The present systematic review was carried out according to the Preferred Reporting Items for Systematic review and Meta-Analyses (PRISMA) standards [5].

A systematic literature search selection (Fig. 1) was conducted on PubMed database on June 5, 2021, with this search strategy: (COVID-19) AND (H1N1 influenza) showing 101 results. To narrow this search, the following inclusion criteria were selected: English language; published in a scholarly peer-reviewed journal; full-length articles were further elected. Another useful criterion to further refine the research was to focus on the type of manuscript: review, systematic review, and meta-analysis (Table 1).

Fig. 1.

Flow of the systematic review, according to PRISMA guidelines.

A critical appraisal of the collected studies was conducted, analyzing titles and abstracts, as well as a hand search of reference lists were carried out by two researchers (GB and RLR) excluding the following topics: treatment, public health measures and perception of the general population or healthcare personnel about their quality of life. Data extraction was verified by another investigator (AM). According to these procedures, 54 eligible studies were included in the present review.

This systematic review concerns two pandemic events that occurred 10 years apart. Thus, it includes studies that were published in a time frame of 10 years. Therefore, study results should be interpreted taking into account that the accuracy of clinical and diagnostic procedure have changed over the years. As far as SARS-CoV-2 is concerned, it should be known that the dominant variant at the time of the study was: delta.

The H1N1-09 pandemic began in March 2009 in Mexico with subsequent diffusion in the US and then throughout the world until August 2010, with approximately 110 countries involved [1, 6, 7]. At the end of H1N1-09 pandemic, WHO confirmed 18,500 deaths, although, according to the study by Dawood et al. [8] these numbers were underestimated as the total number of deaths ranged between 151,700 and 575,400. The H1N1-09 virus continues to circulate in humans seasonally [9]. The current COVID-19 pandemic began in November-December 2019 in Wuhan, China, when several cases of pneumonia of unknown etiology were found. It subsequently spread to more than 203 countries and territories. As of 4 June 2021, a total of 171,708,011 cases worldwide have been confirmed since the start of the pandemic and 3,967,151 deaths and 33.689 deaths in Italy [10].

H1N1-09 virus infection (viral taxonomy: Riboviria \guilsinglright Orthornavirae \guilsinglright Negarnaviricota \guilsinglright Polyploviricotin\guilsinglright Insthoviricetes \guilsinglright Articulavirales \guilsinglright Orthomyxoviridae \guilsinglright Alphainfluenzavirus \guilsinglright Influenza A virus pH1N1) was a new strain of virus type A originating from the human, avian and pigvirus groups. Immediately after sequencing, the clinical condition generated by contact with this virus took the name of “swine flu”, because the viral strain was probably transmitted from pigs to humans, although pigs were not involved in the worldwide diffusion of the virus during the pandemic [11, 12, 13, 14]. As regards the current pandemic, the etiological agent, whose genomic sequence was closely related to the SARS-CoV of 2003, has been identified as SARS-CoV-2 (viral taxonomy: Riboviria \guilsinglright Orthornavirae \guilsinglright Pisuviricota \guilsinglright Pisoniviricetes \guilsinglright Nidovirales \guilsinglright Cornidovirineae \guilsinglright Coronaviridae \guilsinglright Orthocoronavirinae \guilsinglright Betacoronavirus \guilsinglrightSarbecovirus \guilsinglright severe acute respiratory syndrome-related coronavirus-2) [12]. This virus likely originated in bats but may have been amplified in an intermediate host before transmission to humans represented by pangolins [13, 14, 15].

During pH1N1, the most affected age group included children and young adults with only 5% of cases relating to adults over the age of 51 [1, 16, 17], possibly due to partial immunity to the virus in the elderly population. Similarly, it was observed that the swine flu hospitalization rate decreases as the age of patients increases. The age group most affected by COVID-19 is adults over 40 ($>$70%), while only 10% of cases are under the age of 30 [1, 15]. The COVID-19 hospitalization rate thus increases based on the age of the patients. This positive association shows that individuals over the age of 85 have the highest hospitalization and death rates [18].

As for pathogen transmission efficiency, influenza viruses and coronaviruses are both effective in causing respiratory diseases because they spread easily among humans through oral and nasal droplets. Furthermore, they can also be transmitted through indirect contact with infected surfaces (fomites) [19, 20, 21].

H1N1-09 is an enveloped-spherical virus, 80–120-nm in diameter, with a negative-polarity single-strand RNA, consisting of 8 segments coding for 12–14 proteins. The influenza virus infects the epithelial cells that line the upper respiratory tract (including the nasal tract) to the lower respiratory tract (up to the alveoli). Access to the cells of the upper respiratory tract is determined by the presence of sites rich in sialic acids, to which the hemagglutinin proteins present in human influenza viruses bind. These sites are particularly expressed in the soft palate [22]. The main mechanism of influenza pathophysiology is the result of lung inflammation and impairment caused by direct viral infection of the respiratory epithelium, combined with the effects of lung inflammation caused by host-triggered immune responses that limit the spread of the virus. Alveolar macrophages and endothelial cells appear to play a key role, as inducing exposure of cytokines and viral antigens to the endothelial layer can amplify inflammation, with endothelial cells constituting a major source of pro-inflammatory cytokines. Inflammation can thus progress and spread systemically and manifest itself as multi-organ failure, but these consequences are generally occurred after severe pulmonary impairment. The inability of the lung to perform its primary gas exchange function occurs through mechanisms of airway obstruction, loss of alveolar structure, loss of pulmonary epithelial integrity, and degradation of the extracellular matrix that maintains the structure of the lung [23, 24].

The COVID-19 virus also has the characteristic of being enveloped and spherical, with a diameter of about 120 nm and a positive polarity single-strand RNA, consisting of 1 segment coding for 15 non-structural proteins, 4 structural proteins (spike, envelope, membrane glycoproteins, and nucleocapsid protein), 8 accessory proteins [25]. For entry into the host cell, this species of human coronavirus binds to the angiotensin 2 converting enzyme (ACE2); individual variation in the expression and/or polymorphisms of ACE2 can influence susceptibility to infection by the COVID-19 phenotype [26, 27]. This interaction then triggers ACE2 endocytosis along with the COVID-19 virion and fusion of the viral membrane and host cell. Simultaneously, the viral spike protein is exposed to endosomal proteases which lead to its cleavage at two different sites: the first removes the S1 subunit, while the second occurs within the S2 subunit and causes exposure of the fusion peptide. The viral packet is thus released into the host cytoplasm, where it usurps the cellular mechanism producing new viral particles. In this perspective, lung tissue represents the ideal candidate for virus action due to its large surface area, which makes it highly susceptible to inhaled viruses and the conspicuous expression of ACE2 by Type II alveolar epithelial cells (pneumocytes) [28].

The development and progression of COVID-19 continue with major pathological mechanisms such as direct virus-induced cytotoxicity in ACE2-expressing cells, dysregulation of the RAAS (renin-angiotensin-aldosterone system) as a consequence of virus-mediated ACE2 down-regulation, dysregulation of immune responses, endothelial cell injury and thrombosis and fibrosis. This is because of the internalization of the virion-ACE2 promoting the accumulation of Ang II, with consequent production of receptors for TNF-$\alpha$ and IL-6 and activation of macrophages in a pro-inflammatory state, which evolves towards the well-known clinical condition of “cytokine storm” [29]. Furthermore, the virus nucleocapsid protein can interact with Smad3 to prevent apoptosis of infected host cells, promoting tissue fibrosis mediated by transforming growth factor (TGF)-$\beta$ [30]. Type II alveolar epithelial cells renew themselves autonomously and express high levels of ACE2 and are thus constantly oriented towards viral entry and replication, which induces a vicious cycle of tissue damage and repair that can eventually result in areas being replaced, in turn responsible for the exchange of gases in non-functional fibrotic tissue.

For pH1N1, the incubation period was 1.5–3 days, while the incubation period for COVID-19 is usually longer (2–14 days), averaging 5.2 days [31, 32]. In both cases, fever and respiratory symptoms are the dominant clinical picture; dyspnea affects subjects with severe COVID-19 and pandemic H1N1-09 indifferently, followed by cough, fatigue, myalgia, arthralgia, and headache. Rhinorrhea, sore throat, thoracic pain, and sputum production were more common during the H1N1-09 pandemic, whereas dry cough, diarrhea, and vomit were more frequent among COVID-19 patients [33].

Clinical specimens including a throat or nasopharyngeal swab, saliva, or lower respiratory tract aspirate sample are indicated for laboratory diagnosis of H1N1-09 and COVID-19 infections [34, 35, 36]. Additionally, whole blood or serum/plasma samples can be collected for seroconversion assessment. Rapid techniques for detecting the influenza virus include immunofluorescence and enzyme immunoassays. For both viruses, RT-PCR is the gold standard technique.

At imaging, the most significant differences between the two pathological pictures were more evident on CT examinations, where linear opacification, pleural thickening, vascular enlargement, and crazy-paving signs, are more severe with COVID-19 pneumonia, while bronchiectasis and pleural effusion, are more common in patients with H1N1-09 pneumonia. Other imaging findings, including peripheral or peribronchial and perivascular distribution, ground-glass opacity (GGO), consolidation, subpleural line, air bronchogram, and bronchial distortion, were not significantly different between the two patient groups [37, 38, 39, 40] (Fig. 2, Ref. [40]).

Fig. 2.

Radiological pattern comparison. (A–C) with red arrow, the “crazy paving” sign. It consists of scattered or diffuse ground-glass attenuation with superimposed interlobular septal thickening and intralobular lines, the typical pattern among others of pathologies which pattern includes Pneumocystis carinii pneumonia, mucinous bronchioloalveolar carcinoma, pulmonary alveolar proteinosis, sarcoidosis, nonspecific interstitial pneumonia, organizing pneumonia, exogenous lipoid pneumonia [40]. (B–D) GGO pattern.

Regarding the radiological diagnosis of COVID-19, however, further clarification is required, as several studies have reported sensitivity of RT-PCR tests for SARS-CoV-2 between 37% and 83%, while chest sensitivity to CT for COVID-19 has been reported between 80% and 90% and specificity between 82.9% and 96% [41]. In this respect, in March 2020 a standardized disease diagnosis-probability system was proposed, the COVID-19 Reporting and Data System (CO-RADS), with a range that goes from CO-RADS 1 (COVID-19 is highly unlikely. CT is normal or findings indicate non-infectious diseases such as congestive heart failure, sarcoidosis, histoplasmosis, neoplasm, UIP or fibrotic NSIP) to CO-RADS 6 (Patient with PCR positive and bilateral GGO) [42].

Histopathological analysis was performed on formalin-fixed paraffin-embedded (FFPE) lung samples collected by the same operator from two corpses with similar characteristics (Case 1: male between 50–60 years; mute history; positive swab for COVID-19 after entering the ER; died shortly after arriving at the hospital; Case 2: male between 50–60 years; mute history; positive swab for H1N1-09 after access to the emergency room; died 48 hours after entering the local hospital) during autoptic procedures performed at the Section of Legal Medicine, University of Foggia (Fig. 3).

Fig. 3.

Macroscopic findings. COVID-19: massive pulmonary edema at parenchymal level (A) and at airway opening (C). H1N1-09 (B–D): the forceps indicate widespread subpleural hemorrhages.

Standard hematoxylin and eosin (H&E) staining and modified Masson’s trichrome stain [40] were performed (Fig. 4). For the immunohistochemical staining, after a first phase common to all the antibodies used and consisting in the preliminary pre-treatment for antigenic unmasking, subsequent application of the primary anti-staining antibody with Mayer’s hematoxylin, the following anti-human antibodies were used: anti-nucleocapsid anti-COVID (anti Coronavirus -FIPV3-70 Santa Cruz Biotechnology, Inc., Dallas, TX, USA); anti-human fibrinogen (Dako A/S, Glostrup, Denmark); anti-human CD61 (Dako A/S, Glostrup, Denmark); anti-IL6 (Santa Cruz Biotechnology, Inc., Dallas, TX, USA); anti-HIF-1a.

Fig. 4.

Histopathological H&E staining and modified Masson’s trichrome stain comparison. COVID-19 demonstrated the subacute organizing phase of DAD (A–G, 10$\times$) with microthrombi (C–E, 20$\times$) and H1N1-09 showed the acute or exudative phase with hyaline membranes (B, 20$\times$) with edema (B, 10$\times$–D, 20$\times$–F, 20$\times$), mild interstitial and alveolar inflammatory infiltrates, revealing a more marked involvement of the interstitium by the COVID-19 infection (H, 10$\times$).

Furthermore, a qualitative method was used for the evaluation of each immunohistochemical staining [43, 44, 45].

The strength of the immunohistochemical staining weighed on different investigated areas was classified attributing values as follows: no detectable staining = 0, weak staining = 1, moderate staining = 2, strong staining = 3 (Fig. 5).

Fig. 5.

Immunohistochemical comparison (20$\mathrm{\times }$). COVID-19 showed the following grading: (A) anti-COVID-19 = 2; (C) anti-human fibrinogen = 3; (E) anti-human CD61 = 3; (G) anti-IL6 = 2; (I) anti-HIF-1a = 2. H1N1-09 revealed the subsequent grading: (B) anti-COVID-19 = 0; (D) anti-human fibrinogen = 0; (F) anti-human CD61 = 0; (H) anti-IL6 = 1; (J) anti-HIF-1a = 0. These findings demonstrate that COVID-19 infection is associated with a greater extent of thromboinflammation than H1N1-09. This condition, which can be explained by a more marked tropism of the virus for endothelial cells, would in turn explain the deeper cellular hypoxia induced in cases of COVID-19 infection, demonstrated by the positivity to HIF-1a.

Diffuse Alveolar Damage (DAD) is the typical finding of the histological analysis of viral ARDS induced by COVID-19 and H1N1-09 and represents the severe form of the Acute Lung Injury (ALI) spectrum. DAD is caused by “lesions of the cells of the endothelial and alveolar lining that led to the exudation of liquids and cells”, which culminates in the physical destruction of the alveolo-capillary membrane and, thus, in the inability to exchange gas [46].

Samples from autopsies conducted on individuals who died from H1N1-09 and COVID-19, demonstrated that the acute or exudative phase of DAD was the predominant pulmonary histological picture (hyaline membranes with edema, mild interstitial inflammatory infiltrates and pneumocytes desquamated with reactive hyperplasia of pneumocytes) [47, 48, 49].

In detail, the histological changes most frequently found in the lungs during the H1N1-09 pandemic were intra-alveolar inflammatory infiltrates, consisting of macrophages, polymorphonuclear cells and lymphocytes scattered between areas of edema, hemorrhage and fibrin deposits. By contrast, inflammatory infiltrates found in the lungs of COVID-19 patients were dominated by macrophages, associated with thickening of the alveolar walls and partial loss of histological architecture [37]. In addition, in just over half the COVID-19 cases, the presence of organizing fibrosis has been described and reported in the histopathological examination, indicating an early transition to the subacute organizing phase. However, the important almost pathognomonic, feature of lung damage in COVID-19 patients is the presence of microthrombi [50, 51], resulting in speculation that COVID-19 has a predilection for endothelial cells. Vascular thrombosis and micro-thrombosis are frequent findings in DAD, resulting from local inflammation even in the absence of a state of systemic hypercoagulability, which occurs early on in ARDS of various causes, but would be more verifiable in cases of COVID-19 infection (Figs. 4,5). According to some authors, this phenomenon is nine times greater and it is associated with phenomena of neovascularization [52] and hemorrhages with different severity ranging [53]. Perivascular inflammation was also described as defining feature of the virus-induced systemic disease [54, 55].