ISOLATION AND IDENTIFICATION OF FUNGI FROM SELECTED BREAD

SOLD IN CAFETERIA OF COLLEGE OF AGRICULTURE

SCIENCE AND TECHNOLOGY

ABSTRACT

Bread is one of the most widely consumed staple foods in Nigeria and is susceptible to fungal contamination during production, handling, storage, and sale. This study investigated the isolation and identification of fungi from selected bread samples sold in the cafeteria of the  College of Agriculture, Science and Technology (CAST), Obubra, in Cross River State. Ten (10) bread samples comprising sliced packaged bread, unpackaged loaves, and burger rolls were purchased aseptically from the cafeteria and analysed using standard mycological procedures. Fungal isolation was carried out on Potato Dextrose Agar (PDA) and Sabouraud Dextrose Agar (SDA), and identification was based on macroscopic colony characteristics and microscopic morphological features. Results revealed that all ten samples (100%) harboured at least one fungal species. A total of four genera of fungi were identified: Aspergillus (90%), Penicillium (80%), Rhizopus (60%), and Mucor (30%). The specific isolates identified were Aspergillus niger, Aspergillus flavus, Penicillium expansum, Penicillium citrinum, Rhizopus stolonifer, Rhizopus microsporus, and Mucor mucedo. Aspergillus niger was the most frequently encountered isolate. Mean pH and moisture content of the samples were 5.83 ± 0.22 and 35.60 ± 2.44%, respectively. The high prevalence of mycotoxigenic fungi such as Aspergillus flavus and Penicillium expansum poses significant public health risks, particularly with regard to the production of aflatoxins and patulin. The findings underscore the need for improved hygiene practices in bread storage and handling in institutional cafeterias, as well as stringent regulatory oversight of food outlets within academic environments. It is recommended that cafeteria operators adopt refrigerated storage for bread products and that consumers be educated on the risks of consuming mould-contaminated food.

Keywords: Bread, Fungi, Contamination, Aspergillus, Penicillium, Mycotoxins, Food Safety, Cafeteria

TABLE OF CONTENTS

Declaration                                                                            ii

Certification                                                                         iii

Dedication                                                                            iv

Acknowledgements                                                               v

Abstract                                                                                vi

Table of Contents                                                                vii

List of Tables                                                                        ix

List of Abbreviations                                                             x

CHAPTER ONE: INTRODUCTION                               1

1.1 Background of the Study                                                 1

1.2 Statement of the Problem                                                3

1.3 Aim and Objectives of the Study                                    4

1.4 Research Questions                                                         4

1.5 Significance of the Study                                                5

1.6 Scope and Limitations of the Study                                5

1.7 Methodology Overview                                                   6

CHAPTER TWO: LITERATURE REVIEW                  7

2.1 Bread: Composition and Consumption Patterns in Nigeria                                                                                   7

2.2 Bread Contamination                                                       8

2.3 Food Spoilage Fungi                                                       9

2.4 Common Fungi Associated with Bread                        10

2.4.1 Aspergillus                                                                 10

2.4.2 Penicillium                                                                 11

2.4.3 Rhizopus                                                                     12

2.4.4 Mucor                                                                         12

2.5 Public Health Implications of Fungal Contamination of Bread                                                                                   13

2.6 Empirical Review of Related Studies                            14

2.7 Research Gap                                                                 15

CHAPTER THREE: MATERIALS AND METHODS 16

3.1 Study Area                                                                     16

3.2 Sample Collection                                                         16

3.3 Materials and Apparatus                                                17

3.4 Culture Media Preparation                                            17

3.5 Isolation of Fungi                                                          18

3.6 Identification of Fungal Isolates                                    19

3.7 Microscopic Examination                                             19

3.8 Incubation Procedures                                                   20

3.9 Determination of pH and Moisture Content                  20

3.10 Data Analysis                                                              20

CHAPTER FOUR: RESULTS AND DISCUSSION     21

4.1 Physicochemical Properties of Bread Samples             21

4.2 Frequency of Fungal Isolates Across Bread Samples   21

4.3 Macroscopic and Microscopic Characteristics of Isolates                                                                                             22

4.4 Occurrence of Fungi by Bread Type                             23

4.5 Discussion                                                                     23

CHAPTER FIVE: SUMMARY, CONCLUSION AND RECOMMENDATIONS                                                  27

5.1 Summary                                                                       27

5.2 Conclusion                                                                     28

5.3 Recommendations                                                         29

References                                                                          30

LIST OF TABLES

Table 4.1: Physicochemical Properties (pH and Moisture Content) of Bread Samples                                                 21

Table 4.2: Frequency of Fungal Isolates Across Bread Samples                                                                               21

Table 4.3: Macroscopic and Microscopic Characteristics of Identified Fungal Isolates                                                    22

Table 4.4: Occurrence of Fungi by Bread Type                  23

LIST OF ABBREVIATIONS

ADA         Aspergillus Differential Agar

CAST        College of Agriculture Science and Technology

CFU         Colony Forming Unit

FAO         Food and Agriculture Organization

HACCP       Hazard Analysis and Critical Control Points

NAFDAC      National Agency for Food and Drug Administration and Control

PDA         Potato Dextrose Agar

PH          Potential of Hydrogen (pH)

SDA         Sabouraud Dextrose Agar

SD          Standard Deviation

WHO         World Health Organization

CHAPTER ONE

INTRODUCTION

1.1 Background of the Study

Food safety remains one of the most critical public health challenges in developing nations, including Nigeria. Among the various food categories susceptible to microbial contamination, bakery products particularly bread occupy a position of significant concern owing to their widespread consumption, nutrient-rich composition, and inherent susceptibility to spoilage (Awujo et al., 2022). Bread is a staple food consumed daily by millions of Nigerians across all socioeconomic strata, making its microbiological safety a matter of paramount importance.

Bread is prepared primarily from wheat flour, water, yeast, salt, and various additives. Its nutritional profile, which includes carbohydrates, proteins, fats, vitamins, and minerals, creates an ideal substrate for the proliferation of microbial agents — especially fungi (Garba et al., 2022). Fungal contamination of bread occurs along the entire supply chain, from the raw materials and production environment to packaging, transportation, storage, and point-of-sale handling. In institutional settings such as university cafeterias, these risks are amplified by high product turnover, suboptimal storage conditions, and inadequate temperature regulation.

Fungi implicated in bread spoilage predominantly belong to the genera Aspergillus, Penicillium, Rhizopus, and Mucor (Kareem, 2023). These organisms are ubiquitous in the environment and can colonise bread substrates under ambient conditions. Beyond causing visible spoilage characterised by mycelial growth, off-odours, and textural changes, certain spoilage fungi are capable of producing mycotoxins — secondary metabolites with documented carcinogenic, mutagenic, and immunosuppressive properties in humans and animals (Pitt & Hocking, 2009). Notable mycotoxins associated with bread-spoilage fungi include aflatoxins (produced by Aspergillus flavus and Aspergillus parasiticus), ochratoxin A (produced by Aspergillus ochraceus), and patulin (produced by Penicillium expansum).

In Nigerian academic institutions, cafeterias serve as central food outlets for students, academic staff, and administrative personnel. The bread sold in these outlets is often sourced from local artisanal bakeries operating under varying degrees of compliance with food safety standards. Studies have documented poor sanitation conditions in Nigerian bakeries, inadequate personnel hygiene practices, and the use of sub-standard ingredients, all of which contribute to elevated microbial loads in finished bread products (Muhammad & Galadima, 2022). The College of Agriculture Science and Technology (CAST) cafeteria, like many institutional food service environments, is exposed to these challenges.

The isolation and identification of fungi from bread sold in the CAST cafeteria is therefore not merely an academic exercise but a necessary investigation with direct implications for the health and well-being of the institution's community. By characterising the mycobiota of cafeteria bread, this study provides an evidence base for targeted interventions aimed at improving food safety standards within the institution.

1.2 Statement of the Problem

Despite growing awareness of mycotoxin-related health risks, fungal contamination of bread sold in institutional settings in Nigeria remains inadequately investigated. The CAST cafeteria serves a sizeable population of students and staff who consume bread daily, often without awareness of potential fungal hazards. The warm and humid tropical climate of Nigeria accelerates fungal growth on baked products, particularly when bread is stored in open, unrefrigerated display counters — a common practice in Nigerian cafeterias and food kiosks.

Previous studies conducted in other Nigerian states have consistently demonstrated high fungal loads in commercially produced and locally baked bread (Awujo et al., 2022; Garba et al., 2022; Muhammad & Galadima, 2022). However, data specifically addressing the mycological quality of bread sold in the CAST cafeteria are non-existent. This knowledge gap precludes the formulation of institution-specific food safety policies and exposes cafeteria patrons to poorly characterised microbial risks.

Furthermore, the increasing student population in Nigerian higher institutions means that more individuals are relying on institutional food outlets for their daily nutritional needs. A single outbreak of mycotoxicosis traceable to contaminated cafeteria food could have devastating health consequences and institutional repercussions. There is therefore an urgent need to generate baseline data on the fungal quality of bread sold within this institution.

1.3 Aim and Objectives of the Study

The aim of this study is to isolate and identify fungi from selected bread samples sold in the cafeteria of the College of Agriculture Science and Technology.

The specific objectives are to:

•       Collect bread samples from the CAST cafeteria and assess their physicochemical properties (pH and moisture content);

•       Isolate fungal contaminants from the bread samples using standard mycological techniques;

•       Identify the isolated fungi based on macroscopic colony characteristics and microscopic morphological features;

•       Determine the frequency and distribution of fungal isolates across different bread types;

•       Discuss the public health implications of the identified fungal species and compare findings with related studies.

1.4 Research Questions

This study seeks to answer the following research questions:

1.    What are the physicochemical properties (pH and moisture content) of bread samples sold in the CAST cafeteria?

2.    Which fungal species are associated with bread sold in the CAST cafeteria?

3.    What is the frequency and prevalence of fungal isolates across different bread types?

4.    What are the macroscopic and microscopic characteristics of the isolated fungi?

5.    What are the public health implications of the fungi identified in relation to mycotoxin production?

1.5 Significance of the Study

The findings of this study are significant for several reasons. First, they provide baseline mycological data for the CAST cafeteria, which can inform institutional food safety policies and standard operating procedures for bread handling and storage. Second, the identification of mycotoxigenic fungi such as Aspergillus flavus and Penicillium expansum will alert cafeteria management and consumers to the potential health risks associated with mould-contaminated bread.

Third, this study contributes to the growing body of scientific literature on food safety in Nigerian academic institutions, a relatively underexplored area compared to commercial food markets and bakeries. The data generated will be valuable to food scientists, public health practitioners, nutritionists, and regulatory bodies such as the National Agency for Food and Drug Administration and Control (NAFDAC).

Finally, the study will serve as a reference point for future investigations into mycological contamination of other food products sold in institutional settings and may stimulate interest in broader food safety audits within the institution.

1.6 Scope and Limitations of the Study

Several limitations are acknowledged in this research. The cultural and morphological identification methods intended to be employed may not discriminate between closely related species or cryptic fungal taxa that require molecular tools for definitive identification. The study also does not quantify mycotoxin levels in the samples, which would require high-performance liquid chromatography (HPLC) or enzyme-linked immunosorbent assay (ELISA) analyses beyond the scope of an undergraduate project. Additionally, seasonal variation in fungal contamination was not captured, as sampling was conducted within a single semester period.

1.7 Methodology Overview

Ten bread samples will be aseptically collected from the CAST cafeteria and transported to the Microbiology laboratory for processing. The samples will be analysed for pH and moisture content prior to mycological examination. Fungal isolation will be performed on Potato Dextrose Agar (PDA) and Sabouraud Dextrose Agar (SDA) using the dilution plating method for crumb samples and direct surface plating for crust samples. Cultures will be incubated at 25°C for 5–7 days. Identification of fungal isolates will be based on colony morphology, pigmentation, texture, and lactophenol cotton blue-stained microscopic preparations. Data were presented as frequency and percentage occurrence and analysed descriptively.