Abstract
The diaphragm is an unique skeletal muscle separating the thoracic and abdominal cavities with a primary function of enabling respiration. When abnormal, whether by congenital or acquired means, the consequences for patients can be severe. Abnormalities that affect the diaphragm are often first detected on chest radiographs as an alteration in position or shape. Cross-sectional imaging studies, primarily CT and occasionally MRI, can depict structural defects, intrinsic and adjacent pathology in greater detail. Fluoroscopy is the primary radiologic means of evaluating diaphragmatic motion, though MRI and ultrasound also are capable of this function. This review provides an update on diaphragm embryogenesis and discusses current imaging of various abnormalities, including the emerging role of three-dimensional printing in planning surgical repair of diaphragmatic derangements.
DEVELOPMENT OF THE DIAPHRAGM
Development of the diaphragm occurs during weeks 4–12 of embryonic life (Figure 1 and Supplementary video 1).1, 2 The septum transversum, a thin mesodermal plate that provides scaffolding and forms the floor of the pericardial cavity, separates the thorax and abdomen. The pleuroperitoneal folds FORM next and are positioned laterally in the embryo. Subsequently, the post-hepatic mesenchymal plate (PHMP) forms. This structure grows in a laterodorsal direction to close the pleuroperitoneal canals and seals the thorax from the abdomen. Ingrowth of progenitor cells from somites form the muscular portions of the diaphragm. Recent research found that abnormal development of the post-hepatic mesenchymal plate due to genetic derangements is central in the formation of congenital diaphragmatic hernias.3–5
Figure 1.
Final image of normal diaphragm development illustration (Please see supplemental video for full illustration).
CONGENITAL ABNORMALITIES OF THE DIAPHRAGM
Congenital hernias
Bochdalek hernia
Bochdalek hernias result from incomplete closure of the embryonic pleuroperitoneal membrane. Despite the name, they typically occur through posterolateral defects in the diaphragm that are separate from the foramen of Bochdalek.6 The defects also occur medially and are of variable size. These hernias are seen more commonly on the left, an observation that has been attributed to earlier closure of the right pleuroperitoneal membrane7 and protection by the liver.8
In the neonatal period, a large congenital diaphragmatic hernia is a surgical emergency.9 The initial chest radiograph usually reveals a space-occupying lesion of the hemithorax and contralateral shift of the mediastinum, caused by herniation of the abdominal contents into the chest. Morbidity and mortality are related to the degree of underlying pulmonary hypoplasia. Prenatal diagnosis is possible with the use of fetal ultrasonography or MRI.10
In adults, a Bochdalek hernia is usually asymptomatic and discovered incidentally by chest radiography or CT. Such incidental hernias may be more frequent on the right side.11 The concomitant diaphragmatic defect is easily seen on CT or MRI. Other radiologic modalities also may demonstrate the hernia contents: barium studies may reveal herniated bowel loops; intravenous urography may reveal a herniated kidney; and radionuclide imaging may demonstrate hepatic or splenic herniation. If bowel or organs are involved, there is a risk of bowel obstruction and strangulation (Figure 2).12 Small focal diaphragmatic defects, with or without herniated fat or viscera, may be seen in more than 5% of adults on CT.13, 14 Their increasing incidence with age and emphysema strongly suggests that most such abnormalities are acquired and are not true congenital Bochdalek hernias.14
Figure 2.
Coronal CT of a congenital Bochdalek hernia (a), initially contains non-obstructed large bowel with subsequent development of bowel obstruction (b).
Morgagni hernia
Foramen of Morgagni hernias are related to maldevelopment of the embryologic septum transversum with failure of fusion of the sternal and costal fibrotendinous elements of the diaphragm.7, 15 In contrast to the true Bochdalek hernia, a hernia sac of peritoneum and pleura surrounds the contents of a Morgagni hernia.16 Morgagni hernias are often associated with obesity and most often right-sided, probably because left-sided defects are covered by the heart and pericardium.
Morgagni hernias usually come to clinical attention as asymptomatic right cardiophrenic angle masses on chest radiographs. The finding of omental vessels coursing across the parasternal diaphragmatic defect facilitates CT diagnosis (Figure 3). The actual defect may be difficult to identify because of its typically small size. CT readily permits distinction from other causes of cardiophrenic angle masses by revealing abdominal viscera, omental fat and vessels peripheral to the diaphragm in the lower anterior chest. As with Bochdalek hernias, multiplanar MRI is occasionally useful.
Figure 3.
Axial and coronal CT images (a, b) of a large Morgagni hernia demonstrate mesenteric vessels and mesenteric fat extending from the parasternal portion of the diaphragm to the herniated bowel loops filling the right lower hemithorax. The white arrows indicate the hemidiaphragm and the red arrow denotes the vessels.
Accessory diaphragm
Accessory, or duplicated, diaphragm is a rare congenital anomaly in which a thin, fibromuscular membrane is attached to the diaphragm anteriorly and courses posterocranially to attach to the posterior rib cage.17–22 CT or MRI may be helpful in suggesting the diagnosis and identifying associated congenital anomalies, particularly pulmonary hypoplasia.21
ACQUIRED ABNORMALITIES AFFECTING THE DIAPHRAGM
Acquired hernias
Hiatal hernia
Hiatal hernia is the most frequently encountered diaphragmatic hernia in adults. Acquired enlargement of the esophageal hiatus and laxity of the phrenoesophageal ligament are etiologic factors, often associated with conditions resulting in increased intra-abdominal pressure, such as obesity and pregnancy.7, 16 Sliding hiatal hernias (Type I) are more common than the paraesophageal (Type II) variety, in which the stomach herniates up alongside the lower esophagus while the gastroesophageal junction remains underneath the diaphragm.
On chest radiography, hiatal hernias are depicted as lower posterior mediastinal, retrocardiac soft tissue masses, often containing an air-fluid level. The diagnosis can be confirmed by a barium esophagogram, although this is rarely necessary. Very large hernias can become incarcerated or undergo volvulus (Figure 4).23 Multidetector CT or multiplanar MRI can be useful to define the contents of large hiatal hernias when operative repair is planned.
Figure 4.

Upper gastrointestinal series demonstrates a large paraesophageal hernia, manifesting as an intrathoracic stomach. The gastric fundus (F) is rotated posteroinferiorly, consistent with organoaxial rotation.
In fact, cross-sectional imaging serves as the springboard from which three-dimensional (3D) models are created.24 3D models can assist in critical surgical planning for complex hernias, enabling individualized medicine since each patient’s abnormality is mapped out precisely prior to surgery (Figure 5). Moreover, 3D models are essential in designing and testing innovative reconstruction surgeries. Consulting with the surgical team, the important anatomy to be printed is selected on a patient’s MRI and/or CT. The 3D modeling team segments the requisite anatomy and creates a computerized 3D image of the model. The 3D modeling program typically allows fusion of different structures from an MRI, CT or both to achieve the best anatomic depiction. The optimal material to construct the 3D model is selected. The model is printed by a multimaterial 3D printer, which deposits layers of photopolymers that are subsequently hardened by ultraviolet light. After washing and drying, the model is ready for review by multidisciplinary teams for surgical planning and for educating trainees and patients.
Figure 5.
Presurgical planning for a patient with a large paraesophageal hernia utilizing 3D modeling starts with segmentation of the necessary structures (a), followed by construction of a 3D computer model (b). Final product after supporting material has been removed (c). 3D, three-dimensional.
Traumatic hernia
Traumatic diaphragmatic hernia usually results from either blunt or penetrating injury. Diaphragm rupture has been recognized in less than 6% of blunt trauma survivors.25–28 It affects the left hemidiaphragm more frequently, possibly because of protection of the right hemidiaphragm by the liver or inherently greater weakness of the left hemidiaphragm.29 Infrequently, bilateral diaphragmatic rupture occurs.25–28,30–32 Blunt traumatic tears can involve any portion of the diaphragm,33 although they usually involve the posterior central aspect of a hemidiaphragm and extend radially, or they can result in disruption of the posterolateral attachments.25, 34 Blunt traumatic defects are usually large, often more than 10 cm in length.15, 27,31,35 Penetrating trauma due to stab wounds most often affects the left hemidiaphragm because most people are right-handed, whereas gunshot wounds affect both sides equally.31 Penetrating wounds are usually less than 2 cm in length.35Though exceedingly rare, diaphragm rupture and hernia may occur spontaneously from sudden increases in abdominal pressure, such as with vomiting, parturition, coughing, and vigorous physical exertion,36 or after thoracoabdominal surgery.37
No reliable clinical signs or symptoms indicating diaphragmatic rupture have been identified.26 Abdominal injuries managed conservatively can be monitored by imaging, precluding identification of diaphragmatic tears that would have been detected during exploratory laparotomy. Early diagnosis, therefore requires a high degree of suspicion, which is important because the pleuroperitoneal pressure gradient can cause defects to enlarge over time,31, 38 with eventual bowel obstruction, incarceration, and strangulation.33,39–43
Detection sensitivities for diaphragmatic rupture on chest radiographs is typically less than 50% (range from 20 to 71%).25–28,31,32,39 The disparate sensitivities from prior studies is likely multifactorial, including simultaneous lung pathology that mask or mimic diaphragmatic rupture and delayed presentation when certain studies may have accounted for initial radiographs only. Herniation of hollow viscera into the chest and intrathoracic stomach with a nasogastric tube (Figure 6) are the most specific radiographic signs. Diaphragm rupture should be suspected whenever radiographs reveal apparent elevation of a hemidiaphragm, although this can be caused by atelectasis, eventration, diaphragm paralysis, or subpulmonic pleural effusion. Right-sided rupture is more difficult to detect radiographically; one study suggests that a 4–5 cm or greater elevation of the right hemidiaphragm relative to the left should be considered highly suspicious for right-sided rupture in the setting of blunt trauma.37
Figure 6.

Frontal chest radiograph of a male after a motor vehicle accident shows the nasogastric tube coiled in the herniated intrathoracic stomach.
Evidence of rupture may not be present on the initial radiograph but may develop on subsequent studies, thus serial imaging is helpful.26, 27,32,40 Other radiographic findings of diaphragm injury include basal lung opacity and abnormal hemidiaphragm contour. Rarely, herniated omental fat can simulate pleural fluid on chest radiographs.44 Radiographic findings are absent or nonspecific in most penetrating injuries, and early diagnosis typically requires direct inspection.45
The sensitivity and specificity of CT in diagnosing traumatic diaphragmatic hernias is approximately, 80% (range from 61 to 100%) and above 90% (range from 77 to 100%) respectively, better for left- than for right-sided ruptures.46–49 The disparate results seem to steam from the fact that most studies lack adequate power as surgically proven diaphragmatic rupture is rare. Intrapericardial herniation occurs rarely but may be demonstrated by CT.50, 51 The presence of any one of these signs indicate a substantial chance of diaphragm rupture:34–49,52–57
Contact of the upper third of the liver on the right or the stomach or bowel on the left with the posterior ribs (dependent viscera sign) (Figure 7)
Abdominal structures external to the diaphragm (Figure 7)
Focal constriction of the hernia contents (collar or hourglass sign) (Figure 7)
A focal bulge along the diaphragm (or cottage loaf sign, a variation of the collar sign) (Figure 8)
Linear lucency across the liver along the torn edge of the hemidiaphragm (band sign)
Abrupt discontinuity of the diaphragm (Figure 8), with or without visceral herniation (cautious with this sign as small defects are frequently seen in asymptomatic persons scanned for indications other than trauma)
Inability to identify the diaphragm (absent diaphragm sign) in an area where it does not contact another organ and should normally be seen
Acute arterial extravasation of contrast at the level of the diaphragm
Diffuse or focal thickening of the diaphragm Figure 9
Injured diaphragm curled inward from its normal course parallel to the body wall (dangling diaphragm sign)
Figure 7.
Initial (a) and post-admission (b) radiographs obtained in a male, who fell 30 feet show new apparent elevation of the left hemidiaphragm suspicious for interval development of a diaphragmatic hernia. Multiplanar reformatted CT (c–e) images reveal herniation of the stomach through a traumatic defect in the medial left hemidiaphragm (white arrows at margins of defect). Note constriction of stomach by the defect (collar sign) and contact of herniated portion of stomach with posterior chest wall (black arrows denote the dependent viscera sign).
Figure 8. .

Frontal chest radiograph in a female after a car accident shows normal diaphragmatic position. CT image obtained on admission shows thickening of the right crus (arrows) that can be interpreted as an indirect sign of traumatic rupture. Imaging obtained 1 week later reveals herniation of the liver with narrowing at the level of the diaphragm defect (collar sign).
Figure 9. .
Posteroanterior (a) chest radiographs of a patient who had sustained a left upper quadrant stab wound 2 years earlier shows apparent focal elevation of the anterolateral left hemidiaphragm (hump sign). CT image (b) shows the stomach (St) passing through defect in left hemidiaphragm (arrows at margins of defect). Note edema within fat adjacent to the anterior margin of the defect. A strangulated hernia was found at surgery.
Isolated small defects from penetrating trauma, in the absence of other injuries, may be difficult to detect in the absence of herniation. Such small defects may result in delayed herniation if they are not detected and repaired (Figure 7), therefore, laparoscopic or thoracoscopic evaluation may be indicated with certain injuries.58, 59 Cross sectional imaging such as MRI or CT with multiplanar reformatting (Figure 7) can be helpful.52 Use of MRI is generally limited to nonemergent settings where delayed diagnosis is sought.28,60–63
Other imaging techniques such as ultrasound can be of value in assessing the right hemidiaphragm by depicting the free edge of the diaphragm as a flap within pleural fluid29 or by demonstrating liver herniated into the chest.64 Scintigraphy can demonstrate traumatic herniation of the liver or spleen. Fluoroscopic upper or lower gastrointestinal studies may demonstrate herniated segments, particularly in delayed presentations.
Abnormalities of diaphragm position
Apparent or actual hemidiaphragm elevation has numerous causes (Table 1). Conversely, causes of hemidiaphragm depression (Table 2) are more limited.
Table 1.
Causes of hemidiaphragm elevation
| Unilateral |
| Volume loss (atelectasis, lobar collapse, partial lung resection, radiation fibrosis, congenital pulmonary hypoplasia, pleural encasement by tumor) |
| Eventration |
| Abdominal disease (dilated stomach or colon, hepatomegaly, splenomegaly, subphrenic abscess) |
| Phrenic nerve paralysis |
| Splinting (rib fracture, pneumonia, infarction, abscess, cholecystitis, peritonitis) |
| Mimics (subpulmonic pleural effusion, large pleural mass, diaphragmatic hernia) |
| Single-lung transplantation for pulmonary fibrosis |
| Phrenoplasty |
| Bilateral |
| Volume loss (suboptimal inspiration, supine positioning, atelectasis, lung resection, pulmonary fibrosis) |
| Abdominal mass effect (obesity, pregnancy, marked bowel dilation, ascites, hepatosplenomegaly, large abdominal tumor) |
| Eventration |
| Subpulmonic pleural effusion (mimics hemidiaphragm elevation) |
| Neuromuscular disease (quadriplegia, multiple sclerosis, amyotrophic lateral sclerosis, Guillain-Barré syndrome, myasthenia gravis, Eaton-Lambert syndrome, muscular dystrophy, steroid or alcohol myopathy, rhabdomyolysis) |
| Connective tissue disease (fibrosis in rheumatoid arthritis, scleroderma, and ankylosing spondylitis; weakness in systemic lupus erythematosus, polymyositis) |
| Endocrine and metabolic disorders (hypothyroidism, hyperthyroidism, Cushing’s disease, hypokalemia, hypophosphatemia, hypomagnesemia, metabolic alkalosis) |
| Phrenic nerve paralysis |
Table 2.
Causes of hemidiaphragm depression
| Unilateral |
| Large pneumothorax |
| Asymmetrical bullous emphysema |
| Large pleural effusion |
| Foreign body aspiration |
| Congenital lobar emphysema |
| Single-lung transplantation for emphysema |
| Bilateral |
| Chronic obstructive pulmonary disease (emphysema, asthma) |
| Deep inspiration (young, thin person) |
| Bilateral large pneumothorax |
| Bilateral large pleural effusion |
| Mechanical ventilation at high pressures |
| Cystic fibrosis |
| Pulmonary histiocytosis X |
| Lymphangioleiomyomatosis |
Paralysis of the diaphragm
Diaphragm paralysis may result from an abnormality at any point along its neuromuscular axis, may be unilateral or bilateral, and has numerous potential causes.7,65–68 Invasion by a malignant neoplasm and phrenic nerve trauma related to surgery (stretch, crush, or transection) (Figure 10) are common causes, although many cases are idiopathic. Central nervous system conditions also have been associated with bilateral hemidiaphragm paralysis. Hypothermic injury of the phrenic nerve related to the use of cold topical cardioplegia during coronary artery bypass surgery can lead to hemidiaphragm paralysis, usually on the left, which may persist for longer than 1 year.67 Diaphragm weakness without paralysis can be found in numerous conditions, including myopathies, connective tissue diseases, and various endocrine and metabolic disorders (Table 1).
Figure 10.

Radiograph obtained 1 month after left carotid endarterectomy shows left hemidiaphragm elevation that persisted and was suspicious for cervical phrenic nerve injury. Fluoroscopy confirmed left phrenic nerve paralysis.
Fluoroscopy is the most efficient method of assessing diaphragm motion. Other methods include ultrasound, comparison of radiographs obtained in full inspiration and expiration and MRI. With improved technology, MRI “sniff testing” is possible (Figure 11, Supplementary video 2). Unlike fluoroscopy, MRI offers the advantage of superior depiction of extradiaphragmatic pathologies while imparting no radiation on the patient. Similarly, sonography has recently become more popular for anatomic and functional diaphragmatic evaluation due to its radiation-free and mobile nature where exams can be performed by bedside. Ultrasound’s portability is particularly useful in assessing patients with respiratory failure following cardiothoracic surgery, where early intervention can facilitate successful weaning from mechanical ventilation and prevent ventilator-associated as well as lengthy hospitalization comorbidities. Ultrasound has been shown to be similar in accuracy to most other imaging modalities in diaphragm assessment.69 Unlike fluoroscopy which assesses the anterior central tendon, sonography focuses on the posterior and lateral diaphragm which are the muscular components innervated by the phrenic nerve and move more with respiration. Sonographic techniques have been described previously with B mode measuring diaphragmatic thickening while M mode assessing diaphragmatic excursion (Figure 12).69 A diaphragm thickening of less than 20% is proposed to be the cut-off for paralysis. Normal diaphragmatic excursion is greater than 2.5 cm (mean 3–5 cm, range 2–10 cm), but excursion of less than 3 cm is fairly frequent.8,70–75 Unequal excursion of the hemidiaphragms is common; the difference is usually less than 1.5 cm.73, 74 Asynchronous motion of the hemidiaphragms is not unusual.73, 76,77 M mode can also evaluate diaphragmatic velocity, which is correlated with muscle strength (Figure 12). Decrease diaphragm excursion on M mode has been shown to predict weaning failure equal to the rapid shallow breathing index.78
Figure 11.

Image from a MRI sniff test (Please see Supplementary Video 2, for full illustration) demonstrating MRI’s capability for visualizing extradiaphragmatic pathologies (note the left pleural effusion) in additional to assessing diaphragmatic function, which may explain or are associated with patient’s symptoms.
Figure 12.
M-mode illustration of right diaphragmatic paralysis (a). Notice the flattened waveform for diaphragmatic excursion on the right compared to the left (b). “y” denotes the amplitude of excursion (from trough to peak) and “x” represents the time frame used for diaphragm contraction (from trough to peak), which can be used to calculate velocity of movement. Velocity = “y” cm / “x” s.
Similar to most institutions, our fluoroscopic protocol is typically performed with the patient erect, but occasionally supine positioning is used as it stresses the diaphragm by removing the aid of gravity during inspiration and may increase the sensitivity of the test. The unilaterally paralyzed hemidiaphragm paradoxically moves upward on inspiration and downward on expiration, passively following changes in intrapleural and intra-abdominal pressure.7, 76 In bilateral paralysis, both hemidiaphragms move upward on inspiration, concomitant with inward rather than normal outward movement of the abdominal wall.79 However, a paralyzed hemidiaphragm may show a slight descent on slow, deep inspiration due to passive stretching as the rib cage expands. The sniff test is used to confirm that abnormal hemidiaphragm excursion is caused by paralysis rather than unilateral weakness. For the sniff test, the patient inhales rapidly and forcefully through the nose with the mouth closed. This normally produces a rapid, brief descent of both hemidiaphragms. Paradoxical upward motion of an entire hemidiaphragm (in oblique or lateral projection) of greater than 2 cm is consistent with hemidiaphragm paralysis.70
Several potential difficulties may limit the fluoroscopic assessment of diaphragm paralysis. Diaphragm motion may be diminished due to inflammatory processes such as pneumonia, pleuritis, pleural effusion, peritonitis, and subphrenic abscess, so fluoroscopic assessment is best delayed until such reversible conditions that may affect the diaphragm have resolved. Complete eventration may be difficult or impossible to distinguish from diaphragm paralysis,8 and severe weakness or fatigue may appear identical to bilateral paralysis on fluoroscopy.7 Although some patients with bilateral paralysis show the typical paradoxical upward motion of both hemidiaphragms during a deep inspiration or sniff, normal inspiratory descent of the diaphragm can be mimicked in those patients who perform a compensatory maneuver of actively exhaling below functional residual capacity using their abdominal muscles, then inhaling by relaxing the abdominal muscles, which causes passive descent of the diaphragm.80 This effect can be detected by carefully observing abdominal motion during breathing,81 and it can be minimized by performing the examination with the patient in the recumbent position, which eliminates the assistance of gravity.82 The diagnosis of diaphragm paralysis also can be difficult in patients with severe hyperinflation from chronic obstructive pulmonary disease, in whom the normal diaphragm moves very little, or in weak, debilitated patients who cannot produce a strong inspiratory effort or forceful sniff.
In some patients with phrenic nerve injury, even though fluoroscopy demonstrates paralysis, the paralysis may not be permanent. Regeneration of phrenic nerve fibers may lead to partial or complete recovery of diaphragm function over time. Based on the normal rate of peripheral nerve regeneration, this usually occurs within 1 year.83
Tumors of the diaphragm
Primary tumors of the diaphragm are very rare with benign and malignant tumors occurring with similar frequency and no predilection for laterality.80 The most common benign lesion of the diaphragm is a simple cyst (Figure 13), typically bronchogenic or mesothelial in origin.84 A mesothelial cyst can be found in both adults and children and typically resolve with or without ethanol injection.85 Lipomas are the most common solid benign masses (Figure 13). Lipomas measure fat density on CT and demonstrate signal loss on fat suppressed MRI. Unlike Bochdelak hernias, diaphragm lipomas do not disrupt diaphragm integrity. Diaphragm cysts and lipomas are generally asymptomatic, but when large in size, both can cause symptoms such as cough, pain, or dyspnea.
Figure 13.
Benign tumors of the diaphragm. CT (a) showing a small fat density lesion in the crus of the left hemidiaphragm consistent with a lipoma. T2 weighted (b) and post contrast T1 weighted (c) sequences demonstrate another small non-enhancing lesion with fluid signal consistent with a cyst (arrows).
Most malignant tumors of the diaphragm are sarcomas of fibrous or muscular origin.86 The most common malignant tumor is rhabdomyosarcoma (Figure 14), although these tumors are more likely to occur in the head and neck or urogenital regions. Rhabdomyosarcoma originate from primitive mesenchymal cells for skeletal muscle differentiation, with four major histologic types including embryonal cell (55%), botryoid (5%), alveolar (20%), and undifferentiated or pleomorphic (20%). Other tumors of the diaphragm include schwannoma,87 chondroma,88 pheochromocytoma,89 endometriosis,90 and hemangiopericytoma.91
Figure 14.
Posteroanterior (a) radiograph demonstrates an abnormal opacity along the cardiophrenic angle. Axial (b) CT reveals a heterogeneously enhancing mass centered on the anterior right hemidiaphragm with underlying mass effect on the liver. Final pathology is rhabdomyosarcoma.
The appearance of soft tissue tumors on cross-sectional imaging is usually nonspecific. Furthermore, because the diaphragm is a thin structure, the diaphragmatic origin of a mass may be difficult to confirm. Pulmonary, pleural, or abdominal tumors are far more common than primary diaphragm tumors so suspicion for metastasis should be high when encountering a diaphragm lesion. Thoracic or abdominal tumors may secondarily involve the diaphragm by direct extension.7 However, if thoracic or abdominal masses abut the diaphragm without traversing it on imaging studies, definitive diagnosis of invasion cannot be made.
CONCLUSION
Advancements in imaging have improved diagnosis of and aided in treatment of complicated diaphragmatic derangements. Increased awareness and recognition of what each imaging modality has to offer and familiarity with characteristic radiologic findings is crucial in diagnosis, guiding management and preventing complications in this population.
ACKNOWLEDGMENTS
We would like to acknowledge Ms Sonia Watson and Andrea Moran for their editorial contributions as well as Dr James Zheng from University of Saskatchewan for supplying figure 12.
Contributor Information
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