Review Article Creative Commons, CC-BY
Review on Some Virulence Factors Associated with Campylobacter Colonization and Infection in Poultry and Human
*Corresponding author: Wafaa MM Hassan, Reference laboratory for veterinary quality control on poultry production (RLQP), Animal health research institute, Dokki, Giza, Egypt.
Received: June 28, 2019; Published: July 08, 2019
Campylobacter is one of the most important four global diarrheal diseases. It is considered to be the most common bacterial cause of human gastroenteritis in the world causing a disease called Campylobacteriosis. In developing countries, Campylobacteriosis in children under the age of 2 years are especially frequent and sometimes resulting in death . Mainly C. jejuni and C. Coli are well recognized causes of human campylobacteriososis with symptoms ranging from mild watery diarrhea to serious neuropathies . Poultry (particularly chicken and contaminated raw chicken carcasses) is considered to be the main source for human campylobacteriososis. Other sources such as water, raw milk, Cattle, sheep, pigs, cats, dogs, vehicles, rodents and insects are known as possible sources for not only human but also poultry Campylobacteriosis. After being colonized by Campylobacter spp. Chicken in contrast to human, do scarcely develop pathological lesions . The high body temperature of Poultry species provides an optimal environment for the growth of thermophilic Campylobacter species particularly C. jejuni and C. coli which make poultry constitute the main source of human Campylobacteriosis 
Campylobacter spp. are Gram negative rods, 0.5 - 8μm long and 0.2 - 0.5μm wide with characteristically curved, spiral, or S-shaped cells; coccal forms may be seen under sub-optimal conditions. They generally have a single polar unsheathed flagellum at one or both ends. The motility of the bacteria is characteristically rapid and darting in corkscrew fashion, a feature by which their presence among other bacteria can be detected by phase-contrast microscopy [5,6].
On Skirrow or other blood-containing agars, characteristic Campylobacter colonies are slightly pink, round, convex, smooth and shiny, with a regular edge. On charcoal-based media such as mCCDA, the characteristic colonies are greyish, flat and moistened, with a tendency to spread, and may have a metallic sheen. Campylobacter spp. require microaerobic conditions consisting of (5% O2, 10% CO2 and 85% N2) . They neither ferment nor oxidase carbohydrates. Energy is obtained from amino acids or tricarboxylic acid cycle intermediates, not carbohydrates. Some species, particularly C. jejuni, C. coli and C. lari, are thermophilic, grow optimally at 42°C .
Despite over 30 years of research, Campylobacteriosis is the most prevalent bacterial cause of foodborne infection in many countries including in the EU and the USA . As mentioned before that poultry species are important reservoirs for the transmission of Campylobacter species and their high body temperature provides an optimal environment for the growth of the organism . It is important to explore further the relationships between certain Campylobacter virulence genes and their capacity for survival in poultry meat, and hence their contribution to the incidence of Campylobacteriosis  and the large genetic diversity of Campylobacter must be considered in epidemiological evaluations and microbial risk assessments of Campylobacter in poultry [11,12].
As a first step, colonization of the intestine requires the ability to move into the mucus layer covering the intestinal cells. Campylobacter motility is conferred by the polar flagella, which together with their ‘cork-screw’ shape allow them to efficiently penetrate this mucus barrier [13,14]. The most important virulence factor that has been studied and well characterized in Campylobacter spp. was the flagellin, which is encoded by the flaA gene . The global regulator, CsrA (Carbon starvation regulator) gene, has been well characterized in several bacterial genera and is known to regulate a number of independent pathways via a post transcriptional mechanism, but remains relatively uncharacterized in the genus Campylobacter .
Many of virulence genetic factors connected with Campylobacter invasiveness are placed on the pVir plasmid for example, virB11 gene that encodes the IV secretory system protein. It has been showed that strains with mutation in the virB11 sequence have much lower adhesion and penetration ability in vitro in comparison to original strains, as well as lower pathogenicity 0069n vivo the plasmid gene virB11 . One of the most important genes responsible for Campylobacter invasion is the CiaB (Campylobacter invasive antigen B) gene which is known to be involved in the translocation of Campylobacter into host cells for the purpose of host cell invasion and also plays a significant role in cecal colonization in chicken . The invasion-associated marker (iam) gene is one of most important factors responsible for Campylobacter invasion of host cell and this gene was first reported  and was detected in 85% of invasive strains and 20% of non-invasive strains. The pldA gene is also related to cell invasion and is responsible for the synthesis of an outer membrane phospholipase that is important for cecal colonization [18,20]. That gene encodes proteins associated with increased bacterial invasion on cultured epithelial cells . Cytolethal distending toxin (CDT) in which CdtB subunit is the active toxic unit. CdtA and CdtC required for CDT binding to target cells and for the delivery of CdtB into the cell interior . The toxin is retrograde transported into the nuclear compartment, where the CdtB subunit exhibits type I DNase activity. Cellular intoxication induces DNA damage and activation of the DNA damage response, which results in arrest of the target cells in the G1 and/or G2 phases of the cell cycle and activation of DNA repair mechanisms, cellular distention and nuclear enlargement, and Cdc2 and ataxia-telangiectasia- mutated protein (ATM) phosphorylation. Cells that fail to repair the damage will senesce or undergo apoptosis . Considering the important role that toxins have in the pathogenesis of Campylobacteriosis and other infections, all knowledge generated in this area will serve to propose and develop new strategies for the control of pathogens . The thermal stress response of bacteria is mostly carried out by the induction of the expression of heat shock proteins (HSPs). These HSPs have an important function in thermotolerance as well as in the response to other stresses by acting as chaperones to promote the folding of most cellular proteins and proteolysis of potentially deleterious, misfolded proteins. Several HSPs have been identified in C. jejuni, including the GroESL, DnaJ, DnaK and ClpB proteins [25-28]. However, a role in C. jejuni pathogenesis has only been demonstrated for the DnaJ protein, as a C. jejuni dnaJ mutant was unable to colonize chickens . The importance of the C. jejuni thermal stress response is also indicated by the link between thermoregulation and chicken colonization through the racR regulatory protein  Cited by . dnaJ was detected in 100% of all chicken fecal samples examined while there is a difference in human samples with detection rate of 98% . Relatively similar results were obtained in Egypt by  who confirmed these results with gene expression of dnaJ and using 23srRNA as a housekeeping gene. On the other hand our results in human samples agreed with  who detected dnaJ gene in 46% and 50% of human C. jejuni and C. coli samples respectively although there are some differences in results of dnaJ from chicken samples which came in a rate of 69% and 70% for C. jejuni and C. coli respectively , detected dnaJ gene in 100% of Human C. jejuni and C. coli. High detection of dnaJ gene in chicken than human host that reported by many authors confirmed the data that revealed importance of dnaJ gene in broiler cecal colonization by . Two-component regulatory system, RacR-RacS (reduced ability to colonize) system, that is involved in a temperature-dependent signalling pathway was identified . A mutation of the response regulator gene racR reduced the organism’s ability to colonize the chicken intestinal tract and resulted in temperature-dependent changes in its protein profile and growth characteristics. Authors added that C. jejuni dnaJ gene is adjacent to and under the transcriptional control of racR.  detected racR gene in 98.2 and 100% of C. jejuni isolates from human and broiler respectively , reported partially similar results in C. jejuni but not C. coli. They detected racR gene in 84.9% and 95.6% of C. jejuni from human and poultry respectively , detected racR gene in 100% of C. jejuni isolated from Human diarrheal patients in Bangladesh , reported racR gene in 98.3 % of C. jejuni isolates from children’s ≤14 years who were treated for diarrhoea at emergency rooms in north-eastern Brazil , detected racR gene in 95% and 0% of human C. jejuni and C. coli respectively. And in 76% and 79% of chicken C. jejuni and C. coli respectively. Similar results revealed by  in Egypt.
Infection with C. jejuni usually causes uncomplicated gastroenteritis; however, in rare cases can lead to the Guillain-Barré syndrome (GBS), a post infectious immune-mediated disorder of the peripheral nerves and nerve roots . The global incidence of GBS ranges from 0.4 to 4.0 (median 1.3) cases per 100,000 people annually, occurring slightly more often in adolescents and young adults . Results of  review analysis suggest that 31% of 2,502 GBS cases included in this review are attributable to Campylobacter infection. Molecular mimicry between lip oligosaccharides (LOS) present on the cell wall of C. jejuni and gangliosides found in the human nervous system is thought to play a critical role in the pathogenesis of C. jejuni-related GBS . The wlaN, cgtB and waaC are LOS (lipo-oligosaccharides) associated genes while wlaN and cgtB are involved in β-1,3 galactosyltransferase production. These two genes are associated with waaC gene which encodes heptosyltransferase I . The waaC gene, which encodes heptosyltransferase I, is responsible for transferring the first l-glycerod- manno- heptose residue to the inner core of LOS . The wlaN gene, which encodes a beta-1,3 galactosyltransferase, is responsible for biosynthesis GM1-like structure whereas cgtB (which encodes another beta-1,3 galactosyltransferase) catalyzes the biosynthesis of the carbohydrate moieties analogous to GM2 . Sialyltransferase encoded by the cst-II gene in C. jejuni is associated with risk of developing GBS . On the other hand, the cst-II gene has been linked to the invasiveness of C. jejuni for intestinal epithelial cells . C. jejuni gene ggt encoding the periplasmic gamma-glutamyl transpeptidase (GGT) seems to play a pivotal role in the enteric colonization. GGT has been shown in chicken model to be important in long lasting gut colonization, and in vitro it has been shown that GGT plays a significant role in C. jejuni-mediated apoptosis [48,49] detected cst-II and ggt genes in 83.6% and 32.7% of 55 examined Campylobacter jejuni human origin isolates and 40% and 5.5% of 55 Campylobacter jejuni broiler meat origin isolates in Chile , detected cgtB, wlaN and waaC genes in 7.69%, 30.77% and 57.69% of isolates respectively in Bangladesh , detected wlaN and cgtB in 20% and 6.7% of 30 C. jejuni isolates from Patients with Diarrhea in Rosario, Argentina.
Campylobacter epidemiology results should be liked with its virulence gene characterization. Although the molecular basis of pathogenicity of Campylobacter has not been fully elucidated, several virulence factors have been identified based on in vitro and in vivo studies. For example, flaA, cadF, CsrA for adhesion. iam, virB11, ciaB and pldA (invasion). CDT (CdtA, CdtB and CdtC) (cytotoxicity). dnaJ (heat shock protein). racR (reduced ability to colonize). cgtB, waaC, cstII, wlaN & ggt (ganglioside mimicry)..
- WHO (2018) Campylobacter/ World Health Organization.
- Khoshbakht R, Tabatabaei M, Shirzad Aski H, Hosseinzadeh S (2014) Occurrence of virulence genes and strain diversity of thermophilic campylobacters isolated from cattle and sheep faecal samples. Iranian Journal of Veterinary Research 15(2): 138-144.
- Pielsticker C, Glunder G, Rautenschlein S (2012) Colonization Properties of Campylobacter jejuni in Chickens. Eur J Microbiol Immunol 2(1): 61- 65.
- Noormohamed A, Fakhr MK (2014) Prevalence and antimicrobial susceptibility of Campylobacter spp. in oklahoma conventional and organic retail poultry. Open Microbiol J 8: 130-137.
- Vandamme P, Debruyne L, De Brandt E, Falsen E (2010) Reclassification of Bacteroides ureolyticus as Campylobacter ureolyticus comb. nov., and emended description of the genus Campylobacter. Int J Syst Evol Microbiol 60(Pt 9): 2016-2022.
- WJ Snelling, M Matsuda, JE Moore, JSG Dooley (2005) Campylobacter jejuni. Letters in Applied Microbiology 41(4): 297-302.
- OIE (2008) OIE Terrestrial Manual Chapter 2. 9. 3; Campylobacter jejuni and Campylobacter coli. pp. 1185-1191.
- Garrity GM, Bell J A, Lilburn T (2005) Family II. Helicobacteraceae fam. nov. In: Brenner DJ, et al. (Eds) Bergey’s Manual of Systematic Bacteriology, (The Proteobacteria), Part C (The Alpha-, Beta-, Delta-, and Epsilon proteobacteria). New York, USA. pp. 1168.
- Bolton DJ (2015) Campylobacter virulence and survival factors. Food Microbiol 48: 99-108.
- Abu Madi M, Behnke JM, Sharma A, Bearden R, Al Banna N (2016) Prevalence of Virulence/Stress Genes in Campylobacter jejuni from Chicken Meat Sold in Qatari Retail Outlets. PLoS One 11(6): e0156938.
- Alter T, Weber RM, Hamedy A, Glünder G (2011) Carry-over of thermophilic Campylobacter spp. between sequential and adjacent poultry flocks. Vet Microbiol 147(1-2): 90-95.
- Vidal AB, Colles FM, Rodgers JD, McCarthy ND, Davies RH, et al. (2016) Genetic Diversity of Campylobacter jejuni and Campylobacter coli Isolates from Conventional Broiler Flocks and the Impacts of Sampling Strategy and Laboratory Method. Appl Environ Microbiol 82(8): 2347- 2355.
- Szymanski CM, King M, Haardt M, Armstrong GD (1995) Campylobacter jejuni motility and invasion of Caco-2 cells. Infect Immun 63(11): 4295- 4300.
- Haag LM, Fischer A, Otto B, Grundmann U, Kühl AA, et al. (2012) Campylobacter jejuni infection of infant mice: acute enterocolitis is followed by asymptomatic intestinal and extra-intestinal immune responses. Eur J Microbiol Immunol (Bp) 2(1): 2-11.
- Hermans D, Van Deun K, Martel A, Van Immerseel F, Messens W, et al. (2011) Colonization factors of Campylobacter jejuni in the chicken gut. Vet Res 42: 82.
- Fields JA, Thompson SA (2012) Campylobacter jejuni CsrA complements an Escherichia coli csrAmutation for the regulation of biofilm formation, motility and cellular morphology but not glycogen accumulation. BMC Microbiol 12: 233.
- Bacon DJ, Alm RA, Burr DH, Hu L, Kopecko DJ, et al. (2000) Involvement of a plasmid in virulence of Campylobacter jejuni 81-176. Infect Immun 68(8): 4384-4390.
- O Cróinín T, Backert S (2012) Host epithelial cell invasion by Campylobacter jejuni: Trigger or zipper mechanism? Front Cell Infect Microbiol 2: 25.
- Carvalho AC, Ruiz Palacios GM, Ramos Cervantes P, Cervantes LE, Jiang X , et al. (2001) Molecular characterization of invasive and noninvasive Campylobacter jejuni and Campylobacter coli isolates. J Clin Microbiol 39(4): 1353-1359.
- Reddy S, Zishiri OT (2018) Genetic characterization of virulence genes associated with adherence, invasion and cytotoxicity in Campylobacter spp. isolated from commercial chickens and human clinical cases. Onderstepoort J Vet Res 85(1): e1-e9.
- Ghorbanalizadgan M, Bakhshi B, Kazemnejad Lili A, Najar Peerayeh S, Nikmanesh B (2014) A molecular survey of Campylobacter jejuni and Campylobacter coli virulence and diversity. Iran Biomed J 18(3): 158- 164.
- Lara Tejero M, Galán JE (2002) Cytolethal distending toxin: limited damage as a strategy to modulate cellular functions. Trends Microbiol 10(3): 147-152.
- Guerra L, Guidi R, Frisan T (2011) Do bacterial genotoxins contribute to chronic inflammation, genomic instability and tumor progression, FEBS J 278(23): 4577-4588.
- Méndez Olvera ET, Bustos Martínez JA, López Vidal Y, Verdugo Rodríguez A, Martínez Gómez D (2016) Cytolethal Distending Toxin From Campylobacter jejuni Requires the Cytoskeleton for Toxic Activity. Jundishapur J Microbiol 9(10): e3559.
- Konkel ME, Kim BJ, Klena JD, Young CR, Ziprin R (1998) Characterization of the thermal stress response of Campylobacter jejuni. Infect Immun 66(8): 3666-3672.
- Thies FL, Karch H, Hartung HP, Giegerich G (1999) The ClpB protein from Campylobacter jejuni: molecular characterization of the encoding gene and antigenicity of the recombinant protein. Gene 230(1): 61-67.
- Thies FL, Karch H, Hartung HP, Giegerich G (1999) Cloning and expression of the dnaK gene of Campylobacter jejuni and antigenicity of heat shock protein 70. Infect Immun 67(3): 1194-1200.
- Thies FL, Weishaupt A, Karch H, Hartung HP, Giegerich G (1999) Cloning, sequencing and molecular analysis of the Campylobacter jejuni groESL bicistronic operon. Microbiology 145(Pt 1): 89-98.
- Brás AM, Chatterjee S, Wren BW, Newell DG, Ketley JM (1999) A novel Campylobacter jejuni two‐component regulatory system important for temperature‐dependent growth and colonization. J Bacteriol 181(10): 3298-3302.
- AHM Van Vliet, JM Ketley (2001) Pathogenesis of enteric Campylobacter infection. Journal of Applied Microbiology 90(S6): 45S-56S.
- Datta S, Niwa H, Itoh K (2003) Prevalence of 11 pathogenic genes of Campylobacter jejuni by PCR in strains isolated from humans, poultry meat and broiler and bovine faeces. J Med Microbiol 52(Pt 4): 345-348.
- Mekky AAA (2019) Molecular Characterization of main virulence factors responsible for campylobacter colonization in poultry and Human.
- Cho HH, Kim SH, Min W, Ku BK, Kim YH (2014) Prevalence of virulence and cytolethal distending toxin (CDT) genes in thermophilic Campylobacter spp. from dogs and humans in Gyeongnam and Busan, Korea. Korean Journal of Veterinary Research 54(1): 39-48.
- Ziprin RL, Young CR, Stanker LH, Hume ME, Konkel ME (1999) The absence of cecal colonization of chicks by a mutant of Campylobacter jejuni not expressing bacterial fibronectin-binding protein. Avian Dis 43(3): 586-589.
- Bardoň J, Pudová V, Koláčková I, Karpíšková R, Röderová M, et al. (2017) Virulence and antibiotic resistance genes in Campylobacter spp. in the Czech Republic. Epidemiol Mikrobiol Imunol 66(2): 59–66.
- Talukder KA, Aslam M, Islam Z, Azmi IJ, Dutta DK, et al. (2008) Prevalence of Virulence Genes and Cytolethal Distending Toxin Production in Campylobacter jejuni Isolates from Diarrheal Patients in Bangladesh. J Clin Microbiol 46(4): 1485-1488.
- Quetz Jda S, Lima IF, Havt A, Prata MM, Cavalcante PA, et al. (2012) Campylobacter jejuni infection and virulence-associated genes in children with moderate to severe diarrhoea admitted to emergency rooms in northeastern Brazil. J Med Microbiol 61(Pt 4): 507-513.
- Thakur S, Zhao S, McDermott PF, Harbottle H, Abbott J, et al. (2010) Antimicrobial resistance, virulence, and genotypic profile comparison of Campylobacter jejuni and Campylobacter coli isolated from humans and retail meats. Foodborne Pathog Dis 7(7): 835-844.
- Bax M, Kuijf ML, Heikema AP, van Rijs W, Bruijns SC, et al. (2011) Campylobacter jejuni Lipooligosaccharides Modulate Dendritic Cell- Mediated T Cell Polarization in a Sialic Acid Linkage-Dependent Manner. Infect Immun 79(7): 2681-2689.
- Hadden RD, Gregson NA (2001) Guillain-Barré syndrome and Campylobacter jejuni infection. Symp Ser Soc Appl Microbiol 30(Suppl): S145-54.
- Poropatich KO, Walker CL, Black RE (2010) Quantifying the association between Campylobacter infection and Guillain-Barré syndrome: a systematic review. J Health Popul Nutr 28(6): 545-552
- Ang CW, Jacobs BC, Laman JD (2004) The Guillain-Barré syndrome: a true case of molecular mimicry. Trends Immunol 25 (2): 61-66.
- Müller J, Schulze F, Müller W, Hänel I (2006) PCR detection of virulence associated genes in Campylobacter jejuni strains with differential ability to invade Caco-2 cells and to colonize the chick gut. Vet Microbiol 113(1- 2): 123-129.
- Klena JD, Gray SA, Konkel ME (1998) Cloning, sequencing, and characterization of the lipopolysaccharide biosynthetic enzyme heptosyltransferase I gene (waaC) from Campylobacter jejuni and Campylobacter coli. Gene 222(2): 177-185.
- Linton D, Gilbert M, Hitchen PG, Dell A, Morris HR, et al. (2000) Phase variation of a beta-1, 3 galactosyltransferase involved in generation of the ganglioside GM1-like lipo-oligosaccharide of Campylobacter jejuni. Mol Microbiol 37(3): 501-514.
- van Belkum A, van den Braak N, Godschalk P, Ang W, Jacobs B, et al. (2001) A Campylobacter jejuni gene associated with immune-mediated neuropathy. Nat Med 7(7): 752–753.
- Louwen R, Heikema A, van Belkum A, Ott A, Gilbert M, et al. (2008) The sialylated lipooligosaccharide outer core in Campylobacter jejuni is an important determinant for epithelial cell invasion. Infect Immun 76(10): 4431-4438.
- Barnes IH, Bagnall MC, Browning DD, Thompson SA, Manning G, et al. (2007) Gamma-glutamyl transpeptidase has a role in the persistent colonization of the avian gut by Campylobacter jejuni. Microb Pathog 43(5-6): 198-207.
- González Hein G, Huaracán B, García P, Figueroa G (2013) Prevalence of virulence genes in strains of Campylobacter jejuni isolated from human, bovine and broiler. Braz J Microbi 44(4): 1223–1229
- Nahar N, Rashid RB (2018) Genotypic Analysis of the Virulence and Antibiotic Resistance Genes in Campylobacter species in silico. Journal of Bio analysis and Biomedicine 10(1): 13-23.
- Casabonne C, Gonzalez A, Aquili V, Subils T, Balague C (2016) Prevalence of Seven Virulence Genes of Campylobacter jejuni Isolated from Patients with Diarrhea in Rosario, Argentina. International Journal of Infection 3(4): e37727.