthermore, necrotizing pneumonitis pattern seen on these patients is the likely cause of mortality in these patients. Although clinical ARDS pattern was noted in all these patients, it was well correlated in lung pathology in all these cases

Basavraj, A.

Bharadwaj, R.

Borse, R. T.

Ghorpade, S. V.

Jamkar, A. V.

Kadam, D. B.

Mave, Vidya

Mishra, A. C.

Prasad, H. B.

Puranik, S. C.

Sangle, S. A.

Umarji, P. B.

English

Publications of the IAS Fellows

498  © JAPI • August 2011 • VOL. 59
Original Article
Retrospective Analysis of Necropsy Findings in Patients 
of H1N1 and their Correlation to Clinical Features
HB Prasad*, SC Puranik**, DB Kadam***, SA Sangle†, RT Borse†, A Basavraj†, PB Umarji†,  
Vidya Mave‡, SV Ghorpade#, R Bharadwaj$, AV Jamkar%, AC Mishra^
*Associate professor Medicine, **Professor of Pathology, ***Professor of 
Medicine, †Associate Professor Medicine, ‡Director, BJ Medical College 
Clinical Research site, #Professor and Head Department of  Chest and 
tB, $Professor and Head Department of  Microbiology, %Dean, B.J. 
.Medical College, Pune, ^ Director, National Institute of  Virology, Pune
Received: 27.04.2010; Accepted: 10.12.2010
Abstract
India reported its first case of H1N1 in July 2009 in Pune and since then, the number of reported cases and deaths 
exploded in India. since very little data is available about histopathological findings in patients of H1N1 fatal cases 
in India, a retrospective chart analysis of necropsy findings of  15 cases of 2009 H1N1 fatal cases was performed. 
Common clinical features were fever, cough , and breathlessness followed by sore throat and rhinorrhea. 
Common lung findings were mononuclear cell infiltration, thick alveolar septae, intraalveolar hemorrhage . the 
other findings were congested pulmonary blood vessels, pulmonary edema, cytomegaly, fibrin accumulation 
and formation of eosinophilic membrane. these findings are suggestive of diffuse alveolar damage ( DAD) and 
DAD with hemorrhage.  All patients who underwent necropsy had radiographic findings suggestive of unilobar 
or multilobar pneumonia. this clinical finding can be correlated pathologically in these patients as all of them 
had either polymorphonuclear or mononuclear infiltrate. Furthermore, necrotizing pneumonitis  pattern  seen 
on these patients is the likely cause of mortality in these patients. Although clinical ARDs pattern was noted in 
all these patients, it was well correlated in lung pathology in all these cases. 
Introduction 
the first two cases of H1N1 influenza were  confirmed by the centers of disease control and prevention (CDC) on April 21, 
2009 in children from California who reported febrile respiratory 
illnesses.1  When the Mexican Ministry of Health reported an 
increase in severe pneumonia cases and deaths in young adults 
in April of 2009, CDC confirmed that it is due to the same strain 
of the virus.2,3  On June 11, 2009, the World Health Organization 
raised the pandemic alert level to phase 6, indicating that a global 
pandemic had began.4 As of september 20, 2009, human infection 
with 2009 H1N1 virus had been identified in 191 countries and 
territories.5
A recent meta-analysis showed that 2009 pandemic influenza 
has affected 30% of children less than 18 years and has resulted 
in ICu admissions and deaths in 35% and 13% of patients 
respectively.6  Evaluations by CDC of lung specimens of 77 fatal 
cases revealed that 29% had a bacterial co-infection.7  A recent 
autopsy report from Brazil also showed that in fatal H1N1 cases, 
pathology is restricted to lungs.8 
India reported its first case of H1N1 in July 2009 in Pune and 
since then, the number of reported cases and deaths exploded in 
India. Since very little data is available about histopathological 
findings in patients of H1N1 fatal cases in India, a retrospective 
chart analysis of necropsy findings of   fifteen  fatal  cases of 
2009 H1N1  was performed.  
Methods
We conducted a retrospective chart analysis of all 15   fatal 
cases of H1N1 influenza over the age of 12 years who had 
necropsy performed between August 2009 and November 2009 
in BJ Medical College (BJMC) and sassoon general Hospital 
(SGH), Pune, India. Institutional ethics committee approval 
was obtained. 
Data Collection
All clinical data were collected by reviewing the medical 
charts from BJMC-sgH. We collected demographic and clinical 
characteristics of all fatal cases of H1N1 influenzae during the 
study period. A person was defined as infected with 2009 H1N1 
influenza based on laboratory confirmation of the presence of 
H1N1 specific viral nucleic acid in a sputum or nasopharyngeal 
specimen collected at any time during hospitalization.  All patient 
specimens were sent to the National Institute of Virology (NIV), 
a national reference virology laboratory in Pune, India, where 
reverse-transcriptase PCR assay was performed, according to the 
protocol recommended by the u.s. Centers for Disease Control 
and Prevention (CDC).9
Tissue Preparation
Necropsy was performed in BJMC-sgH pathology 
department. Both lung and liver tissue was processed to identify 
pathology. tissue fragments were formalin-fixed, paraffin-
embedded, and hematoxylin and eosin–stained.
Results
Fifteen patients died during the study period and necropsy 
of lung and liver were performed. the demographic and 
clinical characteristics of these patients are shown in table 1. 
the median age was 34 years (range13-50 years), and 40% were 
females and 60% males. sixty percent of patients had one or 
more of co-morbidities. the most common co-morbidity noted 
in fatal cases were pregnancy, obesity, Rheumatic Valvular Heart 
Disease (RVHD), Chronic Obstructive  lung disease (COPD), and 
Cerebrovascular Accident (CVA).  the most common symptoms 
at presentation in decreasing order were fever, cough and 
shortness of breath. The most common radiologic findings were 
fluffy infiltrates  followed by confluent opacities. Most of the 
patients had PO2 <60 on admission and 2 patients had PO2>60 
while PO2/FiO2 ratio in all patients was less than 150 suggestive 
of (acute  respiratory  distress syndrome) ARDs.
All  these  patients   were  mechanically  ventilated, 40%  of 
patients died within 48 hours of hospitalization while 60% of 
patients were mechanically ventilated for >48 hours before death. 
© JAPI • August 2011 • VOL. 59    499
the changes on patients who died within 1-4 days of 
hospitalization were mononuclear cell infiltration, congested 
pulmonary blood vessels, large giant cells, pulmonary edema, 
fibrin accumulation, desquamation and ARDs. Common 
histopathological findings beyond 4 days of hospitalization 
were thick alveolar septae, lung collapse, polymorphonuclear 
cell infiltration.
upon comparing the clinical features with necropsy, 33% 
patients who presented with hemoptysis also had intraalveolar 
haemorrhage. All patients had mono or polymorpho nuclear 
infiltrates consistent with pneumonia on radiography. All the 
cases showed ARDs on blood gas examination, with involvement 
of more than 3 segments on radiography in 80% of cases but on 
necropsy only 2 patients had ARDs. 
the median duration of symptoms before death was 9 days 
(range, 2-12days). All patients were admitted to intensive care 
unit and were receiving antiviral and combination antibiotics. 
the most common complication noted was  ARDs.  
The necropsy findings of these patients are shown in Table 
2. Common findings were mononuclear cell infiltration, thick 
alveolar septae, and intraalveolar haemorrhage (Figs. 1 and 
2) and were seen in 80% of patients. The other findings were 
congested pulmonary blood vessels, pulmonary edema, 
cytomegaly, fibrin accumulation and formation of eosinophilic 
membrane. These findings were suggestive of diffuse alveolar 
damage (DAD) and were seen in all patients.  Acute respiratory 
distress syndrome (ARDs) and desquamation was seen in two 
patients. Liver tissue showed fatty changes and portal triaditis 
in all  patients (Fig. 3).
Table 1 : Demographic and clinical characteristics of 15 fatal 
H1N1 cases from Pune, India
Characteristic Number (%) N=15
Female sex  4 (40%)
Median age 34
Ethnicity
Hindu  15(100%)
Muslim NONE
Co-morbidity 9(60%)
Pregnancy 4 (26,66%)
COPD 1  ( 6.6%)
Obesity 1 ( 6.6%)
Cerebrovascular event  1 ( 6.6%)
Rheumatic valvular disease 1 (6.6%)
HIV/AIDs 1  (6.6%)
Symptoms on admission
Fever 14  (93.33%)
Cough 14 (93.33%)
Rhinorrhea 2 (13.33%)
shortness of breath  6(40%)
sore throat  3(20%)
Bodyache 4(26.66%)
Radiologic Findings
Unilobar infiltrates 3(20%)
Multilobar infiltrates 5(33.33%)
Hospital Course
Mechanical ventilation 15(100%)
Intensive care unit admission 15(100%)
Antiviral treatment within 48 hours 15(100%)
Antibiotic treatment on admission 15(100%)
Renal Failure None
Fig. 3 : Liver showing fatty change and portal triaditis
Fig. 1 : Lung alveoli showing interstitial mononuclear infiltration 
with thickened alveolar septae (Interstitial pneumonitis)
Fig 2 : Lung alveoli showing hyaline membrane formation and 
pulmonary edema.
Table 2 : Common histopathological findings on necropsy
Histopathological  Finding Number of Patients (%)
Mononuclear  infiltrates 12(80)
thick  alveolar  septae 12(80)
Intraalveolar   hemorrhage 12(80)
Polymorphonucler infiltration 9(60)
Pulmonary  edema 5(33.33)
Congested  pulmonary  vessels 3(20)
Collapse  of  alveoli 3(20)
Fibrin  accumulation 3(20)
Hyaline  membrane  formation 2(13.33)
Large  giant cells 2(13.33)
Cytomegaly 2(13.33)
Eosinophilic membrane 2(13.33)
ARDs 2(13.33)
500  © JAPI • August 2011 • VOL. 59
Discussion
Our report on necropsy findings of fatal cases of H1N1 
influenzae shows that these patients had mononuclear cell 
infiltration, thick alveolar septae, and intra alveolar haemorrhage. 
The other findings were congested pulmonary blood vessels, 
pulmonary edema, cytomegaly, fibrin accumulation and 
formation of eosinophilic membrane. these findings was 
suggestive of diffuse alveolar damage (DAD). This report is the 
first report from India describing lung and liver pathology in 
patients who died secondary to 2009 H1N1.
the median age of our fatal cases were 34 years and is similar 
to prior reports. the clinical features of these patients were 
comparable with previous reports of 2009 H1N1influenza.10-12 
In consistent with previous reports, 60% of our fatal cases had 
a co-existing condition.  Majority of our patients showed clinical 
picture consistent with ARDs.
Viral infections can cause bronchitis, bronchiolitis and 
pneumonia. Most viruses that cause pneumonia infect epithelial 
cells exclusively or in addition to infecting  other  cells. the 
predominant tissue response is interstitial pneumonia.  the walls 
of air spaces are thickened by edema, congestion and an infiltrate 
of mononuclear cells. The  air spaces  contain edema, fibrin 
and  scanty  cellular  exudates  of   monocytes,  macrophages, 
occasional  neutrophils  and  sloughed  epithelial  cells.13 
In fatal cases, hyaline membrane are usually present  along 
alveolar  spaces.  Alveolar epithelial cells are  enlarged  and 
hyperplastic,  Patients  who have  been ill for some days  show 
signs of repair   and  organization similar  to  organizing  bacterial 
pneumonia  such as  exudate in alveolar  space  being  invaded 
by  fibroblasts  and converted to buds  of fibrous tissue  similar 
to  bacterial  pneumonia.  Foci of hyperplasia and metaplasia to 
a squammous or bronchiolar type are found in alveoli. Discrete 
foci of necrosis are also seen.10   
the present study shows that H1N1 caused extensive 
involvement of the lungs, and the changes range from 
mononuclear cell infiltration to formation of hyaline membrane 
and thick alveolar septae. These findings are suggestive of 
diffuse alveolar damage (DAD).  As a result most of the patients 
presented with extensive pneumonia and ARDS. These findings 
were also reported in previous autopsy report from Brazil as well 
as previous pandemics.8,14-19 the liver changes noted in our series 
are mainly limited to fatty changes and portal triaditis which is 
not a primary consequence of viral infection.
In our series, 4 women were pregnant and showed evidence 
of extensive lung involvement. the major pathology noted was 
same as non-pregnant patients. these changes were also noted 
in prior reports.8 
All patients who underwent necropsy had radiographic 
findings suggestive of unilobar or multilobar pneumonia. This 
clinical finding can be correlated pathologically in these patients 
as all of them had either polymorphonuclear or mononuclear 
infiltrate. Furthermore, necrotizing pneumonitis pattern seen on 
these patients is the likely cause of mortality in these patients. 
Although clinical ARDS pattern was noted in all these patients, 
it was  not  well correlated in lung pathology in all these cases. 
the main limitation of our study is that we do not have data 
on the presence of bacterial co-infection on necropsy specimens. 
However, the correlation between presence of neutrophils and 
bacterial infection is established.18 the other main limitation is 
that we did not perform complete autopsy of these fatal cases. 
since the study is retrospective in nature thus clinical comparison 
may be limited and biased. Despite these limitations, to our 
knowledge, our study is the first study describing the lung 
pathology in adult H1N1 fatal cases in India.
In summary, the extensive lung involvement in H1N1 
infection is the likely cause of mortality in adult H1N1 cases. 
Our report adds to the current literature on natural progression 
of H1N1 infections as well as lung pathology in fatal H1N1 
infection cases.  
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Retrospective analysis of necropsy findings in patients of H1N1 and their correlation to clinical features

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Retrospective analysis of necropsy findings in patients of H1N1 and their correlation to clinical features

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