Planned versus actual coverage of the survey.
\\n\\n
More than half of the publishers listed alongside IntechOpen (18 out of 30) are Social Science and Humanities publishers. IntechOpen is an exception to this as a leader in not only Open Access content but Open Access content across all scientific disciplines, including Physical Sciences, Engineering and Technology, Health Sciences, Life Science, and Social Sciences and Humanities.
\\n\\nOur breakdown of titles published demonstrates this with 47% PET, 31% HS, 18% LS, and 4% SSH books published.
\\n\\n“Even though ItechOpen has shown the potential of sci-tech books using an OA approach,” other publishers “have shown little interest in OA books.”
\\n\\nAdditionally, each book published by IntechOpen contains original content and research findings.
\\n\\nWe are honored to be among such prestigious publishers and we hope to continue to spearhead that growth in our quest to promote Open Access as a true pioneer in OA book publishing.
\\n\\n\\n\\n
\\n"}]',published:!0,mainMedia:null},components:[{type:"htmlEditorComponent",content:'
Simba Information has released its Open Access Book Publishing 2020 - 2024 report and has again identified IntechOpen as the world’s largest Open Access book publisher by title count.
\n\nSimba Information is a leading provider for market intelligence and forecasts in the media and publishing industry. The report, published every year, provides an overview and financial outlook for the global professional e-book publishing market.
\n\nIntechOpen, De Gruyter, and Frontiers are the largest OA book publishers by title count, with IntechOpen coming in at first place with 5,101 OA books published, a good 1,782 titles ahead of the nearest competitor.
\n\nSince the first Open Access Book Publishing report published in 2016, IntechOpen has held the top stop each year.
\n\n\n\nMore than half of the publishers listed alongside IntechOpen (18 out of 30) are Social Science and Humanities publishers. IntechOpen is an exception to this as a leader in not only Open Access content but Open Access content across all scientific disciplines, including Physical Sciences, Engineering and Technology, Health Sciences, Life Science, and Social Sciences and Humanities.
\n\nOur breakdown of titles published demonstrates this with 47% PET, 31% HS, 18% LS, and 4% SSH books published.
\n\n“Even though ItechOpen has shown the potential of sci-tech books using an OA approach,” other publishers “have shown little interest in OA books.”
\n\nAdditionally, each book published by IntechOpen contains original content and research findings.
\n\nWe are honored to be among such prestigious publishers and we hope to continue to spearhead that growth in our quest to promote Open Access as a true pioneer in OA book publishing.
\n\n\n\n
\n'}],latestNews:[{slug:"stanford-university-identifies-top-2-scientists-over-1-000-are-intechopen-authors-and-editors-20210122",title:"Stanford University Identifies Top 2% Scientists, Over 1,000 are IntechOpen Authors and Editors"},{slug:"intechopen-authors-included-in-the-highly-cited-researchers-list-for-2020-20210121",title:"IntechOpen Authors Included in the Highly Cited Researchers List for 2020"},{slug:"intechopen-maintains-position-as-the-world-s-largest-oa-book-publisher-20201218",title:"IntechOpen Maintains Position as the World’s Largest OA Book Publisher"},{slug:"all-intechopen-books-available-on-perlego-20201215",title:"All IntechOpen Books Available on Perlego"},{slug:"oiv-awards-recognizes-intechopen-s-editors-20201127",title:"OIV Awards Recognizes IntechOpen's Editors"},{slug:"intechopen-joins-crossref-s-initiative-for-open-abstracts-i4oa-to-boost-the-discovery-of-research-20201005",title:"IntechOpen joins Crossref's Initiative for Open Abstracts (I4OA) to Boost the Discovery of Research"},{slug:"intechopen-hits-milestone-5-000-open-access-books-published-20200908",title:"IntechOpen hits milestone: 5,000 Open Access books published!"},{slug:"intechopen-books-hosted-on-the-mathworks-book-program-20200819",title:"IntechOpen Books Hosted on the MathWorks Book Program"}]},book:{item:{type:"book",id:"86",leadTitle:null,fullTitle:"Optoelectronic Devices and Properties",title:"Optoelectronic Devices and Properties",subtitle:null,reviewType:"peer-reviewed",abstract:"Optoelectronic devices impact many areas of society, from simple household appliances and multimedia systems to communications, computing, spatial scanning, optical monitoring, 3D measurements and medical instruments. This is the most complete book about optoelectromechanic systems and semiconductor optoelectronic devices; it provides an accessible, well-organized overview of optoelectronic devices and properties that emphasizes basic principles.",isbn:null,printIsbn:"978-953-307-204-3",pdfIsbn:"978-953-51-4906-4",doi:"10.5772/618",price:159,priceEur:175,priceUsd:205,slug:"optoelectronic-devices-and-properties",numberOfPages:676,isOpenForSubmission:!1,isInWos:1,hash:"aa120b5e14a2c88603b54cc31a2d953e",bookSignature:"Oleg Sergiyenko",publishedDate:"April 19th 2011",coverURL:"https://cdn.intechopen.com/books/images_new/86.jpg",numberOfDownloads:85967,numberOfWosCitations:57,numberOfCrossrefCitations:25,numberOfDimensionsCitations:52,hasAltmetrics:1,numberOfTotalCitations:134,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"May 19th 2010",dateEndSecondStepPublish:"June 16th 2010",dateEndThirdStepPublish:"October 21st 2010",dateEndFourthStepPublish:"November 20th 2010",dateEndFifthStepPublish:"January 19th 2011",currentStepOfPublishingProcess:5,indexedIn:"1,2,3,4,5,6,7",editedByType:"Edited by",kuFlag:!1,editors:[{id:"58036",title:"Dr.",name:"Oleg",middleName:null,surname:"Sergiyenko",slug:"oleg-sergiyenko",fullName:"Oleg Sergiyenko",profilePictureURL:"https://mts.intechopen.com/storage/users/58036/images/5755_n.jpg",biography:"Oleg Yu. Sergiyenko received his BS and MS degrees from Kharkiv National University of Automobiles and Highways, Kharkiv, Ukraine, in 1991 and 1993, respectively. He received his PhD degree from Kharkiv National Polytechnic University in 1997.\nHe has written 81 papers in control systems, robot navigation, 3D coordinate measurement, and SHM. He is an editor of two books, holds two patents in Ukraine and Mexico, and is a reviewer for various journals. 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A Description of the Pathways and Organelles Involved on These Processes",doi:"10.5772/intechopen.86046",slug:"how-does-the-main-infective-stage-of-em-t-cruzi-em-enter-and-avoid-degradation-in-host-cells-a-descr",body:'\n
Trypanosoma cruzi, the etiological agent of Chagas disease, causes an anthropozoonosis discovered and characterized by Dr. Carlos Chagas in 1909 [1] and recognized by the World Health Organization (WHO) as one of the three most neglected tropical diseases of the world [2, 3, 4]. Nowadays, up to 8 million people are estimated to be infected with Chagas disease only in the Americas. Patients who do not receive treatment can develop severe cardiac debility and gastrointestinal organ dysfunction and may die, and 25 million are at risk of contracting the disease [3, 4]. Due to population migration and specific modes of transmission, Chagas disease is spreading beyond its natural geographical boundaries and becoming a global problem [5]. Although the protozoan has three major developmental stages, only two are capable of infecting mammals (trypomastigotes and amastigotes), and the trypomastigote stage is the main T. cruzi vector [6]. Recent data indicate the existence of an infective epimastigote-like stage observed in axenic cultures as well as in the invertebrate host [7]. It is important to point out that before publication of this article, it was assumed that replicating epimastigotes present in the insect gut are not infective to mammalian host. During the vector infection (caused by a hematophagous insect of the family Reduviidae), metacyclic trypomastigotes [8], which penetrate the vertebrate host (several mammals, including man), are released along with their excreta coming in contact with conjunctiva areas or through small lesions in the own site of the bite (favored by the itch caused after the insect’s bite). In turn, metacyclic trypomastigotes are able to invade virtually all cell types in the vertebrate host, especially muscle cells, fibroblasts, and macrophages [6]. At this moment, the intracellular cycle of T. cruzi begins, where the firing of several signaling cascades culminates with the closure of the parasitophorous vacuole (PV) where the parasite is found [9, 10]. After the PV closure, the process of differentiation of the parasite from the trypomastigote stage to the amastigote stage begins. At the same time, fragmentation of the PV membrane takes place most probably due to the increased concentration of the Tc-Tox perforin-like protein produced by the parasite [11]. After the destruction of the vacuole, the parasite, in the process of differentiation, will be found in the cytoplasm of the host cell where it will initiate its multiplication and subsequent differentiation for trypomastigotes culminating in the rupture of the host cell (Figure 1) [13]. The whole process of formation of the parasitophorous vacuole until its rupture counts on the participation of several organelles of the host cell. Among these, the best characterized is the participation of host cell endosomes and lysosomes. It is the fusion of these organelles with the PV membrane that probably allows the increase or expansion of the PV. In addition, this process is also responsible for the generation of an acidic environment within the PV, which probably will potentiate the action of Tc-Tox and PV membrane fragmentation [13]. Wilkowsky and colleagues [14] have shown that early and late endosomes were critical for vacuole formation. In addition, other organelles responsible for the production of proteins and energy (endoplasmic reticulum (ER), Golgi complex (GC), and mitochondria) have also been observed during the initial infection process [11]. In this chapter we will discuss the available data on the process of parasite-host cell recognition, triggering of the internalization process, and biogenesis of the PV. A better understanding of all the processes may identify new potential targets to block parasite invasion and may constitute alternative ways to treat Chagas disease.
\nIntracellular cycle of T. cruzi. (A) Attachment of the trypomastigote. (B) Internalization and recruitment and fusion of host cell lysosomes. (C) Parasitophorous vacuole is formed and lysosomal content is released. (D) Differentiation of trypomastigote to amastigotes. (E) Parasitophorous vacuole membrane disintegration. (F) Amastigote division into cytoplasm. (G) Differentiation of amastigotes into trypomastigotes. (H) Liberation of parasites into extracellular [12].
The trypomastigote stage of T. cruzi has the ability to invade both professional phagocytic cells and nonprofessional phagocytic mammalian cells either in vitro or in vivo [15, 16, 17]. The kinetics of the T. cruzi’s intracellular cycle varies according to the strain, including time duration (Y strain, e.g., has an intracellular cycle that is completed between 5 and 7 days), since the initial interaction with trypomastigotes, triggering a signaling cascade that culminates with the formation and closure of the PV where the parasite will be located for some time [18].
\nIn order for the interaction process and consequent internalization of the parasite to occur, there must be a recognition between molecules present on the surface of both cells. These processes are complex and involve several adhesion molecules (Figure 2), signaling events, and proteolytic activities [10, 19]. Some of these molecules have been identified as participants in the adhesion and invasion processes, such as gp35/50 [20], gp82 (whose expression varies according to the T. cruzi strain analyzed) [21], and gp90 [22]. Both gp82 and gp90 are the main metacyclic stage-specific surface molecules and are extremely important to oral infection [21, 23]. However, while glycoprotein 90 is known as a negative modulator of metacyclic trypomastigote invasion [24, 25], gp82 binds to an unidentified receptor in host cell surface mediating a calcium-dependent signaling pathway that will be discussed below [20, 26]. Although the receptors presented in host cell surface that recognize gp82 and gp90 are not known, some data reinforce they are different since the target cell’s interaction motif of both glycoproteins is distinct [26]. It is important to point out that these molecules are not present in tissue culture-derived trypomastigotes. At this stage, the molecules described as present in the membrane are gp85 (recognizes extracellular matrix), Ssp3, shed acute-phase antigen (SAPA) (trans-sialidase (TS)), oligopeptidases (serine proteases), and penetrin, which bind to heparin, heparan sulfate, and collagen [10].
\nSchematic model of molecules involved in T. cruzi-host cell interaction [10].
Expression of a family of GPI-anchored glycoproteins, termed trans-sialidases/neuraminidases, present mainly on the surface of trypomastigotes is capable of modifying the exposure of surface glycoconjugates both in host cells and in the parasite itself. T. cruzi trans-sialidases (TS) are “shed” from the parasite membrane to the external medium through microvesicles. Microvesicles have a complex lipid bilayer and are responsible for carrying several molecules derived from the parasite, such as lipids, proteins, and nucleic acids [27, 28, 29]. Trypomastigote TS protein structure is formed by two major regions: an N-terminal catalytic region and a C-terminal region. C-terminal presents 12 amino acids repeated in tandem, named SAPA. TS can be active or inactive. Inactive TS are lectin-like proteins and are capable to bind SAcs and β-galactose [30]. Active TS are modified sialidases which, in addition to cleaving sialic acid, may transfer from sialoglycoconjugates of the host cell to β-galactose of T. cruzi glycoconjugates [31]. The TS gene family is complex, consisting of fourteen members divided into two groups: one translates TS present in stages present in mammalian hosts (trypomastigotes), and another translocates TS present in specific vector stages (epimastigotes). Although biochemical studies do not demonstrate the presence of TS at the amastigote stage, immunocytochemical assays have shown that TS can indeed be expressed in amastigotes [32, 33, 34]. Pereira et al. [35] observed that about 20–30% of cultured trypomastigotes had trans-sialidases/neuraminidases on their surface and these parasites were more invasive than the population that did not express the enzyme. Expression of trans-sialidases in T. cruzi is directly related to infectivity/virulence of the parasite since proteome studies indicate that different strains exhibit significantly different amounts of TS and TS-containing microvesicles and that host cells incubated with larger quantities containing microvesicles of TS before infection with trypomastigotes will generate a greater infection [36]. Results regarding virulence had already been related to TS expression through the analysis of virulent and non-virulent strains by transcriptome. In this work it can be observed that the expression of TS-coding genes during the differentiation process from intracellular amastigote to trypomastigote (end of intracellular cycle) is much higher in the virulent than in the non-virulent strain. Associated with this the transfection of avirulent strain with gene allowing the constitutive expression of TS also makes the release of trypomastigotes faster [37]. The TS family also includes members that have no enzymatic activity but which may also be involved in recognition between the parasite and the host cell, such as gp85 [38]. Todeschini and colleagues [39] demonstrated that inactive enzymes of the TS family are sialic acid-binding proteins and terminal β-galactopyranose (βGalp) residues. In relation to gp85/trans-sialidase, San Francisco and colleagues [40] demonstrate that this protein plays a fundamental importance in invasion since its depletion causes a decrease in T. cruzi virulence. The same type of result was reported by Pascuale et al. [41] since inactive TS expression in trypomastigotes of a strain that does not express these TS (iTS null) allowed a better invasion and increase of the parasitic load in mice demonstrating that the inactive form may act alternatively or complementing the active TS in pathogenesis.
\nSialic acid from the host cells has a crucial importance in intracellular cycle of T. cruzi, a parasite that does not have the ability to synthesize sialic acids. This molecule plays an important role in protecting T. cruzi from lysis by serum factors and also acting in interaction with host cell. Mucins and TS function as substrate and enzyme, respectively, and sialylated mucins are localized in microdomain regions of trypomastigotes [42]. During the interaction process involving macrophages, the presence of sialic acid on the surface of trypomastigotes hinders the invasion process, since the removal of these residues through the use of neuraminidase or their blockage through the use of periodic acid or lectins from Limax flavus or Limulus polyphemus increases adhesion and internalization rates. It is possible that the presence of desialylated parasites in macrophages may increase due to the recognition of galactose/N-acetyl galactosamine receptors present on the surface of macrophages [15, 16]. In relation to sialic acid present on the surface of macrophages, there has been an increase in the entry of trypomastigote forms in cells that expose this residue when compared to cells that do not expose galactose [15, 43]. The presence of sialic acid on the surface of trypomastigotes does not yet have a fully known function though it is believed that sialic acid helps adhesion and penetration into non-phagocytic cells [34, 44].
\nIt is currently discussed that any class of molecules exposed on the surface of mammalian cells has a great receptor potential for molecules exposed on the surface of T. cruzi [19]. Most of the receptor classes have carbohydrates in their composition, such as galactose, mannose, sialic acid residues [19, 44, 45], and lectin-type proteins such as galectins (binding to carbohydrate residues present on the surface of the parasite). Pineda et al. [46] described binding data from different human galectins (gal-1, gal-3, gal-4, gal-7, and gal-8) to different strains of T. cruzi belonging to the six different strains (DTUs). It has been observed that all galectins bind preferentially to the infective stages (amastigotes and trypomastigotes) and that many can promote higher rates of adhesion and infection to host cells and higher rates of infection to mice. In relation to galectin-1, it was observed that the presence of this glycoprotein in human and murine cardiomyocytes is able to prevent infection with trypomastigotes, one more data that goes against the modulating role of galectin in the process of internalization of trypomastigotes [47].
\nGalectin-3, a protein abundant in the cytoplasm of epithelial cells and macrophages, has also been described as a participant in the immune response and infection processes (in addition to the recognition process between the host and T. cruzi) [48, 49]. In addition, it has also been demonstrated that galectin-3 accumulates in both the parasite and phagosome entry regions and in tubules and vesicles that would derive from the endosomal system, thus suggesting that this protein is also an excellent marker of the lysis process of the PV containing this parasite [50]. The absence of galectin-3, in addition to increasing intracellular replication in vitro, is able to increase parasitemia in vivo by decreasing the secretion of pro-inflammatory cytokines and increasing cardiac fibrosis [51], which confirms the important role of this glycoprotein in the pathogenesis of Chagas disease.
\nIn addition to all the molecules already mentioned, they also act as endothelin-1 receptors and bradykinin receptors. Both are used by trypomastigotes in the invasion of cardiovascular cells, being very important in the pathogenesis of Chagas disease [52]. Cytokeratin 18, fibronectin, laminin, and integrin are also recognized by Tc-85, forming a bridge between the parasite and the host cell [19, 53, 54]. Besides, a novel family of T. cruzi surface membrane proteins (designated as TcSMP) was detected in parasite surface and plays some role on host cell invasion by T. cruzi [54].
\nThe mechanisms that lead to the internalization of trypomastigotes appear to be different when one considers the cell type where the internalization will occur. Morphological evidence shows that the parasite invades the host cell by an endocytic process that culminates in the formation of a PV. In cells of the immune system such as macrophages, which are specialized in phagocytosis events, we observe that the process of internalization occurs in two distinct ways. Dvorak and Schumunis [55] initially suggested that trypomastigotes forced the membrane of the host cell in an event where there is energy expenditure by the two cells involved. Nogueira and Cohn [56] observed the formation of projections on the surface of the host cell, which leads to a classical phagocytic process.
\nIn cells considered as nonprofessional phagocytic, such as epithelial cells and fibroblasts, there appears to be a process of internalization where the parasite is the agent of penetration [33]. This process was confirmed by Martins et al. [57], where it was shown that metacyclic trypomastigotes (G and CL strains) require ATP to invade nonprofessional phagocytic cells. In these cell types, two different strategies are known to be involved in the invasion process and formation of the PV: one dependent on lysosomes and another one independent of lysosomes. The lysosomal-dependent pathway, first described by Tardieux et al. [58], was well characterized in nonprofessional phagocytic cell lines. Signals triggered by the recognition between T. cruzi and the host cell lead to the recruitment of lysosomes to the parasite’s entry site, which would actively participate in the invasion process [33]. In addition, lysosome exocytosis would depend (1) on the performance of microtubules [8] and (2) on the regulation of host cell cytoplasmic Ca2+ levels [15]. This process of lysosomal-dependent invasion would occur in about 20% of the parasites [59]. The fusion of the lysosomes with the plasma membrane would thus end up donating the membrane for the formation of the PV in a calcium-dependent process. In addition, a similar Ca2+-dependent lysosomal exocytosis mechanism was observed during injury and repair of the plasma membrane [60]. Tam et al. [61] demonstrated that the mechanism of injury-dependent endocytosis is directly related to the secretion of a specific lysosomal enzyme and acid sphingomyelinase (ASM). ASM would cleave sphingomyelin, an abundant sphingolipid in the outer leaflet of the plasma membrane [62], forming ceramide, which is internalized by the cell [63, 64, 65]. Trypomastigotes are able to take advantage of this mechanism by inducing the formation of these vesicles rich in ceramides, thus facilitating the invasion process [66]. Over the years, this entry mechanism involving plasma membrane repair pathways has been extensively studied by several groups. The participation of lysosomal proteins known as Lamp1 and Lamp2 has been shown to be essential, since the knockout of both proteins decreases the entry of trypomastigotes. This decrease in entry is not accompanied by inhibition of lysosome exocytosis to the repair region as well as phenotypic modification of the host cell or generated PV. It is believed that the decrease in parasite entry is due to problems in caveolin-mediated endocytosis and in calcium efflux [67]. Considering that there are two distinct pools of lysosomes (cortical and internal) in mammalian cells, Hissa and Andrade used cardiomyocytes and observed, through the use of specific inhibitors, that trypomastigotes enter into this cell type mainly recruiting lysosomes from the more internal cell layer (perinuclear) [52]. Some molecules known as SAPs are secreted by microvesicles which are recognized by host cells and promote an efflux of perinuclear lysosomes. These molecules probably act together with gp82, activating Ca2+ pathway and promoting T. cruzi internalization [68]. Another molecule capable to recruit lysosomes to entry site is oligopeptidase also involved in Ca2+ efflux [69]. Using non-phagocytic cells it is described that the entry of metacyclic trypomastigotes is predominantly due to recruitment of lysosomes, whereas entry of cultured trypomastigotes (from the same CL strain) does not involve the participation of lysosomes [70]. Recently, Rodrigues et al. [26] observed that infection of host cells by metacyclic trypomastigotes is associated with lysosome spreading and presence of gp90 (metacyclic trypomastigote surface glycoprotein). Strains expressing low amount of surface gp90 are able to recruit more lysosomes to the site of infection giving rise to a more successful infection.
\nIn all other mechanisms used by the parasites, there would be no recruitment and exocytosis of lysosomes to the entry site, which are classified as lysosomal-independent mechanisms. In these pathways, there is an invagination of the plasma membrane of the host cell with the consequent formation of a PV without the initial presence of lysosomal markers. In 2002, Wilkowsky et al. [14] demonstrated the existence of this lysosomal-independent pathway for the invasion of T. cruzi into two nonprofessional phagocytic cell lines. Using Hela and CHO cells transfected with Rab5, Rab7, or dinamine-GFP, the presence of these endosomal markers in the newly formed vacuole was observed, indicating that some vacuoles fuse first with early and late endosomes and not with lysosomes. Later these events were quantified, and 50% of the parasitophorous vacuoles formed used the plasma membrane of the host cell but were enriched with PI3-kinase action products and negative for endosomal markers; 20% of the other trypomastigote-containing vacuoles were positive for EEA-1 (Rab5 effector and marker of initial endosomes) and Rab5, and approximately 20% of the vacuoles were positive for LAMP-1 (lysosomal marker). Vacuoles from these pathways are matured by the gradual fusion of early endosomes as well as of lysosomes, which allows the complete formation of the PV [16, 19, 71]. Both models of interaction are illustrated in Figure 3. In addition to the endolysosomal system, the cytoskeleton and autophagic processes were also related to the complex formation process of T. cruzi parasitophorous vacuoles, but there is still much controversy about these involvements. The first time that a compound known to interfere with actin polymerization inhibitor was used with the aim of studying the entry of T cruzi was reported by Nogueira and Cohn [56]. Subsequently, Meirelles et al. [72] using the same compound demonstrated that the treatment of chicken macrophages with cytochalasin prevented the entry of the parasite without, however, preventing adhesion. This work was the first to split the entry’s phase of T. cruzi (adhesion and internalization). On the other hand, using another cytochalasin (cytochalasin D—an actin filament polymerization inhibitor) as a tool, it was observed that the infection rate of the parasite was not significantly altered [33]. Subsequently the Tardieux group [58] demonstrated that invasion of T. cruzi into nonprofessional phagocytic cells is significantly enhanced by the depolymerization of the host cell actin cytoskeleton. The rapid reorganization of actin occurs as a response to the trypomastigote stage, suggesting that the direct reorganization of the actin cytoskeleton is a critical step for the entry process [18]. The reduction in the entry process was again observed in other studies, indicating that this divergence may be related to the observed interaction time [73].
\nSchematic model of T. cruzi internalization and parasitophorous vacuole formation using different entry pathways [10].
In relation to the microtubules, their dynamics are important to facilitate T. cruzi invasion targeting lysosomes to entry site [57, 74]. It is also believed that the PV acts as a secondary center for the organization of microtubules, as regards the lysosomal fusion process at the parasite’s entry site [74]. Besides a protein known as a CLASP1 (a microtubule plus-end tracking protein) is described as involved in the internalization of T. cruzi integrating actin pathway with microtubules and helping with the perinuclear localization of PV [75]. Microtubules are also responsible for carrying vesicles such as those positive for Vamp7 (essential for lysosomal fusion and retention of infection) to the entry site. This transport is dependent on the KIF 5 protein (a kinesin) [76]. More recently, Romano et al. [77] have shown that an autophagic protein, LC3, would also be present in the PV membrane, also demonstrating that the induction of autophagy in the host cell (pharmacological or physiological) interferes with the mechanism of trypomastigote invasion. Autophagy mechanism also plays a protective role against T. cruzi infection in mice by activation of a host immune response [78].
\nMany processes are already described as involved in the entry of T. cruzi trypomastigotes. As already discussed in this topic since 1972, Dvorak and Schumunis [55] had already described that the internalization of trypomastigotes in host cells could be by endocytic mechanisms. Endocytic processes are currently divided into different classes: clathrin-mediated endocytosis, endocytosis mediated by membrane microdomains (planar and caveolae), macropinocytosis, and phagocytosis. The first endocytic mechanism described as a participant in T. cruzi entry into host cells was phagocytosis. The participation of this mechanism was described by Nogueira and Cohn [56] through the treatment of several cell types, peritoneal macrophages, L929, HeLa, and embryo fibroblasts of calves with cytochalasin B (a drug that interferes with the extension of actin filaments). Afterward, Barbosa et al. [79] demonstrated by transmission electron microscopy that trypomastigotes are able to bind to cardiac muscle cells and induce the formation of extensive pseudopodia, a typical feature of phagocytic processes. Subsequently several reports showed that actin filaments are essential in other endocytic mechanisms, such as the macropinocytosis. Cytochalasins B and D are quite specific inhibitors of phagocytic activity. When the parasite is internalized via phagocytosis, there is internalization of CR3 receptors, β1 integrin, lysosomal membrane glycoproteins (Lpg), and Fc receptors (the latter appears only when trypomastigotes are opsonized). The participation of toll-2 receptors (“toll-like receptors 2”), as well as membrane components containing galactosyl, sialoconjugate, and glycoconjugate residues [80, 81, 82], is also demonstrated. In general phagocytosis is also a process that can be divided into different subtypes that are morphologically distinct. Using scanning electron microscopy, it was demonstrated that during internalization of trypomastigotes, the plasma membrane of peritoneal macrophages can cover the parasite in a juxtaposed way, with bilateral projections of plasma membrane forming a funnel-like structure that can follow the entire extent of the parasite’s body, culminating in its total internalization (similar to what is described as classical phagocytosis). It has also been observed in the development of structures similar to the initial stages of trigger phagocytosis or macropinocytosis, in addition to the formation of structures described as coiled-type phagocytosis [19, 82, 83].
\nThe participation of membrane rafts in the invasion of T. cruzi has also been demonstrated [84, 85]. Regions of membrane microdomains (rafts) are small, dynamic, cholesterol-rich membrane invagination regions, where sphingolipids, GM1 gangliosides, and caveolae (caveolae are a special type of membrane rafts) concentrate. These regions are known as signaling hotspots because they contain several proteins that can be deposited by triggering signaling cascades. This topic (signaling activation in T. cruzi entry process) will be discussed later in this chapter. More recently it has been demonstrated that T. cruzi could also use another endocytic mechanism in the invasion of its host cell: macropinocytosis. [86]. This endocytic pathway involves the internalization of large areas of plasma membrane along with significant amounts of extracellular fluid. It is important to point out the participation of dynamin as a key protein for the formation and consequent release of the early PV from the plasma membrane. Its inhibition using dynasore or its blockage through an overexpression of a dominant-negative mutant of dynamin inhibits the internalization of trypomastigotes, demonstrating that GTPase activity is also important [71, 86].
\nThere are several external factors that can regulate different types of cellular responses. For these responses to occur, it is necessary that a conformational change of several proteins takes place, which means that they can now interact with other molecules leading to their activation, transducing the signal and amplifying it. The conformational change is usually dependent on the action of protein kinases or protein phosphatases. The case of the process of interaction between Trypanosoma cruzi and host cells is not different since it is also an external process that will require an internal response. From this perspective, several studies have focused on the investigation of different pathways that coordinate the invasion of T. cruzi and that modulate the gene expression of the host cell in response to this process [85, 86].
\nT. cruzi seems to exploit an infinity of cell surface receptors, secondary messengers, and transcription factors of different pathways to ensure its invasion and survival [86, 87, 88, 89]. Among the signaling events, the best studied is that which leads to a calcium release in the host cells. This calcium release is one of the main responsible for regulating the process of invasion of T. cruzi [9, 18]. Three different models have already been proposed by different groups as being responsible for the activation of this signaling pathway. Among the described models, two involve peptidases such as oligopeptidase B and cruzipain, and the third one involves a membrane glycoprotein called gp82. Although all three mechanisms activate calcium firing, none of them are correlated. The first proposed model is based on the activation of the serine peptidase called oligopeptidase B where this enzyme present in the trypomastigote cytosol cleaves an inactive precursor to generate an active calcium agonist that is released by the parasite and binds to the receptor present on the surface of the host cell. This receptor is coupled to G protein which stimulates phospholipase C activity generating inositol 3,4,5-trisphosphate which binds to its receptor releasing calcium [90]. The second mechanism, proposed by Scharfstein et al. [91], is based on the secretion of cruzipain through the flagellar pocket region. This protein binds to a kininogen molecule that is cleaved into short kinin molecules, which in turn bind to the bradykinin receptor by stimulating the release of calcium from IP3. Cruzipain is also capable to regulate arginase activity increasing T. cruzi survival inside the cell through an increase in the production of IL-10 and TGF-β [92]. The model based on the activation of the glycoprotein gp82 is known to be bidirectional since it has been shown that a peak of calcium is generated not only in the host cell but also in the parasite itself. In the case of activation from this glycoprotein, the receptor is not yet known [90].
\nAs previously described one of the activated pathways is TGF-β, where molecules secreted by trypomastigotes stimulate TGF-β receptors and activate the transcription of genes regulated by this molecule [93]. Activation of this pathway is involved in Chagas disease fibrosis development [94, 95, 96]. TGF-β was first described as being activated through cruzipain, but it is now known that trypomastigotes are capable of exposing phosphatidylserine to the outer layer of the plasma membrane. This exposure would be responsible for triggering the TGF signaling pathway in macrophages, based on the phosphorylated Smad2 nuclear translocation, leading to inhibition of iNOS in infected macrophages. This event would favor intracellular survival of the parasite [97]. More recently, the mechanism used to favor this intracellular survival was proposal by Calvet and colleagues [98] using cardiomyocytes as host cell model. In these cells the TGF-β receptor (TβRII) is localized in cardiomyocyte’s costameres, which are also rich in vinculin and associated with cytoskeleton (known as a signaling domain). Its activation potentiates Smad2 phosphorylation. When T. cruzi infection is established, the cytoskeleton is disorganized, disrupting TβRII striations and decreasing Smad2 phosphorylation making cardiomyocytes less responsive to exogenous TGF-β stimulation.
\nPhosphorylation of protein tyrosine kinases is an important step in the regulation of a variety of eukaryotic cell signaling pathways [99]. In professional phagocytes, the entry of T. cruzi into macrophages is inhibited by treatment with genistein, a tyrosine kinase inhibitor [79]. It has also been shown that trypomastigotes of the Y strain stimulate tyrosine phosphorylation of a large number of proteins [100]. Pretreatment of fibroblasts with genistein does not inhibit entry of the parasite [18], suggesting different roles for protein tyrosine kinases in T. cruzi invasion of the Y strain in professional phagocytic cells or nonprofessional phagocytic cells. In cardiomyocytes and cardiac fibroblasts, tyrosine kinases appear to be essential to infection. Tyrosine kinase C (TrkC) is recognized by T. cruzi parasite-derived neurotrophic factor (PDNF) through neurotrophin receptor culminating in the entry of trypomastigotes into cardiac cells, while TrkA activation by the same ligand in the same cell types leads to a decrease in oxidative stress [101]. In this same sense of protection from the T. cruzi infection, other signaling pathways are also activated as, for example, the pathways of Erk11/Erk2 and Jak/STATs [102]. Other protein kinases also participate in T. cruzi invasion in host cells, such as protein kinase C (PKC), MAP kinases, and phosphatidylinositol 3-kinases (PI3-K) [86, 88, 97]. Recently the regions known as membrane microdomains have been described as signaling platforms. These regions are capable of recruiting a wide range of proteins involved in signal transduction processes. These proteins may include tyrosine kinase receptors and protein kinases such as PI3 kinase, protein kinase C, Src kinase (Lyn and Fyn) family proteins, FAK, bradykinin receptors, GTP (Rac, Rho, and Ras), and adapter proteins (Vav, Sos, and Shc). Some of these molecules have already had their share in the process of invasion of T. cruzi elucidated, while others have not. Proteins such as those from the Src kinases family (responsible for ITAM phosphorylation, which is essential for initiating the signal transduction cascade that triggers pathogen growth) and adapter proteins such as Vav, although not demonstrated as participants in this process, have been described by Vieira et al. [88] as possible phosphorylated proteins during the T. cruzi invasion in macrophages. Signaling pathways described as involved in macropinocytosis pathway (Pak1 and PKC pathways) are important to the intracellular development of infection [19]. More recently, Wnt signaling also has been shown to be an important pathway to immunomodulatory functions during T. cruzi infection, regulating the control of parasite replication. Activation of Wnt pathway is important to avoid a production of pro-inflammatory cytokines and indoleamine 2,3-dioxygenase activity by T. cruzi, helping to control the infection [103].
\nAfter the internalization process, T. cruzi resides obligatorily, and temporarily, inside a parasitophorous vacuole [10]. Over the years, several groups have described that the formation of the vacuole membrane is a complex process related to the participation of numerous molecules, depending also on the type of host cell involved in the process, i.e., phagocytic lines or nonprofessional phagocytic lines. As regards the formation of the PV membrane, it has been previously discussed in this text that several components of the host cell have already been described as components of the cytoskeleton [19, 73, 104, 105]. The organelle known as endoplasmic reticulum (ER) has been described in the literature as a donor membrane for phagosome formation, binding directly to the base of this phagosome in formation [106]. Recently, our group demonstrated by electron tomography (followed by 3-D reconstruction) and fluorescence videomicroscopy that the endoplasmic reticulum (ER) participates in the process of formation of the vacuole from initial moments until its complete maturation. The participation or not of ER in this process may be directly related to the input mechanism used by the parasite at the time of the invasion [104].
\nThe Golgi complex (CG) plays a central role in eukaryotic cells, serving as an intermediary and bidirectional axis of protein and lipid trafficking in the endo-membrane system [107]. In the case of T. cruzi, Carvalho et al. [82] described that after long infection times, there would be no change in GC distribution, remaining in the perinuclear region. Recently, transmission electron microscopy showed an intimate proximity between the CG and the parasite-containing vacuole. These data are of extreme relevance due to the function of GC in eukaryotic cells, i.e., because the membrane of the PV containing the parasite is still in the process of formation, requiring lipids and proteins from the host cell that would aid its closure as well as its expansion. As regards mitochondria, apparently there is no direct link between this organelle and the vacuole [104].
\nAs discussed above trypomastigotes of T. cruzi use different molecules and mechanisms to invade a host cell. Regardless of the chosen mechanism (participation of plasma membrane components or lysosomes at the site of invasion), the parasite will be located inside a vacuole.
\nInside the PV the trypomastigotes release the enzyme trans-sialidase/neuraminidase, which is responsible for the removal of the sialic acid residues from the vacuole membrane. This removal makes it sensitive to the action of another enzyme, homologous to factor 9 of human complement, Tc-Tox. The lysis of the vacuole membrane by the action of Tc-Tox may be associated with the formation of pores in this membrane, which, together with the secretion by the trans-sialidase/neuraminidase parasite, will lead to the complete fragmentation of the PV membrane [35, 108].
\nDespite the attempt to determine the exact moment of the T. cruzi exit process from the parasitophorous vacuole, this step has not yet been well elucidated. Since 1989, however, our group has been trying to chart the paths necessary for this discovery. Initially Carvalho and De Souza [109] demonstrated by transmission electron microscopy that the lysing process of the PV membrane begins to occur in the first 2 hours of interaction of the trypomastigote stage in peritoneal macrophages, already suggesting the occurrence of fusion of lysosomes, after entry of the parasites. More recently, Reignault and colleagues [50] demonstrate that galectin-3 decorates T. cruzi vacuole acting as an important marker to be used also for the study of parasitic vacuole lysis of T. cruzi, as it also undergoes a process of disorganization with consequent exit of the parasite into the cytoplasm of the host cell. Transcriptome studies have shown an extensive remodeling of the intracellular T. cruzi in the first 4 hours of trypomastigote invasion (until the parasitophorous vacuole disintegration), and these modifications can be associated with a regulation of the initial step of host cell invasion [110]. Some modifications occurring in intracellular differentiation processes (trypomastigotes to amastigotes and subsequently to trypomastigotes again) may be involved in the process of destruction of PV. One of the modifications that seems to be involved with this process is the increase in the enzymatic activity of glutamine synthase during the amastigogenesis process. This enzyme is normally involved in the process of handling excess ammonia, and given the fact that the main energy source of the intracellular forms of T. cruzi is from amino acids, the ammonium generation is high. Marim et al. [111] observed that the activity blockage of this enzyme impair the progression of the intracellular cycle (amastigotes did not differentiate for trypomastigotes). In this case the role of the enzyme is to regulate the intracellular pH by controlling the content of intravacuolar ammonium (generated by the consumption of amino acids in this reduced space). It is this acidification process followed by the pH control that allows the enzymatic activity that culminates in the release of parasite in the cytoplasmic environment allowing the infection to continue [112].
\nHost cell invasion and parasite internalization are important steps in the evolution of parasitism by several pathogens. These processes present at least two important advantages: protection against the host immune response and access to a microenvironment rich in metabolic products. Substantial progress has been made in understanding the roles of proteins in infection and invasion by T. cruzi. Host cell intracellular signaling can combat the infection; but it can also favor parasite entry. Parasites hijack the host immune response, phagocytosis, ECM, and antiparasitic proteins for their own survival, replication, and immune evasion purposes. The complex networks are interconnected and require extensive study to identify intracellular rearrangements that facilitate parasite internalization. A multidisciplinary approach is necessary to a better understanding of parasite-host interaction and will be critical to better understand Chagas disease physiopathology, diagnosis, and treatment.
\nWe thank all authors who work with this theme and for the contribution during the last years.
\nThe authors declare “no conflict of interest.”
\nResearch methodology is the path through which researchers need to conduct their research. It shows the path through which these researchers formulate their problem and objective and present their result from the data obtained during the study period. This research design and methodology chapter also shows how the research outcome at the end will be obtained in line with meeting the objective of the study. This chapter hence discusses the research methods that were used during the research process. It includes the research methodology of the study from the research strategy to the result dissemination. For emphasis, in this chapter, the author outlines the research strategy, research design, research methodology, the study area, data sources such as primary data sources and secondary data, population consideration and sample size determination such as questionnaires sample size determination and workplace site exposure measurement sample determination, data collection methods like primary data collection methods including workplace site observation data collection and data collection through desk review, data collection through questionnaires, data obtained from experts opinion, workplace site exposure measurement, data collection tools pretest, secondary data collection methods, methods of data analysis used such as quantitative data analysis and qualitative data analysis, data analysis software, the reliability and validity analysis of the quantitative data, reliability of data, reliability analysis, validity, data quality management, inclusion criteria, ethical consideration and dissemination of result and its utilization approaches. In order to satisfy the objectives of the study, a qualitative and quantitative research method is apprehended in general. The study used these mixed strategies because the data were obtained from all aspects of the data source during the study time. Therefore, the purpose of this methodology is to satisfy the research plan and target devised by the researcher.
The research design is intended to provide an appropriate framework for a study. A very significant decision in research design process is the choice to be made regarding research approach since it determines how relevant information for a study will be obtained; however, the research design process involves many interrelated decisions [1].
This study employed a mixed type of methods. The first part of the study consisted of a series of well-structured questionnaires (for management, employee’s representatives, and technician of industries) and semi-structured interviews with key stakeholders (government bodies, ministries, and industries) in participating organizations. The other design used is an interview of employees to know how they feel about safety and health of their workplace, and field observation at the selected industrial sites was undertaken.
Hence, this study employs a descriptive research design to agree on the effects of occupational safety and health management system on employee health, safety, and property damage for selected manufacturing industries. Saunders et al. [2] and Miller [3] say that descriptive research portrays an accurate profile of persons, events, or situations. This design offers to the researchers a profile of described relevant aspects of the phenomena of interest from an individual, organizational, and industry-oriented perspective. Therefore, this research design enabled the researchers to gather data from a wide range of respondents on the impact of safety and health on manufacturing industries in Ethiopia. And this helped in analyzing the response obtained on how it affects the manufacturing industries’ workplace safety and health. The research overall design and flow process are depicted in Figure 1.
Research methods and processes (author design).
To address the key research objectives, this research used both qualitative and quantitative methods and combination of primary and secondary sources. The qualitative data supports the quantitative data analysis and results. The result obtained is triangulated since the researcher utilized the qualitative and quantitative data types in the data analysis. The study area, data sources, and sampling techniques were discussed under this section.
According to Fraenkel and Warren [4] studies, population refers to the complete set of individuals (subjects or events) having common characteristics in which the researcher is interested. The population of the study was determined based on random sampling system. This data collection was conducted from March 07, 2015 to December 10, 2016, from selected manufacturing industries found in Addis Ababa city and around. The manufacturing companies were selected based on their employee number, established year, and the potential accidents prevailing and the manufacturing industry type even though all criterions were difficult to satisfy.
It was obtained from the original source of information. The primary data were more reliable and have more confidence level of decision-making with the trusted analysis having direct intact with occurrence of the events. The primary data sources are industries’ working environment (through observation, pictures, and photograph) and industry employees (management and bottom workers) (interview, questionnaires and discussions).
Desk review has been conducted to collect data from various secondary sources. This includes reports and project documents at each manufacturing sectors (more on medium and large level). Secondary data sources have been obtained from literatures regarding OSH, and the remaining data were from the companies’ manuals, reports, and some management documents which were included under the desk review. Reputable journals, books, different articles, periodicals, proceedings, magazines, newsletters, newspapers, websites, and other sources were considered on the manufacturing industrial sectors. The data also obtained from the existing working documents, manuals, procedures, reports, statistical data, policies, regulations, and standards were taken into account for the review.
In general, for this research study, the desk review has been completed to this end, and it had been polished and modified upon manuals and documents obtained from the selected companies.
The study population consisted of manufacturing industries’ employees in Addis Ababa city and around as there are more representative manufacturing industrial clusters found. To select representative manufacturing industrial sector population, the types of the industries expected were more potential to accidents based on random and purposive sampling considered. The population of data was from textile, leather, metal, chemicals, and food manufacturing industries. A total of 189 sample sizes of industries responded to the questionnaire survey from the priority areas of the government. Random sample sizes and disproportionate methods were used, and 80 from wood, metal, and iron works; 30 from food, beverage, and tobacco products; 50 from leather, textile, and garments; 20 from chemical and chemical products; and 9 from other remaining 9 clusters of manufacturing industries responded.
A simple random sampling and purposive sampling methods were used to select the representative manufacturing industries and respondents for the study. The simple random sampling ensures that each member of the population has an equal chance for the selection or the chance of getting a response which can be more than equal to the chance depending on the data analysis justification. Sample size determination procedure was used to get optimum and reasonable information. In this study, both probability (simple random sampling) and nonprobability (convenience, quota, purposive, and judgmental) sampling methods were used as the nature of the industries are varied. This is because of the characteristics of data sources which permitted the researchers to follow the multi-methods. This helps the analysis to triangulate the data obtained and increase the reliability of the research outcome and its decision. The companies’ establishment time and its engagement in operation, the number of employees and the proportion it has, the owner types (government and private), type of manufacturing industry/production, types of resource used at work, and the location it is found in the city and around were some of the criteria for the selections.
The determination of the sample size was adopted from Daniel [5] and Cochran [6] formula. The formula used was for unknown population size Eq. (1) and is given as
where n = sample size, Z = statistic for a level of confidence, P = expected prevalence or proportion (in proportion of one; if 50%, P = 0.5), and d = precision (in proportion of one; if 6%, d = 0.06). Z statistic (Z): for the level of confidence of 95%, which is conventional, Z value is 1.96. In this study, investigators present their results with 95% confidence intervals (CI).
The expected sample number was 267 at the marginal error of 6% for 95% confidence interval of manufacturing industries. However, the collected data indicated that only 189 populations were used for the analysis after rejecting some data having more missing values in the responses from the industries. Hence, the actual data collection resulted in 71% response rate. The 267 population were assumed to be satisfactory and representative for the data analysis.
The sample size for the experimental exposure measurements of physical work environment has been considered based on the physical data prepared for questionnaires and respondents. The response of positive were considered for exposure measurement factors to be considered for the physical environment health and disease causing such as noise intensity, light intensity, pressure/stress, vibration, temperature/coldness, or hotness and dust particles on 20 workplace sites. The selection method was using random sampling in line with purposive method. The measurement of the exposure factors was done in collaboration with Addis Ababa city Administration and Oromia Bureau of Labour and Social Affair (AACBOLSA). Some measuring instruments were obtained from the Addis Ababa city and Oromia Bureau of Labour and Social Affair.
Data collection methods were focused on the followings basic techniques. These included secondary and primary data collections focusing on both qualitative and quantitative data as defined in the previous section. The data collection mechanisms are devised and prepared with their proper procedures.
Primary data sources are qualitative and quantitative. The qualitative sources are field observation, interview, and informal discussions, while that of quantitative data sources are survey questionnaires and interview questions. The next sections elaborate how the data were obtained from the primary sources.
Observation is an important aspect of science. Observation is tightly connected to data collection, and there are different sources for this: documentation, archival records, interviews, direct observations, and participant observations. Observational research findings are considered strong in validity because the researcher is able to collect a depth of information about a particular behavior. In this dissertation, the researchers used observation method as one tool for collecting information and data before questionnaire design and after the start of research too. The researcher made more than 20 specific observations of manufacturing industries in the study areas. During the observations, it found a deeper understanding of the working environment and the different sections in the production system and OSH practices.
Interview is a loosely structured qualitative in-depth interview with people who are considered to be particularly knowledgeable about the topic of interest. The semi-structured interview is usually conducted in a face-to-face setting which permits the researcher to seek new insights, ask questions, and assess phenomena in different perspectives. It let the researcher to know the in-depth of the present working environment influential factors and consequences. It has provided opportunities for refining data collection efforts and examining specialized systems or processes. It was used when the researcher faces written records or published document limitation or wanted to triangulate the data obtained from other primary and secondary data sources.
This dissertation is also conducted with a qualitative approach and conducting interviews. The advantage of using interviews as a method is that it allows respondents to raise issues that the interviewer may not have expected. All interviews with employees, management, and technicians were conducted by the corresponding researcher, on a face-to-face basis at workplace. All interviews were recorded and transcribed.
The main tool for gaining primary information in practical research is questionnaires, due to the fact that the researcher can decide on the sample and the types of questions to be asked [2].
In this dissertation, each respondent is requested to reply to an identical list of questions mixed so that biasness was prevented. Initially the questionnaire design was coded and mixed up from specific topic based on uniform structures. Consequently, the questionnaire produced valuable data which was required to achieve the dissertation objectives.
The questionnaires developed were based on a five-item Likert scale. Responses were given to each statement using a five-point Likert-type scale, for which 1 = “strongly disagree” to 5 = “strongly agree.” The responses were summed up to produce a score for the measures.
The data was also obtained from the expert’s opinion related to the comparison of the knowledge, management, collaboration, and technology utilization including their sub-factors. The data obtained in this way was used for prioritization and decision-making of OSH, improving factor priority. The prioritization of the factors was using Saaty scales (1–9) and then converting to Fuzzy set values obtained from previous researches using triangular fuzzy set [7].
The researcher has measured the workplace environment for dust, vibration, heat, pressure, light, and noise to know how much is the level of each variable. The primary data sources planned and an actual coverage has been compared as shown in Table 1.
Planned versus actual coverage of the survey.
The response rate for the proposed data source was good, and the pilot test also proved the reliability of questionnaires. Interview/discussion resulted in 87% of responses among the respondents; the survey questionnaire response rate obtained was 71%, and the field observation response rate was 90% for the whole data analysis process. Hence, the data organization quality level has not been compromised.
This response rate is considered to be representative of studies of organizations. As the study agrees on the response rate to be 30%, it is considered acceptable [8]. Saunders et al. [2] argued that the questionnaire with a scale response of 20% response rate is acceptable. Low response rate should not discourage the researchers, because a great deal of published research work also achieves low response rate. Hence, the response rate of this study is acceptable and very good for the purpose of meeting the study objectives.
The pretest for questionnaires, interviews, and tools were conducted to validate that the tool content is valid or not in the sense of the respondents’ understanding. Hence, content validity (in which the questions are answered to the target without excluding important points), internal validity (in which the questions raised answer the outcomes of researchers’ target), and external validity (in which the result can generalize to all the population from the survey sample population) were reflected. It has been proved with this pilot test prior to the start of the basic data collections. Following feedback process, a few minor changes were made to the originally designed data collect tools. The pilot test made for the questionnaire test was on 10 sample sizes selected randomly from the target sectors and experts.
The secondary data refers to data that was collected by someone other than the user. This data source gives insights of the research area of the current state-of-the-art method. It also makes some sort of research gap that needs to be filled by the researcher. This secondary data sources could be internal and external data sources of information that may cover a wide range of areas.
Literature/desk review and industry documents and reports: To achieve the dissertation’s objectives, the researcher has conducted excessive document review and reports of the companies in both online and offline modes. From a methodological point of view, literature reviews can be comprehended as content analysis, where quantitative and qualitative aspects are mixed to assess structural (descriptive) as well as content criteria.
A literature search was conducted using the database sources like MEDLINE; Emerald; Taylor and Francis publications; EMBASE (medical literature); PsycINFO (psychological literature); Sociological Abstracts (sociological literature); accident prevention journals; US Statistics of Labor, European Safety and Health database; ABI Inform; Business Source Premier (business/management literature); EconLit (economic literature); Social Service Abstracts (social work and social service literature); and other related materials. The search strategy was focused on articles or reports that measure one or more of the dimensions within the research OSH model framework. This search strategy was based on a framework and measurement filter strategy developed by the Consensus-Based Standards for the Selection of Health Measurement Instruments (COSMIN) group. Based on screening, unrelated articles to the research model and objectives were excluded. Prior to screening, researcher (principal investigator) reviewed a sample of more than 2000 articles, websites, reports, and guidelines to determine whether they should be included for further review or reject. Discrepancies were thoroughly identified and resolved before the review of the main group of more than 300 articles commenced. After excluding the articles based on the title, keywords, and abstract, the remaining articles were reviewed in detail, and the information was extracted on the instrument that was used to assess the dimension of research interest. A complete list of items was then collated within each research targets or objectives and reviewed to identify any missing elements.
Data analysis method follows the procedures listed under the following sections. The data analysis part answered the basic questions raised in the problem statement. The detailed analysis of the developed and developing countries’ experiences on OSH regarding manufacturing industries was analyzed, discussed, compared and contrasted, and synthesized.
Quantitative data were obtained from primary and secondary data discussed above in this chapter. This data analysis was based on their data type using Excel, SPSS 20.0, Office Word format, and other tools. This data analysis focuses on numerical/quantitative data analysis.
Before analysis, data coding of responses and analysis were made. In order to analyze the data obtained easily, the data were coded to SPSS 20.0 software as the data obtained from questionnaires. This task involved identifying, classifying, and assigning a numeric or character symbol to data, which was done in only one way pre-coded [9, 10]. In this study, all of the responses were pre-coded. They were taken from the list of responses, a number of corresponding to a particular selection was given. This process was applied to every earlier question that needed this treatment. Upon completion, the data were then entered to a statistical analysis software package, SPSS version 20.0 on Windows 10 for the next steps.
Under the data analysis, exploration of data has been made with descriptive statistics and graphical analysis. The analysis included exploring the relationship between variables and comparing groups how they affect each other. This has been done using cross tabulation/chi square, correlation, and factor analysis and using nonparametric statistic.
Qualitative data analysis used for triangulation of the quantitative data analysis. The interview, observation, and report records were used to support the findings. The analysis has been incorporated with the quantitative discussion results in the data analysis parts.
The data were entered using SPSS 20.0 on Windows 10 and analyzed. The analysis supported with SPSS software much contributed to the finding. It had contributed to the data validation and correctness of the SPSS results. The software analyzed and compared the results of different variables used in the research questionnaires. Excel is also used to draw the pictures and calculate some analytical solutions.
The reliability of measurements specifies the amount to which it is without bias (error free) and hence ensures consistent measurement across time and across the various items in the instrument [8]. In reliability analysis, it has been checked for the stability and consistency of the data. In the case of reliability analysis, the researcher checked the accuracy and precision of the procedure of measurement. Reliability has numerous definitions and approaches, but in several environments, the concept comes to be consistent [8]. The measurement fulfills the requirements of reliability when it produces consistent results during data analysis procedure. The reliability is determined through Cranach’s alpha as shown in Table 2.
Internal consistency and reliability test of questionnaires items.
K stands for knowledge; M, management; T, technology; C, collaboration; P, policy, standards, and regulation; H, hazards and accident conditions; PPE, personal protective equipment.
Cronbach’s alpha is a measure of internal consistency, i.e., how closely related a set of items are as a group [11]. It is considered to be a measure of scale reliability. The reliability of internal consistency most of the time is measured based on the Cronbach’s alpha value. Reliability coefficient of 0.70 and above is considered “acceptable” in most research situations [12]. In this study, reliability analysis for internal consistency of Likert-scale measurement after deleting 13 items was found similar; the reliability coefficients were found for 76 items were 0.964 and for the individual groupings made shown in Table 2. It was also found internally consistent using the Cronbach’s alpha test. Table 2 shows the internal consistency of the seven major instruments in which their reliability falls in the acceptable range for this research.
Face validity used as defined by Babbie [13] is an indicator that makes it seem a reasonable measure of some variables, and it is the subjective judgment that the instrument measures what it intends to measure in terms of relevance [14]. Thus, the researcher ensured, in this study, when developing the instruments that uncertainties were eliminated by using appropriate words and concepts in order to enhance clarity and general suitability [14]. Furthermore, the researcher submitted the instruments to the research supervisor and the joint supervisor who are both occupational health experts, to ensure validity of the measuring instruments and determine whether the instruments could be considered valid on face value.
In this study, the researcher was guided by reviewed literature related to compliance with the occupational health and safety conditions and data collection methods before he could develop the measuring instruments. In addition, the pretest study that was conducted prior to the main study assisted the researcher to avoid uncertainties of the contents in the data collection measuring instruments. A thorough inspection of the measuring instruments by the statistician and the researcher’s supervisor and joint experts, to ensure that all concepts pertaining to the study were included, ensured that the instruments were enriched.
Insight has been given to the data collectors on how to approach companies, and many of the questionnaires were distributed through MSc students at Addis Ababa Institute of Technology (AAiT) and manufacturing industries’ experience experts. This made the data quality reliable as it has been continually discussed with them. Pretesting for questionnaire was done on 10 workers to assure the quality of the data and for improvement of data collection tools. Supervision during data collection was done to understand how the data collectors are handling the questionnaire, and each filled questionnaires was checked for its completeness, accuracy, clarity, and consistency on a daily basis either face-to-face or by phone/email. The data expected in poor quality were rejected out of the acting during the screening time. Among planned 267 questionnaires, 189 were responded back. Finally, it was analyzed by the principal investigator.
The data were collected from the company representative with the knowledge of OSH. Articles written in English and Amharic were included in this study. Database information obtained in relation to articles and those who have OSH area such as interventions method, method of accident identification, impact of occupational accidents, types of occupational injuries/disease, and impact of occupational accidents, and disease on productivity and costs of company and have used at least one form of feedback mechanism. No specific time period was chosen in order to access all available published papers. The questionnaire statements which are similar in the questionnaire have been rejected from the data analysis.
Ethical clearance was obtained from the School of Mechanical and Industrial Engineering, Institute of Technology, Addis Ababa University. Official letters were written from the School of Mechanical and Industrial Engineering to the respective manufacturing industries. The purpose of the study was explained to the study subjects. The study subjects were told that the information they provided was kept confidential and that their identities would not be revealed in association with the information they provided. Informed consent was secured from each participant. For bad working environment assessment findings, feedback will be given to all manufacturing industries involved in the study. There is a plan to give a copy of the result to the respective study manufacturing industries’ and ministries’ offices. The respondents’ privacy and their responses were not individually analyzed and included in the report.
The result of this study will be presented to the Addis Ababa University, AAiT, School of Mechanical and Industrial Engineering. It will also be communicated to the Ethiopian manufacturing industries, Ministry of Labor and Social Affair, Ministry of Industry, and Ministry of Health from where the data was collected. The result will also be availed by publication and online presentation in Google Scholars. To this end, about five articles were published and disseminated to the whole world.
The research methodology and design indicated overall process of the flow of the research for the given study. The data sources and data collection methods were used. The overall research strategies and framework are indicated in this research process from problem formulation to problem validation including all the parameters. It has laid some foundation and how research methodology is devised and framed for researchers. This means, it helps researchers to consider it as one of the samples and models for the research data collection and process from the beginning of the problem statement to the research finding. Especially, this research flow helps new researchers to the research environment and methodology in particular.
There is no “conflict of interest.”
General requirements for Open Access to Horizon 2020 research project outputs are found within Guidelines on Open Access to Scientific Publication and Research Data in Horizon 2020. The guidelines, in their simplest form, state that if you are a Horizon 2020 recipient, you must ensure open access to your scientific publications by enabling them to be downloaded, printed and read online. Additionally, said publications must be peer reviewed.
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