Campylobacter Coli - Research Proposal Example

?Literature Review The significance of Campylobacter coli in spreading microbial intestinal infections gains worldwide recognition as its properties and virulence are slowly being exposed. Luber, et al. 92003) considered Campylobacter spp. as one of the major causes of infectious intestinal illnesses such as bacterial diarrhoea and gastroenteritis in humans all over the world. A total of 2.4 million reported illnesses and 150 mortalities are caused by the genus Campylobacter every year in the United States (Mead, et al., 1999). Infections particularly from Campylobacter jejuni and C. coli amounts to 420,000 cases of intestinal disease in England alone every year (Tam et al., 2003), while it ranks second in the leading causes of bacterial food poisoning in the United States (Miller, et al., 2006). In addition, the English case-control Infectious Intestinal Disease Study revealed that among the cases in 1993 to 1996, Campylobacter spp. amounts to a percentage of 23% among all detected enteric pathogens from faecal samples (Amar, et al., 2007). There are 11 species of Campylobacter that are pathogenic to humans and animals, with C. jejuni and C. coli as the most frequently isolated species in food products (He, et al., 2010). Gurtler, et al. (2005) emphasized the importance of C. coli in human campylobacter infections as their research uncovered that it comprises about 18.6% of Campylobacter spp. found in most infection cases. According to the World Health Organization (2011), infections caused by Campylobacter spp. is mild but fatal especially to infants and elderly people. It is generally regarded as the foremost cause of gastroenteritis, diarrhoea and other intestinal diseases in developing and developed countries. The strong impact and high prevalence of Campylobacter foodborne illnesses placed the said bacteria to the list of pathogens of high significance in terms of health and socio-economic viewpoint, and the detection of its presence could aid in the control and prevention of future outbreaks. Several techniques are used in the detection of Campylobacter spp. as well as assessing the virulence of the strains. Well-known assays include stress assays using specific media which confer positive or negative results (Raphael, et al., 2005), virulence assays that detect virulent characteristics such as bacterial motility and adherence tests (Silva, et al., 2011), and PCR screening tests to identify the strains at the genotypic level as well as in detecting whether virulence genes are present in particular strains or not (Bang, et al., 2003). It is essential that such assays are done during the earliest stages to confirm the presence of the pathogen, and at the same time assess whether the isolated strains are capable of causing diseases in humans or not, especially since Campylobacter have wide host ranges and could easily contaminate various raw meat products in a short amount of time. The mode of action of Campylobacter involves the colonization of the intestines of live animals such as poultry, cattle, sheep, swine and other animals. Thereafter, the bacteria contaminate the meat produces during slaughtering and packaging of products. In turn, humans who consume the contaminated meat that are either raw or undercooked are can be infected with Campylobacter diseases (Kemp, et al. 2005). Aside from food and contaminated meat products, C. jejuni and C. coli are also present in different sources of water (Tam, et al., 2003). The pervasiveness of this microorganism in poultry, cattle and swine is disturbing, given that the growth and survival characteristics of Campylobacter spp. are capable to withstand extreme conditions. Silva, et al. (2011) noted that Campylobacter spp. are thermophiles and can only cease to grow below 30oC. It is thus important to note that frozen or fresh meat from poultry farms have a high susceptibility to become contaminated, but also free-range and organically-grown poultry as well. Although most cases of intestinal illnesses are self-limiting, advanced cases such as “systemic Campylobacter infections, Campylobacter infections in immunosuppressed patients, and severe or long-lasting Campylobacter infections” (Luber, et al., 2003) essentially need antimicrobial treatment like erythromycin or fluoroquinolones. Gebreyes et al. (2005, p. 766), maintain that C. coli has the “ability to show increased resistance to a greater number of antimicrobials”, and this resistance factor against known and effective antibiotics is an imperative concern since the medications that are commonly used to cure such diseases will be put to waste. It is concluded that the “discrepancies in the antimicrobial resistance rates among Campylobacter isolates originating from poultry and humans support the hypothesis that at least some of the resistant Campylobacter strains causing infection in humans come from sources other than poultry products.” (Luber, et al., 2003). In a study concerning the detection of Campylobacter spp., Vashin, et al. (2009) reported that high levels of bacteria were isolated from different parts of packed frozen poultry giblets in Bulgaria, where C. coli and C. jejuni populated 77.8% and 22.2% of the chicken liver, respectively. A similar research work conducted by Wieczorek, et al. (2012) on the prevalences and molecular characterization of Campylobacter jejuni and C. coli isolated from retail raw meat in Poland, revealed that all of the meat samples originating from poultry were contaminated with Campylobacter spp., with a total of 189 strains of C. coli amounting to 58.9% of the isolates. They concluded that the reason behind this is brought about by the lack of a blast chilling system in the plant, and the lag time before fully-freezing the meat promotes a considerable time for bacterial growth. Research aims The research generally aims to study the virulence of C. coli isolated from a range of poultry sources, including organic poultry meat. Specific aims This research paper aims to characterise the number of C. coli isolates and strains from organic chicken in terms of their virulence potential using the following tests: 1. Stress assays such as acidic tolerance assays, percentage bile salt assays, and aerobic stress assays; 2. Virulence assays such as motility detection assays, autoagglutination tests, and adhesion and cohesion assays; and 3. PCR screening to genotypically identify the C. coli strains and isolates, as well as to detect the presence of any virulent genes in the samples. Methodology The following procedures will be used to assess the virulence potential and in identifying present virulence genes in the sampled C. coli strains: 1. Stress assays This association assay will test for the acidic and percentage bile salt tolerance and aerobic stress of the strains. The acidic stress tolerance assay aims to establish the strains’ properties by finding and measuring the acidity limit that the bacteria could thrive in before undergoing the death phase (Requena, 2012). The bile salt percentage tolerance test aims to compare the maximum and minimum percentage salinity that the strain could tolerate under different bile salts as described (Raphael, et al., 2005). Aerobic stress level assays on the other hand will measure the tolerance limit of the strains by measuring cell viability after oxygen exposure (Wiedmann and Zhang, 2011). 2. Virulence assays Virulence assays will test for motility, autoagglutination of the bacterial cells, as well as adhesion and invasion properties of isolated C. coli strains. Motility tests detect the capacity of the strains of self-movement through the use of semisolid motility medium plates (Raphael, et al., 2005). Autoagglutination tests such as latex agglutination assays and cell adhesion and invasion tests are used to check the presence of bacterial colonies that will grow attached to eukaryote cells after undergoing several treatments (Konkel and Joens, 1989; Silva, et al., 2011; Wassenar and Newell, 2000). The ability for self-movement is to be compared among strains of C. coli since motility increases chances of a wider infection among foodborne pathogens, and the adhesion and autoagglutination methods are important since bacteria mostly attach via cell surface sugars interactions and are therefore affected by the class of sugar residues present on host cells (Cheesbrough, 2006; Sharon, et al., 2009; Silva, et al., 2011). 3. PCR screening PCR screening will be conducted to identify known virulence genes of Campylobacter spp. by the detection of certain virulence and toxin genes, which were already identified in previous studies (Bang, et al., 2003; Hong, et al., 2003). This method of strain detection helps in properly identifying the virulent strain through genotyping, which has higher detection accuracy in comparison with phenotypic levels of identification and thus is integral in confirmatory testing. Time Frame The study will undergo five phases of the research calendar, as shown in the following 2013 Gannt Chart: April May June July August 1 2 2 3 3 3 3 4 4 4 4 5 Legend: 1. Stress assays – April 2013 2. Motility tests –Middle of April to Middle of May 2013 3. Virulence assays – Late April to Middle of June 2013 4. PCR screening – Middle of April to July 2013 5. Thesis complete – August 2013 Read More