The determination of antibiotic residues in breast muscle, liver and gizzard of broilers collected from markets of Nakawa Division. The study used questionnaire as a data collection tool for the antibiotics used by the farmers. Sample extraction was performed where the 4g of each minced sample, 10 mL Phosphate Buffer Saline (pH-6.5) was added and mixed by vortexing using a Centrifuge @ 6000 rpm for 20 min after mixing with 2 mL 30% TCA. Supernatant was collected and filtered by Watman filter paper of 50 µm in thickness and funnel.

An optimized triple-quadrupole mass spectrometer/DA method 6400 ESI was used for identification and quantification of the five antibiotics. The results from the questionnaires showed that 82.2% of respondents use Tetracycline at the poultry farm, 68.8% use Penicillin, 62.2% use Chloramphenicol, 57.8% use Enrofloxacin while only 35.6% use Oxytetracycline. Also 32.67% of the analysed samples had the presence of Oxytetracycline (OTC), Tetracycline, Enrofloxacin, Penicillin G and Chloramphenicol and at least one antibiotic residue was found in Banda and Nakawa with the highest concentration of antibiotics, Ntinda and Luzira had the lowest. Of the positive samples 43.49%, 28.07% and 28.45% were found in liver, breast muscles and gizzard of broilers respectively since the p-value (0.000 < 0.05) showing a significant influence on the presence of antibiotic residues found in liver, breast muscles and gizzard. It was concluded that all poultry sold in different markets, there is presence of antibiotics which are above the maximum residual limits.

CHAPTER ONE: INTRODUCTION

1.1: Background of Study

Poultry are domesticated avian species that can be raised for eggs, meat and/or feathers therefore, covers a wide range of birds, from indigenous and commercial breeds of chickens to Muscovy ducks, mallard ducks, turkeys, guinea fowl, geese, quail, pigeons, ostriches and pheasants (Gerber et al., 2007). However, Chicken is the most common type of poultry in the world and in 2019, chickens accounted for some 93 percent of the world’s poultry population, followed by ducks (4 percent), and turkeys (2 percent) (FAO, 2020). Chickens contribute 90 percent of world poultry meat production, followed by turkeys with 5 percent, ducks with 4 percent and geese and guinea fowl with 2 percent (FAO, 2020). The rest comes from other poultry species. Chickens are classified according to the breeds from broilers, local, to layer etc. Layers are bred for eggs while broilers are chicken bred for meat hence rapidly growth compared to layers. Broiler strains are based on hybrid crosses between Cornish White, New Hampshire and White Plymouth Rock. In broiler production there are two main production phases – keeping of parent stock and production of day-old-chicken (DOC); and growing and finishing of broilers (Gerber et al., 2007). Chickens provide 93 percent of world egg production. At global level, non-chicken poultry species lay 10 percent of eggs in Asia, 1 percent in the Americas (3 percent in Latin America), 0.6 percent in Oceania and 0.5 percent in Europe (Prabakaran, 2003).

Chicken production contribute to improve human nutrition and food security by being a leading source of high-quality protein in form of eggs and meat. It acts as a key supplement to revenue from crops and other livestock enterprises, thus avoiding over dependency on traditional commodities with inconsistent prices (Kondaiah, 2007).

Chicken meat is the most common source of white meat which is richly nutritious (Ahmed & Gareib, 2016). It generally has a low-fat content which is found under the skin. Chicken meat is a delicacy and is popular in many regions and communities all over the world (Mcintyre & Choonara, 2004). The main edible parts are; Breast; which is mostly white meat, Leg; comprises two segments; the drum stick, this is dark meat and is the lower part of the leg, the thigh; this is the upper part of the leg. Wing; comprises three segments; the drumette, shaped like a small drum stick, the middle flat segment containing two bones and the tip sometimes discarded. Others; giblets; containing organs such as the heart, gizzards and liver, the head and the feet (Mcintyre & Choonara, 2004).

In addition, chicken has a high potential to generate foreign exchange earnings through export of poultry products to neighboring countries (Donoghue, 2003). Poultry is highly prized in many social-cultural functions such as dowry and festivities. The poultry industry in Uganda is relatively new. Its major problem is therefore associated with its further expansion, though care must be taken to avoid overproduction (MAAIF, 2019). The industry is characterized by widely diverse methods of production which include the following: village flocks, small scale commercial flocks and large-scale commercial farms (Gerber et al., 2007). Constraints in poultry production include: production related constraints (inadequate access to improved breed, access and affordability of feed, disease control); lack of knowledge and skills, inadequate capital at all levels and marketing. (Doyle, 2006).

Chicken production is conveniently categorized into two; namely; commercial and free-range (Gerber et al., 2007). There are wide variations within these two systems in terms of numbers, types of birds, bio-security and management. The commercial system covers production with improved hybrid breeds (and recently also with local breeds) under intensive confined management of uniform stocks and age-groups primarily for commercial purposes. Categorization may also be based on management systems i.e., intensive, semi-intensive and extensive or free ranging system (Gerber et al., 2007). The intensive system, which is based on specialized breeds, constitutes less than 20% of the total poultry population in Uganda (MAAIF, 2019). This system is found mainly in urban areas, where there are markets for eggs and chicken meat. Producers in this production system aim at using the recommended standard practices, such as breed of choice depending on production objectives, appropriate housing, feeding and health and disease control programs (Kondaiah, 2007). Intensive poultry farming was introduced in Uganda in the early 1950s, with the establishment of two government farms at Entebbe and Mbarara whereby under intensive farming methods, a broiler chicken lives less than six weeks before slaughter. Free-range chickens are usually slaughtered at 8 weeks and organic at around 12 weeks (Kondaiah, 2007).

The industry registered rapid development during the 60s, and the country had started exporting poultry products. However, it experienced a sharp decline during the late 70s and early 80s due to civil strife and political turmoil. Although the industry has not yet fully recovered, some progress has been achieved during the last ten years (UBOS, 2008).

Poultry production in Uganda rose by 3.2 per cent in 2012 despite the effect climate change has had on the country’s poultry sector. The increase in production levels has led to people in the country venturing into poultry farming, with the hope of creating a revenue stream. It was also revealed that the number of cattle in the country has been on the rise from 12.1mn in 2010 to 12.46mn in 2011 before rising to 12.84mn last year (UBOS, 2013).

Poultry production has increased from 42.7 million birds in 2010 to 44.3 million in 2011 and 45.9 million last year. This rise is expected to save the country millions of dollars, which otherwise would have been used on the importation of chickens and eggs, mainly from Brazil and South Africa. The increased poultry production is due to the prevention of animal diseases and improvement in livestock production systems because of routine livestock extension interventions. Egg production increased to 27,057 tons (807,634 eggs) compared to 26,269 tons (784,111 eggs) reported back in 2011. Ugachick Poultry Breeders Limited revamps poultry industry in Uganda through production of day-old chicks, high quality animal feeds and other poultry products. (UBOS, 2014)

Only one farm (Ugachick) has the very high bio-security measures that qualify it to be classified as class 1 chicken according to FAO 2020 Ugachick is located near the capital city of Kampala and has major outlets in other main towns such as Jinja and Mbarara, where they also have “out grower’ farmers (UBOS, 2014).

1.2: Problem Statement

The poultry industry in Uganda has experienced commendable growth in recent years, driven primarily by the dominance of chicken in egg and meat production. Despite this positive trajectory, the sector grapples with multifaceted challenges hindering its sustainable expansion and development. The industry’s diverse production methods, ranging from village flocks to large-scale commercial farms, create issues related to standardization, bio-security, and overall management, thereby impeding efficiency and productivity. Various constraints, including limited access to improved breeds, affordable feed, and effective disease control measures, hamper poultry production. Insufficient knowledge, skills, and capital further obstruct the industry’s growth potential, while a classification into commercial and free-range systems exists, the adoption of intensive poultry farming, crucial for urban market demands, remains limited, constituting less than 20% of the total poultry population. Challenges related to breed selection, housing, feeding, and health programs hinder the expansion of intensive poultry farming. Historical setbacks during the late 70s and early 80s due to civil strife and political turmoil continue to impact the poultry industry’s potential for sustained growth and global competitiveness. With only one farm recognized for high bio-security measures, the lack of widespread adoption of stringent bio-security measures poses a significant threat to the industry, especially concerning potential disease outbreaks. Despite increased production, Uganda relies on imports, affecting economic resilience and food security. Striking a balance between supply and demand and avoiding overproduction through strategic planning is crucial.

Furthermore, the potential presence of antibiotic residues in locally sold chicken meat raises serious concerns. Farmers’ failure to observe withdrawal periods before selling poultry birds previously treated with antibiotics exposes consumers to hazardous residues. Currently, there is no established analysis program to investigate antibiotic residues in the chicken meat available in local markets, emphasizing the need for urgent measures to safeguard consumers against the consequences of consuming such residues. A comprehensive approach involving stakeholders, policymakers, and industry players is essential to promote sustainable practices, enhance bio-security measures, provide education and training, and foster an environment conducive to the poultry industry’s growth in Uganda.

1.3: Objectives of the Study

1.3.1 General Objective of the Study

To investigate the presence of antibiotic residues in poultry breast meat, gizzard of broilers and liver collected from selected local markets in Nakawa Division, Kampala Capital City Authority (KCCA).

1.3.2 Specific Objectives of the Study

The specific objectives of this study were;

To establish the types of antibiotics used by poultry farmers in Nakawa Division (KCCA).
To identify and quantify the antibiotic residue in breast muscle, liver and gizzard of broilers collected from selected local markets in Nakawa Division.

1.4: Significance of the Study

The ever-increasing population around Nakawa division, especially the influx of students of Kyambogo University and Makerere University Business School community in this part of Kampala demands a lot of food to support this population. Therefore, the consumption of fast and cheap foods is always on the rise, chicken meat being one of the most available is highly consumed by the inhabitants of Banda, Mbuya, Nakawa market and many towns and commercial centres all over the country. However, with this high demand for chicken, the poultry farmers must use modern methods extensively to maintain production to be able to sustain supply. Hence, the use of antibiotics in production is so unavoidable that there are concerns of chemical residues in the edible chicken products. These chemical residues are likely to cause health risks to the consumers who are mainly students in this area.

These research findings will create awareness of antibiotic residues in chicken meat. This in turn will help to bring out the necessity to establish a regulatory measure to ensure safety of chicken meat consumers around Nakawa Division and other urban markets nationwide by the relevant authorities.

1.5: Scope of the Study

1.5.1: Content Scope

This research study was limited to investigation, identification and quantification of antibiotics used in poultry from selected markets Nakawa Division, Kampala Capital City Authority. This research investigated the presence of Tetracycline, Enrofloxacin, penicillin and chloramphenicol antibiotic residues in randomly collected chicken samples from selected markets in Nakawa Division.

1.5.2: Time Scope

The study was carried out in a period of one month from April to May 2019

1.5.3: Geographical Scope.

The study was carried out in Nakawa Division, Kampala Capital City in the central part of Uganda, Nakawa Division lies in the eastern part of Kampala city, bordering Kira Town to the east, Wakiso District to the north, Kawempe Division to the north-west, Kampala Central Division to the west, Makindye Division across Murchison Bay to the south-west and Lake Victoria to the south.

CHAPTER TWO: LITERATURE REVIEW

2.1: Poultry Production

Poultry production is one of the most important sectors in the agricultural setup of Uganda because it contributes to improve human nutrition and food security by being a leading source of high-quality protein in the form of eggs and meat (FAO, 2020). It acts as a key supplement to revenue from crops and other livestock enterprises, thus avoiding over-dependency on traditional commodities with inconsistent prices. It has a high potential to generate foreign exchange earnings through the export of poultry products to neighbouring countries (MAAIF, 2019).

Poultry is also highly prized in many social-cultural functions such as dowry and festivities. The poultry industry in Uganda is relatively new but the increasing investments in the livestock sector during the last two decades have led to a dramatic increase in the number of exotic chickens such as layers and broilers. However, the major problem is associated with further expansion of the poultry production due to the increasing market base which has led to usage of antibiotics for growth promotion and fighting diseases to meet with the demand and supply locally and internationally (Clarke. B, 2004; Gerber et al., 2007).

2.2: Antibiotics

2.2.1: Definition of Antibiotics

Antibiotics are chemical compounds that kill or inhibit the growth of microorganisms but cause little or no damage to the host,(Guardabassi, 2008).They are naturally produced by microorganisms such as fungi (e.g., penicillin) and bacteria (e.g., tetracycline) or can be semi-synthetically produced (e.g., amoxicillin) or totally synthetically produced (e.g., sulfonamides), (Guardabassi, 2008).

The term of antibiotic refers to a wide range of chemical substances derives from a natural, semisynthetic or synthetic way that affect antibacterial activity, by killing or inhibiting the growth of bacterial pathogens. (Mara C, 2001). Antibiotics have been used in poultry farming in large quantities since the 1940s. Since then, the antibiotics have been used on poultry in large quantities to enhance production in poultry. A wide range of antibiotics are used in poultry not only to treat diseases but also to maintain health, promote growth and enhance feed efficiency (Mara C, 2001).

They are widely used in the prevention and treatment of infectious diseases. Nowadays Antibiotics are widely prescribed for both therapeutic and prophylactic aims against microbial infections and as a growth promoting substances in animal and poultry farms. (Mara C, 2001). The incorrect use of these drugs in veterinary field can lead to the existence of residues in animal derived foods that affect human health (Mara C, 2001).

2.2.2: Classification of Antibiotics

According to Wang, 2012 Antibiotics can be classified in the following ways; Broad spectrum antibiotics where Broad-Spectrum Antibiotics are classes of antibiotics that work against an extensive and wide range of disease-causing bacteria and they target both gram positive and negative bacterial groups and they are often grouped by their abilities to act upon the different bacterial groups, examples include; Doxycycline, Minocycline, Aminoglycosides, Ampicillin, Amoxillin to mention (Wang, 2012). but a few while Narrow spectrum antibiotics are active against a selected group of bacterial types and they can act on either gram positive or negative but not both thus used for the specific infection which when causative organisms are known and examples include; Azithromycin, Clarithromycin, Erythromycin and Clindamycin.

2.2.3 Mode of action of Antibiotics

This involves the disruption of essential processes or structures in the bacterial cell and this is either through killing the bacterium or slowing down bacterial growth thus antibiotics can be bactericidal or bacteriostatic and this is achieved through Inhibition of cell wall synthesis, inhibition of protein synthesis, inhibition of nucleic acid synthesis, inhibition of the synthesis of essential metabolites and injuring the plasma membrane (Wang, 2012).

2.2.4: Common Antibiotics used in Poultry

A wide range of antibiotics are used in poultry not only to treat disease but also to maintain health, promote growth and enhance feed efficiency (Gaudin, 2004). Antibiotic usage has facilitated the efficient production of poultry, allowing the consumer to purchase, at a reasonable cost, high quality meat and eggs (Donoghue, 2003). Broiler chicken is often grown actively with antibiotics to attain maximum weight within a short period of time (Nonga, 2009).

In Uganda penicillin are among the most widely used group of antibiotics (Mitema, 2001). The uncontrolled and unlimited use of these antibiotics may however lead to the accumulation of undesirable residues in the animals treated and their products (Mitema, 2001).

Drug residues in the edible portions of the animal usually occur because the withdrawal period has not been observed (Mitema, 2001). Benzylpenicillin (penicillin G) is widely used in Kenya to treat specific infections and as a prophylactic (Lee, et al. 2001). It is administered as one or more of a variety of salts which are used to prolong the activity of the drug. These can be the soluble sodium or potassium salts or the longer acting procaine and benzathine salts (Lee, et al. 2001). Concern has been expressed over the possible presence of residues of this drug in foods of animal origin due to the occurrence of penicillin hypersensitivity in humans and development and transfer of antibiotic resistance between animals and man (Mitema, et al. 2001).

2.2.4.1: Tetracyclines

The tetracyclines were discovered in the 1940s, they are a family of antibiotics that inhibit protein synthesis by preventing the attachment of aminoacyl-tRNA to the Ribosomal acceptor (A) site (Chopra, 2001). Tetracycline consists of a common four-ring structure to which a variety of side chains are attached (Prescott, 2002). Chlortetracycline and oxytetracycline were the first members of the tetracycline group to be described (Michalova, 2004). Subsequently, a number of important semisynthetic tetracyclines were developed, e.g., doxycycline and minocycline (Michalova, 2004).

Tetracyclines are the most commonly prescribed antibiotics; they have played an important role in veterinary medicine because of their broad-spectrum activity and low cost, tetracyclines (TCs) including tetracycline (TC), Oxytetracycline (OTC), Chlortetracycline (CTC) and doxycycline (DC) are widely used in animals for both prevention, treatment and as feed additives to promote growth. (Mesgariabbasi, 2011).

The widespread utilization of TCs leads to an increasing resistance factor, so accurate monitoring by public health agencies is required (H. Matsumoto, 2000). Three different tetracycline resistance mechanisms have been described; active efflux of the antimicrobial, ribosomal protection and enzymatic inactivation of the drug. All these mechanisms are based on the acquisition of one or several tetracycline resistance determinants, which are widely distributed among bacterial genera. Additionally, mutations in the rRNA, multidrug transporter systems or permeability barriers may be involved in the resistance to several antimicrobials including tetracyclines (Michalova, 2004).

Figure 2.1: Chemical Structure of Tetracyclines

2.2.4.2 β-lactams

The β-lactam group is one of the most important families of antibiotics used in veterinary medicine and has been widely used for decades in animal husbandry (Konieczna, 2007). This group consists of penicillin and cephalosporins. The most common members of the penicillin used in veterinary practice are benzyl penicillin, amoxicillin, ampicillin and penicillin G (Kowalski, 2007). The extensive use of penicillin may cause the presence of their residues in food products of animal origin and may have side effects to consumers (Kowalski, & Konieczna, 2007). Moreover, penicillin residues in food products may be responsible of allergic reactions in humans and promote the occurrence of antimicrobials resistant bacteria (Kowalski, &Konieczna, 2007).