Physiological and molecular triggers

for SARS-CoV membrane fusion and

entry into host cells Jean Kaoru Millet
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 Virology 517 (2018) 3–8

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Virology
journal homepage: www.elsevier.com/locate/virology

Physiological and molecular triggers for SARS-CoV membrane fusion and T

entry into host cells
⁎
Jean Kaoru Millet, Gary R. Whittaker
Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, NY 14853, United States

A R T I C L E I N F O A B S T R A C T

Keywords: During viral entry, enveloped viruses require the fusion of their lipid envelope with host cell membranes. For
Coronavirus coronaviruses, this critical step is governed by the virally-encoded spike (S) protein, a class I viral fusion protein
SARS that has several unique features. Coronavirus entry is unusual in that it is often biphasic in nature, and can occur
Spike protein at or near the cell surface or in late endosomes. Recent advances in structural, biochemical and molecular
Virus entry
biology of the coronavirus S protein has shed light on the intricacies of coronavirus entry, in particular the
Endosomes
Calcium
molecular triggers of coronavirus S-mediated membrane fusion. Furthermore, characterization of the cor-
Fusion peptide onavirus fusion peptide (FP), the segment of the fusion protein that inserts to a target lipid bilayer during
membrane fusion, has revealed its particular attributes which imparts some of the unusual properties of the S
protein, such as Ca2+-dependency. These unusual characteristics can explain at least in part the biphasic nature
of coronavirus entry. In this review, using severe acute respiratory syndrome coronavirus (SARS-CoV) as model
virus, we give an overview of advances in research on the coronavirus fusion peptide with an emphasis on its role
and properties within the biological context of host cell entry.

1. Introduction process, often requiring a specific sequence of interactions (binding to

Coronaviruses are a diverse group of single-stranded plus-sense RNA
viruses belonging to the Coronaviridae family and Nidovirales order (de
Groot et al., 2012). They infect a wide array of mammalian and avian
species, including bats, and have a propensity for interspecies jumping
and zoonotic transmission as exemplified by severe acute respiratory
syndrome coronavirus (SARS-CoV) and more recently by Middle East
respiratory syndrome coronavirus (MERS-CoV) (Graham et al., 2013;
Woo et al., 2009). As coronaviruses possess an envelope, membrane
fusion with host cell membranes is a required and critical step in the
replication cycle allowing for delivery of genomic RNA into the cyto-
plasm, which eventually leads to the start of replication. This critical
early step is being viewed as an attractive target for therapeutic inter-
ventions (White et al., 2008).
Virus entry constitutes a series of interactions between a virion and
its host cell occurring early in the viral life cycle (Boulant et al., 2015;
Grove and Marsh, 2011; Hofmann and Pöhlmann, 2004; Marsh and
Helenius, 2006). For an enveloped virus, these steps allow the virus to
(i) bind to a target host cell, typically via interactions with cellular
receptors, (ii) fuse its envelope with a cellular membrane, either at the
plasma membrane or through the endocytic pathway, and (iii) deliver
its genetic material inside the cell. Virus entry is a finely regulated
receptors and/or co-receptors), triggers or cues (pH, proteolytic acti-
vation), and cellular processes such as endocytosis for successful de-
livery of viral genomic nucleic acids (White and Whittaker, 2016). In-
terestingly, coronaviruses display a large degree of plasticity regarding
the entry pathways they use, which can occur at the plasma membrane
or through the endocytic pathway (Belouzard et al., 2012; Matsuyama
et al., 2005; Nash and Buchmeier, 1997). The mechanisms by which
coronaviruses enter cells depends on the strain and species considered,
along with tissue and cell-type specificities (e.g. receptor and protease
availability, local microenvironment).
For enveloped viruses, a critical player in the entry process is the
viral fusion protein as it mediates the membrane fusion reaction
(Chernomordik and Kozlov, 2008; Colman and Lawrence, 2003;
Harrison, 2008; White et al., 2008; White and Whittaker, 2016). While
the process of merging two distinct lipid bilayers into a single one is a
thermodynamically favorable reaction, it is associated with a high ki-
netic barrier (Harrison, 2008). As such, and because viral fusion pro-
teins facilitate this process, they can be viewed as catalysts for the
membrane fusion reaction. This has been very well studied both
structurally and functionally with the influenza hemagglutinin (HA)
fusion protein (Harrison, 2015). After attachment of HA to sialic acids
cellular receptors the virion is internalized through the endocytic

⁎
Correspondence to: Department of Microbiology & Immunology, C4-127 VMC, Cornell University, Ithaca, NY 14853, United States.
E-mail address: gary.whittaker@cornell.edu (G.R. Whittaker).

https://doi.org/10.1016/j.virol.2017.12.015
Received 31 October 2017; Received in revised form 13 December 2017; Accepted 15 December 2017
Available online 21 December 2017
0042-6822/ © 2017 Elsevier Inc. All rights reserved.
J.K. Millet, G.R. Whittaker
Fig. 1. Structural model of the SARS-CoV spike in pre-fusion uncleaved and monomeric form, based on pdb 3JCL (MHV spike). SARS-CoV S is colored with the S1 domain in blue and
the S2 domain in orange, and is shown in in three different representations: space-filling, surface mesh and cartoon. The fusion peptide/predicted neutralizing epitope is shown green. The
two proteolytic cleavage sites are shown in magenta, with S1/S2 exposed and the fusion peptide-proximal S2′ site protected. An enlarged cartoon depiction of the fusion peptide region is
shown with key negatively charged and hydrophobic residues indicated.
pathway. Because of endosomal acidification, increased H+ ion con-
centration within the endosome forces HA to undergo major con-
formational changes, which allows exposure of the fusion peptide and
its insertion into target cellular membrane. This brings the viral and
endosomal membrane in close proximity. Further conformational
changes of several HA trimers allow merging of the outermost lipid
leaflets forming an intermediate structure called the hemifusion stalk.
This transient structure collapses into an expanding fusion pore al-
lowing release of viral genetic material in the cytoplasm.
In the case of coronaviruses, viral entry into target cells is performed
by the spike (S) envelope glycoprotein, which mediates both host cell
receptor binding and membrane fusion. The S protein is classified as a
class I viral fusion protein (Bosch et al., 2003), which includes the
prototypical influenza virus hemagglutinin (HA) and retrovirus en-
velope (env) proteins (White et al., 2008). Class I viral fusion proteins
form trimers and each monomer can often be divided into two domains,
a receptor-binding domain (e.g. HA1 and gp120 for influenza virus and
HIV, respectively), and a fusion domain (HA2 and gp40). Fusion do-
mains are enriched in alpha-helices and contain regions called heptad
repeats (HR) which are repetitive heptapeptides containing some hy-
drophobic residues and which are involved in the refolding process and
coiling of central helices during membrane fusion. After membrane
fusion has occurred, class I fusion proteins adopt a compact con-
formation, with a well-defined coiled-coil structure called a 6-helix
bundle or 6HB (Belouzard et al., 2012; White et al., 2008). Importantly,
the fusion domain contains a short segment, the fusion peptide (Epand,
2003; Lai et al., 2005; Tamm and Han, 2000; Tamm et al., 2002), ty-
pically composed of 15–25-amino acids, generally hydrophobic in
nature, which becomes anchored to a target membrane when the fusion
protein adopts the pre-hairpin conformation. The fusion peptide plays
an essential role in mediating the membrane fusion reaction as it di-
rectly interacts with lipid bilayers, enabling to disrupt and connect two
apposing membranes. Class I fusion proteins are often proteolytically
activated or primed for fusion by host cell proteases at a specific clea-
vage site that usually forms the boundary between the receptor-binding
and fusion domains (White and Whittaker, 2016). In the case of influ-
enza HA and HIV env proteins, the cleavage event releases the fusion
peptide as it is located at the N-terminal end of the fusion domain.
While coronavirus S proteins possess salient features of class I fusion
proteins, such as being a type I membrane proteins organized in tri-
mers, possessing heptad repeats regions (HR1 and HR2), and requiring
proteolytic cleavage for activation, they differ in several key aspects.
The S proteins form substantially larger trimers, with S monomer size in
the range of 1200–1400 amino acids (~180–200 kDa) compared to
~500 aa for influenza HA and ~800 aa for HIV. Until recently struc-
tural data on the coronavirus S was limited, due mostly to the difficulty
of obtaining X-ray crystallographic structures of intact ectodomains of
4
Virology 517 (2018) 3–8
S. However, recent advances in cryo-electron microscopy (cryo-EM)
have allowed the determination of the structure covering the majority
of the ectodomain of several coronavirus S proteins in their pre-fusion
conformation such as those of the murine hepatitis virus (MHV) (Walls
et al., 2016a), human coronavirus HCoV-NL63 (Walls et al., 2016b),
HCoV-HKU1 (Kirchdoerfer et al., 2016), MERS-CoV and SARS-CoV (Gui
et al., 2017; Yuan et al., 2017). These efforts and studies constitute a
huge step forward in the field as they uncovered the complexity of
coronavirus S proteins and their highly glycosylated nature. The S
protein can be divided into the S1 receptor-binding subunit and S2
fusion domain, usually separated by a cleavage site (S1/S2). However,
coronavirus S proteins possess an additional cleavage site located
within S2 and called (Belouzard et al., 2009, 2012; Millet and
Whittaker, 2014). Not only are coronaviruses S proteins unusual for
harboring multiple cleavage sites, they are also activated by a wide
variety of host cell proteases (Millet and Whittaker, 2015), spanning
different families such as cathepsins (Simmons et al., 2005), trypsin-like
serine proteases such as members of the transmembrane serine protease
(TTSP) family (Bertram et al., 2013; Gierer et al., 2013; Glowacka et al.,
2011; Matsuyama et al., 2010), and the furin-like proprotein con-
vertases (PCs) (Burkard et al., 2014; Millet and Whittaker, 2014). While
the location of the fusion peptide has been debated, most recent data
appears to argue that the segment immediately downstream of S2′
cleavage site is the bona fide fusion peptide. In the cryo-electron mi-
croscopy structures of coronavirus S, the fusion peptide segment ap-
pears to be exposed at the surface of the protein in the pre-fusion state
(Fig. 1), another unique characteristic setting S proteins apart from
other class I viral fusion proteins like the influenza virus HA. While the
fusion peptides of coronaviruses are not as well characterized as the
ones from other prototypical class I fusion proteins like influenza HA or
HIV env, evidence has been steadily accumulating for the identification
of the bona fide fusion peptide.
In this review, we use SARS-CoV as model for studying the me-
chanism of coronavirus-membrane fusion. We aim to give an overview
of the entry pathway of SARS-CoV and connect this to recent advances
in our understanding of the coronavirus spike fusion peptide, high-
lighting its unique features as well as putting these findings back to
their biological context and the apparently biphasic nature of cor-
onavirus entry. An underlying theme that emerges from these studies is
that the flexible characteristic of coronavirus entry pathways is in some
ways imparted by the unique and special features of the coronavirus
fusion peptide.
2. SARS-CoV mediated entry into cells and the activation of
membrane fusion
As with many viruses, SARS-CoV entry into cells is dictated by the
J.K. Millet, G.R. Whittaker
presence of its receptor. During the 2003–2004 SARS epidemic, a
functional viral receptor for SARS-CoV was rapidly identified using a
biochemical approach, where purified S1 was shown to bind human
angiotensin-converting enzyme (ACE2) (Li et al., 2003). The SARS-
CoV/ACE2 interaction has allowed much to be discovered regarding
SARS-CoV tropism changes (Hulswit et al., 2016) and is considered and
essential part of the virus entry mechanism, along with additional and
binding factors, such as CD209L (Jeffers et al., 2004).
While ACE2 is an essential factor in all cell types, subsequent entry
events are more cell-type dependent. In typical cell culture systems, and
following receptor engagement, SARS-CoV is taken up by both clathrin
and non-clathrin pathways (Inoue et al., 2007; Wang et al., 2008), with
entry blocked by inhibitors of endosomal acidification and endosomal
proteases such as cathepsin L (Simmons et al., 2005, 2004). However,
treatment of cells with trypsin or trypsin-like proteases (TTSPs) can also
result in entry and spike-mediated fusion in a pH-independent manner
(Bertram et al., 2011; Matsuyama et al., 2010, 2005; Simmons et al.,
2004). These findings show that in addition to ACE2, host cell proteases
are key activators of SARS-CoV entry, and while the two identified
activation sites (S1/S2 and S2′) are critical, the process of protease
activation is quite complex, possibly depending heavily on cell type
(Reinke et al., 2017). Overall, it is believed that the TTSP-mediated
pathway is the relevant pathway in primary respiratory epithelia, the
site of SARS-CoV infection in vivo (Shulla et al., 2011). These findings
also suggest that low pH in itself is not a driving force for the necessary
conformational events needed for coronavirus fusion, and that, as
suggested by Matsumaya and Taguchi for MHV-2 (Matsuyama and
Taguchi, 2009), additional factors are involved. Related to this, single
particle studies of entry, using feline coronavirus as a model system,
have shown that effects of pH on the rate constant of coronavirus fusion
are negligible (Costello et al., 2013), with pH-dependency of entry
possibly linked more to the enzymatic activity of the activating pro-
tease. One feature of SARS-CoV is that in cell culture, entry only begins
after a lag time of 30 min (Mingo et al., 2015), suggesting that sub-
stantial endosomal maturation is necessary in the endosomal route of
entry. The finding that wild-type MHV and feline coronavirus infection
of HeLa cells also depends heavily on endosomal maturation, including
components such as the VPS/HOPS complex involved in late endosome-
lysosome trafficking, also suggests that endosome maturation (but not
endosomal acidification per se) is a key player in infection across the
Coronaviridae (Burkard et al., 2014). An overarching model, based
primarily on studies with MERS-CoV, is that coronaviruses can enter
cells either following fusion at or close to the cell surface, the so-called
“early” pathway, or following prolonged endocytic trafficking, the
“late” pathway (Earnest et al., 2017).
3. Fusion peptides form the core of viral fusion protein machinery
From the perspective of membrane fusion, viral fusion peptides are
perhaps the most critical region of virus envelope glycoproteins as they
directly interact and disrupt target host cell membranes. Thus, the
corresponding regions have been extensively studied in model viral
fusion proteins. While there are no strict definitions for what constitutes
a viral fusion peptide, a number of criteria have to be met in order to
designate a segment of viral fusion protein a bona fide fusion peptide.
Often, the region corresponding to an FP is composed of hydrophobic
amino acids, particularly enriched in glycine (G) and alanine (A) re-
sidues. It is important to note that fusion peptides can contain a few
charged residues as well as bulky hydrophobic residues such as tryp-
tophans (W). Another feature of fusion peptides is that they correspond
to regions which are extremely sensitive to point mutations in the
context of full-length fusion protein, i.e. that a single residue substitu-
tion within an FP often results in loss of fusion activity. Related to this
characteristic is that although viral fusion protein sequences tend to
vary greatly amongst different viral families, they are extremely well
conserved within a given family. The capacity of a synthetic peptide to
5
Virology 517 (2018) 3–8
induce lipid-mixing and fusion of liposomes is another criterion that
can help in identifying regions hypothesized to be FPs. Furthermore,
the study of the effects of such peptides on lipid bilayer ordering using
powerful spectroscopy techniques such as electron spin resonance
(ESR) has also been successfully used to characterize other cellular and
viral FPs such as those of influenza virus and HIV (Ge and Freed, 2009,
2011; Lai and Freed, 2014, 2015; Pinello et al., 2017).
Within each major classes of viral fusion proteins (I, II, and III) the
associated fusion peptides share basic characteristics. Class I fusion
peptides are usually enriched in alanine and/or glycine residues, and
can either be “N-terminal” (i.e. located immediately downstream of the
activating cleavage site) or “internal” depending on their positions re-
spective to the cleavage site (Apellaniz et al., 2014). For class II and III
fusion proteins, the fusion peptides are not released by proteolytic
cleavage but instead, form so-called internal “fusion loops” which can
be bipartite as in the case of class III fusion proteins (e.g. vesicular
stomatitis virus glycoprotein G) (Apellaniz et al., 2014; Sun et al., 2008;
White et al., 2008). It is important to highlight that it is the anchoring
of the fusion peptides to apposing target membranes that allows for the
formation of a unique biological structure: two lipid bilayers connected
by fusion proteins.
4. Locating the coronavirus S fusion peptide
Identifying and locating the coronavirus fusion peptide has been
most extensively performed using SARS-CoV spike (S) as a model.
Initial work, using a peptide library derived from the SARS-CoV S
protein, showed that three regions (R1, R2, and R3) within the S2 do-
main displayed membrane-interacting properties in experiments mea-
suring vesicle membrane leakage (Guillen et al., 2005). R1 (aa
858–886) was found located upstream of HR1, R2 was identified be-
tween HR1 and HR2 (aa 1077–1092) and R3 (aa 1190–1202) was found
to be situated proximal to the transmembrane region. Mutations within
the region spanning residues 852–883, which approximately corre-
sponds to the above-mentioned R1 region, were found to decrease cell-
cell fusion in S-expression mediated syncytia formation assays (Petit
et al., 2005). Another group identified two regions using the Wimley
and White interfacial hydrophobicity analysis approach. They defined
these regions as WW-I (aa 770–778) and WW-II (aa 864–886) with the
latter almost matching the R1 region identified by Guillen and collea-
gues. WW-I was found to strongly partition into large unilaminar ve-
sicles (LUV) and be most important for fusion. Subsequently, a model
for how different membranotropic segments identified within SARS-
CoV S2 would function has been proposed in which the putative fusion
peptide (for consistency, this FP is referred to here as FP770–788) located
N-terminal of the HR1 region (aa 770–788) is followed by an “internal”
FP or IFP873–888 (aa 873–888) and another segment located upstream of
the transmembrane domain (PTM1185–1202, aa 1185–1202) (Guillen
et al., 2008; Guillén et al., 2008). In this model, all three segments work
sequentially and in concert: the FP first inserts into target membranes,
the IFP would then facilitate the hemifusion process by interacting with
the apposing membrane and the interaction of the PTM with target lipid
bilayer after further conformational changes of spike would also facil-
itate fusion pore expansion. More recently, structural characterization
of these regions has been undertaken (Basso et al., 2016; Mahajan and
Bhattacharjya, 2015). Mahajan and Bhattacharjya have used nuclear
magnetic resonance (NMR) spectroscopy to gain insight into the
structures adopted by isolated peptides corresponding to the above-
mentioned regions FP770–788, IFP873–888, and PTM1185–1202 regions.
They have determined the atomic-resolution structure of these peptides
in presence of dodecylphosphocholine (DPC) detergent micelles by so-
lution NMR (Mahajan and Bhattacharjya, 2015). Basso and colleagues
have used electron spin resonance (ESR) as well as differential scanning
calorimetry (DSC) to probe the interactions of the FP and IFP peptides
with lipid bilayers (Basso et al., 2016).
These initial studies have identified regions of S2 with the capacity
J.K. Millet, G.R. Whittaker
Fig. 2. Predicted model of the CoV fusion peptide region, along with its interaction
with the lipid bilayer. Conserved negatively charged and hydrophobic residues and a
proposed location of Ca2+ ions are shown.
to interact with membranes. They also suggest that several regions
within S2 could act in a concerted fashion to mediate the membrane
fusion process. Further studies have been conducted taking a more
functional approach and considering other criteria that define fusion
peptides, such as proximity to a cleavage site and sequence conserva-
tion (Madu et al., 2009a, 2009b). The starting point for these studies
was the identification of a second cleavage site in the SARS-CoV S S2
domain called S2′ (at R797 residue) (Belouzard et al., 2009). Authors
found that the SARS-CoV S protein was proteolytically activated se-
quentially at the S1/S2 and S2′ sites, and that the latter event was
critical for activating membrane fusion. Using these findings as a basis
for identifying the SARS-CoV fusion peptide, additional work has shown
that the segment located immediately downstream of the SARS-CoV S2′
cleavage site, 798SFIEDLLFNKVTLADAGF815, displayed features of a
fusion peptide. In addition to being proximal to the S2′ cleavage site,
the sequence of this region was found to be remarkably conserved
within the Coronaviridae, with the 800IEDLLF805 motif showing minimal
divergence. Interestingly, the highly conserved 803LLF805 correspond to
the beginning of a major antigenic determinant of SARS-CoV S protein
(aa 803–828), capable of eliciting the generation of neutralizing anti-
bodies (Zhang et al., 2004). Furthermore, mutagenesis analysis of the
S2′-proximal 798–815 segment in the context of full-length protein
demonstrated its importance in mediating membrane fusion. Sub-
sequent structural analysis of the isolated peptide by circular dichroism
(CD) spectral analysis and vesicle lipid mixing assays confirmed the role
of the segment as a fusion peptide (Madu et al., 2009b). More recently
this domain has been subjected to extensive characterization by elec-
tron spin resonance (ESR) spectroscopy (along with CD spectroscopy) to
show membrane ordering and the critical involvement of the LLF motif
6
Virology 517 (2018) 3–8
(Lai et al., 2017). Another mutagenesis study based on cell-cell fusion
and pseudovirion infectivity assays has also found that a pair of highly
conserved cysteines (C822 and C833) and the flanking residues D830
and L831 located C-terminally to the 798–815 region played a critical
role for fusion (Madu et al., 2009a). The small sub-domain formed by
the cysteine pair form a loop structure, a situation that is reminiscent to
the avian sarcoma/leukosis virus subtype A in which a pair of cysteines
flank the internal fusion peptide of the Env glycoprotein (Delos et al.,
2008).
ESR is a powerful tool enabling to analyze membrane-ordering ef-
fects of fusion peptides with lipid bilayers. Using this technique the two
distinct domains downstream of S2′ have recently been characterized.
The 22 amino acids region immediately after the S2′ cleavage site
(798SFIEDLLFNKVTLADAGFMKQY818) were termed FP1 and induced
significant membrane ordering. The work also showed that the sub-
sequent 21 amino acid, disulfide-bonded, domain
(816KQYGECLGDINARDLICAQKF835), termed FP2, also displayed
membrane-ordering properties. Furthermore, these effects of the two
domains were dependent on Ca2+ ions, a situation similar to the cal-
cium-dependency of fusion observed for the Rubella virus E1 envelope
glycoprotein (Dube et al., 2016, 2014). A specific role for calcium in
binding to both the FP1 and FP2 domains of SARS-CoV S was reinforced
by isothermal titration calorimetry (ITC) (Lai et al., 2017). Overall
these data suggest that for coronaviruses the fusion peptide is complex,
possibly forming an extended FP1-FP2 “platform” with a conserved LLF
core, as well as other regions of S2 previously identified as membrane-
interacting (Fig. 2), and suggesting that Ca2+ ions bind to conserved
negatively charged residues in the fusion platform, to comprise the
missing factor required to trigger membrane fusion.
5. The coronavirus fusion peptide in the context of the cell biology
of virus entry
The notion that SARS-CoV fusion peptide structure and function in
vitro is controlled by Ca2+ ions suggests that a similar situation occurs
in vivo. Indeed, SARS-CoV entry is heavily dependent on the Ca2+
concentration of the media in which cells are grown, and is inhibited by
calcium chelators such as BAPTA-AM, which act in endosomes (Lai
et al., 2017). It is also interesting to note that amiodarone, a drug that
blocks endosomal/lysosomal calcium channels, also inhibits SARS-CoV
entry, after endosomal uptake (Stadler et al., 2008). The inhibition by
amiodarone was originally attributed to Ca2+-dependent defects in
endosome trafficking, but may also act more directly in the membrane
fusion process.
Fig. 3. Cartoon model of SARS-CoV entry into
cells. In this model, fusion can take place in two
Ca2+-containing compartments corresponding to the
observed “early” and “late” entry pathways, with
cleavage occurring by distinct activating proteases
following receptor engagement.
J.K. Millet, G.R. Whittaker
In general, the role of Ca2+ and other ions in the context of en-
dosomal trafficking is poorly understood. While extracellular calcium is
high (approximately 1 mM), levels drop rapidly in the lumen of newly
formed endocytic vesicles, due to the action of efflux pumps (Huotari
and Helenius, 2011; Luzio et al., 2007; Scott and Gruenberg, 2010).
However, lysosomes can act as a calcium store via their interaction with
major calcium stores in the endoplasmic reticulum, with levels as high
as 0.5 mM found in lysosomes. During the later stages of endosome
maturation, lysosome-late endosome fusion mediated by the HOPs
complex can also result in substantial levels of Ca2+ in endosomal
compartments. Such trafficking pathways may therefore lead to the
availability of Ca2+ in two distinct points in the coronavirus entry
pathway. First, at the cell surface where TTPS-mediated cleavage can
activate fusion, and second, in “mature” endocytic vesicles where ca-
thepsins can activate fusion.
Overall, we propose a model of SARS-CoV and for coronavirus entry
in general, which integrates the various triggers and activators of the
viral spike protein as follows: 1) Protease cleavage at S1/S2, which
provides a preliminary priming step; 2) Receptor engagement; 3)
Protease cleavage at S2′ to expose the viral fusion peptide (FP1-2); 4)
Ionic changes to promote conformational changes and fusion peptide
insertion. These ionic changes are not exclusively driven by H+ ions,
and we argue that Ca2+ ions are necessary to promote fusion peptide
insertion into the lipid bilayer. Our model encompasses both the
available in vitro and in vivo data, with entry occurring from the ex-
tracellular space (TTSP-driven) – the “early’ pathway, or following
substantial endosome maturation (cathepsin-driven) – the “late
pathway”. Variations on this pathway may occur for some cor-
onaviruses in the situations where furin-like proteases activate viruses
in an intermediate endosomal compartment such as the early endosome
(Burkard et al., 2014; Millet and Whittaker, 2014). A cartoon diagram
of a basic entry model for SARS-CoV is presented in Fig. 3.
Acknowledgments
We thank Ruth Collins, Susan Daniel and members of the Whittaker
and Daniel labs for helpful discussions. This work was supported by the
National Institutes of Health Grants R21 AI1076258 and R21 AI111085.
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Another random document with
no related content on Scribd:
 Motive power.
Volume Max.
Type. h.p. per
m. No. Type. Speed.
Motor.
m/s
Zeppelin 17,700 3 Maybach 150 21
Zeppelin 18,700 3 — 150 21.1
Zeppelin 18,700 3 — 150 22
Zeppelin 18,700 3 — 150 22
Zeppelin 22,000 3 — 150 —
Zeppelin 20,000 3 — 170 —
Zeppelin 18,700 3 — 170 —
Zeppelin — — — — —
Parsifal 4,000 1 Daimler 85 14
Parsifal 7,500 2 N.A.G. 110 15
Parsifal 8,000 2 Maybach 180 18.8
Parsifal 10,000 2 Koerting 200 18.5
Parsifal 8,000 2 N.A.G. 110 16
Parsifal 10,000 4 Maybach 180 —
Siemens-Schückert 15,000 4 Daimler 125 19.8
Schutte-Lanz 19,500 2 Daimler 270 20
(1) Gross-Basenach 5,200 2 Koerting 75 12.5
(2) — 5,200 — — 75 12.5

(1) and (2) as in 1911; since then they have been renovated, and
no doubt their speed and volume are much greater.
We must accept with some reserve the reports that are current in
this respect, and it may be pointed out that in accounts of the doings of
Zeppelin airships in the papers it can be reasonably doubted whether
all the Zeppelins mentioned are in reality Zeppelins. Probably some
are the smaller types, such as the Gross or Parsifal. The word
Zeppelin seems to have become synonymous with a German airship,
and the wounded soldiers or prisoners who are responsible for many
of the stories told would not be likely to have complete knowledge of
the distinctions between classes of airships.
Though what Germany is exactly doing in way of new manufacture
must remain in much fog, still we can form some opinion as to her
preparedness with aircraft on the lighter-than-air principle from our
knowledge of what she possessed last year.
The table on the opposite page will show that her fleet of airships,
including those under construction, was then by no means negligible.
A nation possessing such a fleet of large airships as Germany
does must be provided with sheds (hangars) for their reception in all
parts of the country, and by the table that is appended it will be seen
that in this way last year Germany was very amply provided.
I am indebted to the Aérophile for the following list of German
hangars for dirigibles, with dates of construction and names of
owners:—
 Place and Date of
Proprietors. Observations.
Construction.
Aix-la-Chapelle — Designed for
1914
Allenstein — Designed for
1914
Baden—Baden-Dos — —
(1910)
Berlin—Biesdorf Siemens and Schückert —
(1909)
Berlin—Reinickendorf — —
Berlin—Johannisthal Aëronautical Sport Society —
(1910)
Berlin—Johannisthal — —
(1911)
Berlin—Tegel (1905) Prussian Army —
Berlin—Tegel (1907) Prussian Army —
Berlin—Tegel (1908– Prussian Army —
10)
Bitterfeld (1908) Luffahrtzeug Society —
Bitterfeld (1909) Luffahrtzeug Society —
Braunschweig Airship Harbour Society of Designed for
Brunswick 1914
Cologne — —
Cologne—Leichlingen Rheinwerke Motorluftschiff —
Society
Cologne—Nippes Clouth —
Cuxhaven German Navy Designed for
1914
Dresden City of Dresden Will only hold
one
balloon
Düsseldorf (1910) City of Düsseldorf —
Cologne—Bickendorf Prussian Army —
(1909)
Frankfurt am Main Delay —
(1911)
Friedrichshafen Zeppelin Society —
(1908)
Friedrichshafen— Workshops of the Zeppelin —
Manzell (1900) Society
Gotha (1910) Town of Gotha —
Graudenz — Designed for
1914
Hannover — Designed for
1914
Hamburg— Hamburg Airship Harbour —
Fuhlsbüttel (1911) Society
Hamburg—Hansa — —
Kiel (1910) Union for Motor-Airship Travel —
Königsberg-in- Prussian Army —
Preussen (1911)
Leehr — Designed for
1914
Leipzig Leipziger Luftschiffland —
Flugplatz Gesellschaft
Liegnitz (1913) Prussian Army In construction
Mannheim— — —
Schwetzinger
Mannheim—Rheinau Luftschiffbau Schütte u. Lanz —
(1909)
Metz (1909) Prussian Army —
Potsdam, near Berlin Zeppelin Society — —
(1911)
Posen — Constructing
Schneidemühl — Building
 Strasbourg Prussian Army —
Thorn (1912) — —
Trèves — Building
Waune (1912) Rhenish-Westphalien Flying —
and Sports Club

Such monster airships as the Zeppelin call for a large proportion of

pure hydrogen. This is, indeed, manufactured on a large scale in
Germany. It is produced in quantities by the electro-chemical works at
Bitterfeld, Griesheim, and at Friedrichshafen, specially for the needs of
the Zeppelins at the latter place. There are also works for the
production of very pure hydrogen by electrolysis at Bitterfeld,
Griesheim, Gersthofen, and Dresden.
In the particular way Germany means to use her lighter-than-air
fleet in the present war time will show. If, however, there have not yet
been attempts at any combination of action, individual Zeppelins have
already played the rôle of dreadnoughts of the air. Though their powers
have been no doubt exaggerated, they have been the terror of some
Belgian cities.
Early in the morning of August 25th a Zeppelin airship visited
Antwerp, and drifting silently with the wind steered over the temporary
Royal palace. There it discharged six highly explosive bombs. Not one
found its intended mark, though all fell near the palace. One appears
to have been very near hitting the tower of the cathedral. Though the
bombs failed to attain the object sought, no less than six or seven
persons were victims to the outrage. One struck a private house, killed
a woman, and injured two girls, killed two civic guards, and wounded
another. One bomb fell in the courtyard of the hospital of St. Elizabeth,
tore a hole in the ground, smashed the windows, and riddled the walls.
The Zeppelin repeated its visit early in the morning of September
2nd, but this time with less deadly result. The bombs only wounded the
victims. The experiences of the first visit had given effective warning
against a repetition of aërial invasion. The city had been darkened, and
the airship was attacked from the forts and the high points of the city
as soon as it made its appearance. The crew of the airship seem to
have been struck with panic when it failed to find its bearings over the
darkened city.
It appears they suddenly dropped all their bombs as ballast and
rose quickly out of harm’s way. The bombs used on this occasion were
not of the same type as those used on the previous attempt on the city.
The latter were of high explosive power designed to destroy buildings.
The former were covered by thin envelopes, and held together by
mushroom-shaped rivets. They were filled with iron bolts and nuts, and
were evidently designed for the destruction of human life. It is stated
that this is a type of bomb which has never been used by artillery,
being made on the same model as that used by the notorious French
robber, Bonnet.
In reference to airship raids over cities, it has been suggested in
America that the air in their immediate neighbourhood should be
mined. This could be done by having a number of captive balloons or
kites, the mines on which could be discharged electrically from the
ground. For future wars there will no doubt be devised some form of
travelling aërial torpedoes for destroying the intruding airships. Such
torpedoes would, however, have to be capable of guidance. As has
been pointed out by Mr. W. F. Reid, in 1884, at the siege of Venice, the
Austrians used free balloons for the purpose of dropping bombs upon
the town. The bombs were attached to the balloons in such a way that
after the burning of a certain length of safety fuse, the connection was
severed, and the bomb fell. The length of fuse was calculated
according to the speed of the wind; but, unfortunately, when the
balloons rose, they entered an upper air-current travelling in a different
direction from that below, and many of the bombs burst in the Austrian
lines, whence they had started. Thus it would not be expedient to let
loose ordinary unmanned balloons loaded with timed explosives, even
if the direction of the wind seemed favourable, for their meeting an
approaching airship fleet, as an upper current might bring them back
over the city, where they might do mischief.
It is, however, quite conceivable that in the future aërial torpedoes
may be devised in the shape of unmanned balloons or aëroplanes
controlled by wireless waves of electricity. Those who saw the striking
experiment of steering a small navigable balloon in a large hall entirely
by wireless electric waves must have realised the possibilities which
may thus be opened out in the future.

* * * * *
While writing, the news has come that another Zeppelin has
dropped three bombs on Ostend, the casualty list being one dog. Two
unexploded projectiles were found on a field near Waeragheim. These
were probably thrown from the same airship. They show how
constantly missile throwing from a moving airship may fail to come
near the mark. There is no doubt that to hit particular objects aimed at
from airships is by no means an easy matter. Success would seem to
require considerable training in this particular method of warfare. The
late Colonel Moedebeck, in his well-known pocket-book of aëronautics,
makes the following remarks on the throwing of balloon missiles:—

We may assume that, if handled skilfully, the object aimed

at will be hit very exactly. We must distinguish between the
throw when the airship is at rest and that when it is in motion. In
throwing out while at rest, which is only possible when the
airship can travel against the wind, the following points must be
considered:—

(a) The height of the object.—This may be accurately determined

from the contour lines on the map, or from a determination of its
normal barometric height. Both must be done before starting.
(b) The height of the airship above the object.—The barometric
height is read and reduced to normal conditions. The difference in
heights as found from (b) and (a) gives the height above the object.
(c) The velocity of the wind.—May be read on an anemometer in
the airship, or determined beforehand by captive balloons.
(d) The time of fall.—Given by the law of gravitation from the
determination under (b).
The height of fall = h = gt²/2.
Whence the time of fall t = √(2h/g).
(e) The resistance of the air. R = (γ/g)Fv².
(f) The leeway.—The longer the fall, and the lighter and larger
the falling body, the stronger is the drift. For known missiles, the drift
 for different heights and wind velocities may be determined
practically.
(g) Unsteadiness of the airship.—The irregularity of the pressure
of the wind, and its constant variation in direction, renders it
impossible for the airship to remain perfectly steady.

The elements stated under (b) and (f) must be rapidly

determined, and suitable tables have been prepared for this
purpose. The irregularity of the wind and the peculiarities of the
airship mentioned under (g) render a preliminary trial
necessary. The drift also is determined by this method, before
the large air-torpedo is cast out.
The air-torpedo must be brought by sight vertically over the
object by steering the airship, the value of the mean drift
previously determined being allowed for.
In throwing out a missile while actually travelling, the
velocity of the airship must be taken into account, as well as the
elements (a) to (g) given above, since this velocity is also
possessed by the body thrown out.
The determination of the proper point is now greatly
increased in difficulty. Its position is a function of the relative
height of the airship above the object, of the velocity, and of the
drift, and allowance must be made for all these factors. For this
purpose, motion, either with or against the wind, is the simplest.
On account of the point on the earth over which the missile
must be thrown out not being in general well marked, it is
necessary to use also angles of sight.
The problem before the aëronaut is, then, as follows:—For
a given height, velocity, and drift to find the necessary angle of
depression at which the missile must be thrown out in order that
it may fall on to the object.
The casting out of the missile against the object while
travelling is governed, therefore, by the same rules as those
governing the discharge of a torpedo from a torpedo-boat.
 CHAPTER V
ADVANTAGES AND DISADVANTAGES OF
AIRSHIPS

The chief advantages of aircraft that are lighter than air over those
that are heavier than air in warfare are:—

1. Their speed can be variable.

2. They can hover over a particular point.
3. They can be noiseless by cutting off motive power and
drifting for a while with the wind.
4. They can from their possible size have long range of
action.
5. They can carry considerable weights.
6. They are endowed with sustaining power and stability.

1. Their speed can be variable.

This advantage becomes apparent in cases where they are used
both for scouting and offensive purposes.
In a later chapter it will be pointed out that though the aëroplane
scout has often to make dashes over the enemy, and it would be
thought that from his swift movements his impressions might be
vague, still, in practice, most satisfactory work has been undoubtedly
accomplished. Many, however, will maintain that there are
circumstances when it may be advisable for observers to proceed at
variable speeds. When at a safe height it may be an advantage for
the observers to take their time and leisurely survey the country,
observe, and take photographs. The airship can stealthily travel over
camp and fortress and steal secret after secret of the enemy.

2. They can hover over a particular point.

The fact that the maintenance of the airship in the air does not
depend upon a certain speed being maintained, as is the case with
the heavier-than-air machine, endows it with the property of being
able to hover in fairly calm weather. The hovering power is certainly
an advantage for such offensive operations as dropping bombs.

3. They can be noiseless.

At night it may often be possible to approach over a fortress,
camp or city quite noiselessly at a low altitude by shutting off the
motive power and navigating by means of the natural forces alone.

4. They can from their possible size have long range of action.
From their size and the amount of fuel they can carry it is
possible for them to travel for long distances.
This quality renders them specially fitted for naval purposes,
though possibly in the not very distant future more highly developed
hydroplanes will run them very close.

5. They can carry considerable weights.

The weights large airships can carry is an advantage in offensive
operations. It enables larger stores of bombs to be carried than is at
present possible with aëroplanes. Then several persons can be
carried long distances in the larger airships.

6. They are endowed with sustaining power and stability.

As the envelopes of airships are filled with a gas which lifts and
sustains, the great disadvantage of instability which is the bugbear of
aëroplanists is absent. If engines break down or stop, it does not
necessarily mean that the airship must immediately descend. It can
often remain in the air while the machinery is being repaired.
 But in spite of these advantages airships have very numerous
counterbalancing disadvantages, so marked, indeed, that it seems a
question whether, if the world decided to entirely use aëroplanes in
their place, it would be much the loser.
The principal disadvantages would seem to be:

1. The resistance of the gas-bag.

2. Danger of fire from close combination of petroleum
motor and gas-containing envelope.
3. Danger of fire from self-electrification of surface.
4. Difficulties in the way of applying the propulsive screws
in the most effective position.
5. Difficulties of making gas envelope gas-proof.
6. Great cost of airships.
7. The great amount of personnel needed for the
manipulation of large airships.
8. Great liability of being destroyed by aëroplanes in war.
9. Insufficient power of quickly rising.

1. The resistance of the gas-bag.

From a mechanical point of view it is in opposition to science to
attempt aërial navigation by pushing such a large resisting surface
as the envelope of an airship against the air. In navigating an airship
against the wind, as the latter increases speed is diminished, until a
limit is reached when the motive power will be unavailing. Thus there
are weather limitations to the airship. Not that the aëroplane is
unaffected by the weather. That also has its limits; but recent
practice has shown that the proportion of days when aëroplanes can
fly is considerably larger than those on which airships can venture
forth from their sheds.
This disadvantage of the resistance of surface was very manifest
in the earlier experiments with navigable balloons, when only feeble
motive power was available. For instance, in Count Zeppelin’s
experiments in 1900, his two motors of 16 h.p. could not combat a
greater wind force than about three metres a second. Then airships
could indeed only be called toys. It has only been possible to make
them partially successful concerns by enormously increasing motive
power. At the h.p. figures with which the latest made large airships
have been endowed, the wind limit is much lower than in the case of
the heavier-than-air constructions. Though now airships can
encounter moderate winds, they are still fair-weather instruments.
For the great records of distance established by Count Zeppelin
favourable meteorological conditions have been wisely selected. It
was M. Santos Dumont who first led the way in making airships
something beyond toys. He, in his picturesque and world-alluring
experiments, first dared to encounter winds which in force exceeded
what would be called calm weather. It is exceedingly difficult to
ascertain what are the exact wind forces overcome by a body
moving in air. The measurements have to be taken from a point
independent of the moving body. We generally find this one
important figure omitted in accounts of airship voyages. M. Santos
Dumont’s experiments gave especially favourable opportunity for
ascertaining correct records of the wind forces overcome. Since M.
Santos Dumont so frequently rounded the Eiffel Tower close to the
storey where the meteorological instruments were placed, the writer
obtained from the authorities of the Eiffel Tower a record of the wind
forces registered on all the days of his experiments. A comparison of
those records with those of M. Santos Dumont’s journeys made it
possible to approximately ascertain the highest wind forces he
combated on his journeys round the tower; these were about five
metres a second. M. Santos Dumont, however, appears to have
claimed six metres a second for his highest wind record.
The brothers Lebaudy in their earlier experiments about doubled
the record of Santos Dumont in this respect. As time has gone on
greater advance has been made, though the limit is still represented
by moderate wind.
There is, perhaps, some consolation in this thought for those
who fear raids of an inimical airship fleet. The proverbial windy
nature of our favoured islands is perhaps even more protection than
darkened cities and artillery shot, though it is well indeed not to
neglect the two latter precautions.
Meteorologically speaking, to make a raid with bulky airships
from a distance over these islands would be a very risky
undertaking, fraught with the greatest danger to the occupants of the
airships. It must be remembered that, chiefly owing to the weather,
the history of the Zeppelin may well be called the history of disaster.
For the very reason of its fragility over and over again it has been the
victim of tempest and flame.
The use of aluminium for the framework of the Zeppelins has
been largely responsible for Count Zeppelin’s repeated weather
misfortunes. There has been a fascination about this brittle metal
aluminium for aëronautical work on account of its lightness. Its
employment for aircraft construction, except for trivial purposes, is,
however, a fallacy. That most practical aëronautical engineer, M.
Julliot, in working out his semi-rigid constructions, has never fallen
into the snare of aluminium allurement, wisely using steel instead.
Considering the aluminium framework of the first Zeppelin
constructed was fairly wrecked by the trifling accident of its falling
down from the ceiling of the shed to the floor, it is a wonder that this
species of metal has been retained, to be crumpled up almost like
paper in the many accidents that have occurred.

2. Danger of fire from close combination of petroleum motor and

gas-containing envelope.
In airships of all three types—rigid, semi-rigid, and non-rigid—
this danger is constantly present. There have been examples of
airship conflagrations in mid-air, but the greatest danger of
conflagrations is in descending when the airships have been
overtaken by strong and gusty winds. As has before been stated, fire
has been the great destroyer of the Zeppelins.
The nearer the car containing the motors is placed to the gas
envelope, the greater the fire risk becomes. The Parsifal airship, in
which the car is suspended a considerable distance from the gas-
bag, should in this respect be the safest of all the types of airships
yet constructed.

3. Danger of fire from self-electrification of surface.

This appears to be a great danger in the case of airships whose
gas-bags are made with india-rubber surfaces. No less than two
Zeppelins have been destroyed from this cause. In the case of the
explosion of the gas in a Zeppelin of 1908, when it burst from its
anchorage at Echterdingen, the destruction of the airship appears to
have been caused by electric sparks produced by the friction of the
material of which the gas-bag compartments were made. Colonel
Moedebeck, in the Aëronautical Journal of October, 1908, gave an
expert opinion as to the cause of this accident:—

The balloon material, which is india-rubber coated, has

the peculiar property of becoming electrified in dry air. When
rolled up or creased in any way it rustles, and gives out
electric sparks, the latter being (as shown by the experiments
undertaken by Professor Bonsteim and Captain Dele for the
Berlin Aëronautical Society) clearly visible in the dark.
Now, the lower parts of the material of which the gas-cells
are composed would, owing to the height to which the airship
had ascended (1,100 m.) and the release of gas from the
valves, become creased or folded upon each other, and the
rubbing thus produced would be quite sufficient to generate
the electric sparks above referred to. Under ordinary
circumstances, when the space between the gas-cells and
the outer envelope of the airship is full of atmospheric air,
continually renewed, as when it is in full flight, these sparks
would be harmless enough, but when the ship is at anchor, as
at Echterdingen, this is not necessarily the case.
We know that the carefully made tissue of the Continental
Caoutchouc Company resists the penetration of hydrogen
very strongly, but some may have leaked through into the
space between the cells and the outer envelope, while it
seems very probable that when the mechanics opened the
 valves, and the long axis of the balloon became inclined,
more hydrogen entered this space and an explosive mixture
was formed.
According to the description given by eye-witnesses, the
explosion took place after the forepart of the vessel (dragging
its anchor) struck the ground. The shock thus caused would
have been transmitted to the creased and wrinkled gas-cells,
and the tearing of the material, already in an electrified
condition, might easily have generated sufficient sparks to
detonate the explosive mixture.

Again, in 1912, there was a repetition of this kind of disaster in

the case of the destruction of another Zeppelin, the “Schwaben.” In
this case the framework of the airship had got broken, being battered
about in landing in an adverse wind. The india-rubber-coated bags
were rubbed against each other, with the production of electric
sparks. These either set fire to the gas issuing from one of the gas-
bags or exploded the mixture of air and gas contained in the space
between the gas-bags and outer covering of the airship. Perhaps it
was on account of this accident that gold-beaters’ skin has
sometimes been used for the gas containers of the Zeppelin
airships.

4. Difficulties in the way of applying the propulsive screws in the

most effective position.
Most airships are exceedingly defective in this respect, the
screws being applied to the propulsion of the car and not to the
whole system. The result is that the cumbersome gas-bag lags
behind. Certainly, one of the best points in a Zeppelin was the
attachment of the screws to the airship framework above the cars,
thus securing more advantageous position. This, however, only
amounted to something like half measures. In the case of the ill-fated
airship “La Paix,” the Brazilian aëronaut Severo undoubtedly aimed
at the ideal, though the experiment cost him his life. He devised the
ingenious system of combining balloon and car in one symmetrical
melon-shaped body, through the centre of which passed
longitudinally the shaft which revolved the propelling screws at either
end. The screws were therefore in the position in which to propel the
whole system and not the car only. This, however, necessitated the
introduction of a very small space between the car and balloon
proper. By reason of this very small space the presence of the
petroleum motor in the car could not fail to be dangerous, and was
the cause of the fiery end of Severo’s balloon and the death of the
inventor and engineer. On the morning of the ill-fated May 11th,
1902, Severo and Sachet ascended in “La Paix.” A few moments
after the ascent the balloon exploded, in the words of an eye-
witness, like a crash of thunder, and the occupants were precipitated
to the ground.
In spite of the engineering advantages of Severo’s system no
one has dared to revive the plan.
It has, however, been pointed out by the writer—and the
suggestion elicited the keen interest of the late Professor Langley—
that if electricity could be used as the motive power in an airship the
Severo system could be reasonably revived. Then the electric
motors could be inside the gas-bag. There, electric sparks and
electric heating could do no harm. For it is only the borderland that is
the place of danger, where there are oxygen atoms to combine with
the hydrogen atoms. In the case of a balloon filled with gas it is
surprising to what short distance the danger zone extends. In the
case of the writer’s electric signalling balloons, on one occasion the
ladder framework which supported the incandescent lamps was
being hauled up into the balloon. Through some fault in the
connections there was sparking at the framework just as it had
passed over the dangerous borderland. The sparks went on with
safety. An inch or two lower and there would have been an
explosion!
But on account of the weight of the battery the practical
application of electricity for propelling navigable balloons seems to
be as far off as it was in the days of “La France,” and in airships we
have to continue placing the screws in the wrong place.

5. Difficulties of making gas envelope gas-proof.

 The absence of the knowledge how to obtain a really gas-proof
envelope is, no doubt, one of the greatest difficulties of airship
construction. As has already been pointed out, the gas-holding
quality of gold-beaters’ skin is remarkable. Its cost, however, is fairly
prohibitive in the case of large airships. A material which is a
combination of india-rubber and cotton surfaces is now generally
used for large airships, but this has undoubted disadvantages. India-
rubber is a substance which time, low temperature, and certain
climatic conditions deteriorate. All those who have worked with india-
rubber experimental ballon-sondes (sounding balloons) can testify to
its perishing qualities. Very much can be accomplished with a brand-
new airship. Turned out of a factory it will retain its gas-holding
qualities for a short time excellently. The lapse of time reveals
deterioration and leakiness.
Considering the extreme importance of a varnish that will retain
pure hydrogen for a reasonable time, it is a matter of surprise that
chemists should have almost entirely neglected its production. Mr.
W. F. Reid alone of British chemists seems to have given any serious
thought to the question. In a paper which Mr. Reid read before the
Aëronautical Society of Great Britain, he made some exceedingly
important suggestions in the way of obtaining balloon and airship
varnishes. In case this little volume should fall into the hands of any
chemists who may like to devote their powers of original research to
the production of one missing link in airship construction, the
following quotation from Mr. Reid’s remarks are appended below.

Varnishes may be divided into two classes—those in

which the film solidifies or “dries” by absorption of oxygen
from the air, and those in which the varnish “sets” by the
evaporation of a volatile solvent in which the solid ingredients
have been dissolved. To the first class belong the drying oils,
chiefly linseed oil, for, although there are a number of “drying”
oils, but two or three of them are used commercially in the
manufacture of varnishes. When exposed to the air,
especially in warm weather, linseed oil absorbs oxygen and
forms an elastic translucent mass termed by Mulder “linoxyn.”
This linoxyn has completely lost its oily nature, does not soil
the fingers, and is, next to india-rubber, one of the most
elastic substances known. It possesses but little tensile
strength, however, and can be crumbled between the fingers.
It forms the basis of all linseed oil paint films, and is largely
used in the manufacture of linoleum. Linoxyn, however, is not,
as Mulder supposed, the final product of the oxidation of
linseed oil. When exposed to the air it is still further oxidised,
and then forms a sticky, viscid mass, of the consistency of
treacle and of an acid reaction. This latter property is of
importance because it is due to it that fabrics impregnated
with linseed oil so soon become rotten. In order to hasten the
oxidation of linseed oil it is usually heated with a small
quantity of a lead or manganese compound, and is then ready
for use. No method of preparation can prevent the super-
oxidation of linseed oil, but experience has indicated two
ways of diminishing the evil effects so far as paints and
varnishes are concerned. The first is to mix the oil with
substances of a basic character or with which the acid
product of oxidation can combine. In the case of paints, white
lead or zinc oxide are chiefly used for this purpose. The other
method consists in mixing with the oil a gum resin which
renders the film harder and prevents liquefaction. Such a
mixture of linseed oil and Kauri gum forms an elastic, tough
mass, which is much more durable than the linoxyn alone,
and also possesses greater tensile strength. During oxidation
the linseed oil absorbs about 12 per cent. of its weight of
oxygen, and when the area exposed is very large in
proportion to the weight of the oil the temperature may rise
until the mass catches fire. At a high temperature the super-
oxidation of the oil takes place more rapidly than in the winter,
and I have seen fabrics that had only been impregnated with
an oil varnish for a month cemented together in one sticky
mass, and, of course, completely ruined. When the linseed oil
is thickened by the addition of a gum resin, it is too thick for
direct application, and is thinned down with a solvent, usually
turpentine or a mixture of this with light petroleum. Many
resins and gum resins are used in the manufacture of
varnishes in conjunction with linseed oil, but none of them can
deprive the oil of the defect referred to, and if used in too
large a proportion they become too brittle for balloon
purposes. Both scientific investigation and practical
experience show that any varnish containing linseed oil must
be looked upon with suspicion by the aëronaut, in spite of the
glowing testimonials some manufacturers are always ready to
give their own goods.
When we consider those varnishes which are solutions
and which do not depend upon oxidation for their drying
properties we enter upon a very wide field.
Practically any substance that is soluble in a neutral
solvent and leaves an impermeable film on drying is included
in this class. One of the simplest examples is gelatine in its
various forms, with water as a solvent. Until recently glue or
gelatine would have been useless for our purpose on account
of its ready solubility in water, but now that we are able to
render it insoluble by means of chromic acid or formaldehyde
it comes within the limits of practical applicability. A fabric may
be rendered almost impermeable to gas when coated on the
inside with insoluble gelatine, and on the outside with a
waterproof varnish. Animal membranes are far less
permeable to gases than fabrics coated with varnishes of the
usual kinds. A balloon of gold-beaters’ skin, if carefully
constructed, will retain hydrogen gas for a long time, and if
treated with gelatine that is afterwards rendered insoluble it
becomes practically impermeable. Fabrics treated with
linseed oil varnish, on the other hand, allow gas to pass with
comparative ease. This is not a question of porosity or
“pinholes,” as is sometimes imagined, but a property inherent
to the material. Hydrogen or coal gas is absorbed on the one
side of the film and given off on the other in the same way as
carbonic oxide will pass through cast iron. An inert gas, such
as nitrogen, does not appear to diffuse in this way, even when
there is a considerable difference in pressure between the
two sides of the film. Such a varnished fabric transmits
hydrogen readily, but retains nitrogen, and is perfectly
watertight. In filling up the interstices of a fabric composed of
cellulose the most obvious substance to use would be
cellulose itself, but until recently solutions of this kind were
difficult to obtain. Toy balloons have long been made of
collodion, and are fairly satisfactory, but a cotton fabric
impregnated with pure collodion becomes hard and even
brittle. Celluloid solution, which is collodion with camphor and
a small quantity of castor oil, is more flexible, but, probably on
account of the camphor, is more permeable to hydrogen than
collodion. A variety of collodion known as flexile collodion is a
solution of collodion cotton with a slight addition of castor oil,
and is much to be preferred to any of the preceding forms. In
using it great care must be taken to exclude moisture, as the
presence of this renders the film opaque, in which case it is
always more or less porous. A substance allied to collodion is
velvril material, composed of collodion cotton and nitrated
castor oil. It is tough and flexible, even in thick films, and
gives a good coating to paper or cotton fabric. Unless very
carefully prepared, however, acid products may be generated
from the decomposition of the nitro-compounds present, in
which case the strength of the fabric would suffer. Another
form of cellulose in solution is viscous, which forms a good
coating when applied in a very thin layer, but makes the fabric
harsh and brittle if used in excess. The solutions of this
substance do not keep well and are liable to spontaneous
decomposition.
The difference in flexibility between thin and thick films of
the same materials is very considerable.
Given an elastic, supple cement, such as is afforded by
concentrated solutions of some of the above-mentioned
substances, it is quite possible to cement a tough, close-
grained paper to a cotton fabric of open mesh, and the
compound material thus produced is much more easily
rendered impermeable than the fine cotton fabric now used.