1 Introduction

Coronavirus causes a novel infectious disease otherwise known as COVID-19. It is a major global health crisis of our time that has caused severe illness and death around the globe. It is caused by Severe Acute Respiratory Syndrome Corona 2 Virus (SARS-COV-2 Virus). It was first discovered in Wuhan China in December 2019 and since then has spread across the globe with overwhelming health, social, and economic impact [1]. The World Health Organization (WHO) declared the COVID-19 outbreak a public health emergency of global concern (Pandemic) in January 2020[2]. The disease was first reported in Africa in February 2020, as of 31st December 2020 COVID-19 had infected over eighty-two million people and killed more than 1.8 million worldwide [3]. The economic and social disruption caused by the pandemic was devastating, it has unprecedented challenges to public health. The National Agency for Food and Drug Administration and Control (NAFDAC) regulates the quality of vaccines imported into Nigeria to safeguard public health. Despite the anecdotal belief about countries with emerging economies, in this communication, the Agency provides a summary of the quality control (QC) measures put in place to ensure the public health safety of the citizenry who received the vaccines.

The swift development of effective vaccines against COVID-19 is one of the greatest public health scientific achievements to urgently mitigate severe illness and death caused by the virus. The development of COVID-19 Vaccines involved thousands of scientists across the globe with the collaborative effort of governments, public health agencies, and private pharmaceutical industries working tremendously to develop vaccines to mitigate the scourge of the disease. The Chemistry, Manufacturing, and Control (CMC) involved in the development of vaccines starting from initial research to distribution to vaccination which takes about a decade was reduced to a year from development to vaccination. The development was extremely fast and successful with the emergence of multiple vaccines within one year outbreak of the disease[4]. Several manufacturers obtained emergency use authorization for their products which are now used worldwide for immunization [5]. These vaccines required cold storage and were exported to many developing countries with lower resource limitations. Many low-income countries have limited capacities to verify the quality of these products. This study documents the various processes that NAFDAC used to regulate the quality of the COVID-19 vaccines that were imported for use within Nigeria.

The vaccines were developed using different technologies including the use of whole virus/ bacterium, germ parts/subunit, viral vector, or genetic material DNA or RNA that provides the instructions for making specific proteins to prompt the body to create antibodies that confer immunity to the recipients [6]. The US pharmaceutical company Pfizer in partnership with BioNTech and Moderna use messenger RNA (mRNA) which has never been approved for commercial use in human before the COVID-19 pandemic [7]. Other approved COVID-19 vaccines AstraZeneca, and Janssen Johnson & Johnson use viral vectors or modified versions of a different virus[8].

More than thirty vaccines have been approved by Emergency Use Listing (EUL) in different countries around the world. EUL is a World Health Organization (WHO) initiative which is a risk-based procedure for assessing and listing unlicensed vaccines, therapeutics, and in vitro diagnostics to expedite the availability of the product to the people affected by a public health emergency [9]. During the COVID-19 pandemic, EUL was introduced because there was a need to cut down the timelines of regulatory approvals and approve products that will mitigate illness and reduce death rates. EUL is a fast track, but rigorous process designed to bring impactful products as quickly as possible to all those in need on a risk versus benefit evaluation of the product. The following COVID-19 Vaccines: Covishield, Janssen, AstraZeneca, Pfizer BioNTech, and Moderna vaccines were approved and imported for use in Nigeria via the emergency use listing pathway.

1.1 NAFDAC’s role in public health

The National Agency for Food and Drug Administration and Control (NAFDAC) has the mandate to safeguard the health of the public by ensuring that quality and safe regulated products are manufactured, imported, exported, sold, distributed, and used by the public. The Agency also continuously monitors the quality of thermolabile-regulated products along the supply chain to ensure the stability of the product is maintained throughout its life cycle [10]. NAFDAC performs six critical functions concerning vaccine regulations, these functions include, licensing, inspection, laboratory testing, lot release, surveillance, and clinical trial.

NAFDAC National Control Laboratory for Vaccine and Other Biologicals (NCLVB) was established in 1999 to ensure the safety, efficacy, potency, and lot release of vaccines imported and used in Nigeria. In this study, we considered three functions: laboratory testing, lot release, and surveillance. The purpose of this study was to evaluate the safety, quality, traceability, and lot release of COVID-19 vaccines imported, distributed, and used in Nigeria from March 2021 to March 2022.

1.2 Laboratory testing

One of the key functions of the Agency is to conduct quality analysis on regulated products to make pronouncements regarding their quality, safety, and effectiveness. The public is always anxious to know how extensively vaccines are tested for safety, especially new vaccines, the level of acceptance most times depends on the knowledge of the safety of the vaccine. The standard for testing and monitoring vaccines is higher than it is for most other medicines because they are given to healthy individuals. Vaccines go through vigorous testing for safety and effectiveness before they are approved for public use. This function (Laboratory testing) for vaccines including COVID-19 vaccines is carried out by NCLVB under the Directorate of Laboratory Services in the Agency. In addition to the Agency’s drug laboratory, NCLVB is an ISO 9001:2015 and ISO 17025:2017 certified laboratory, with trained and competent analysts qualified to conduct safety and quality evaluation of vaccines[11].

Upon importation of any batch of the COVID-19 Vaccine, The Federal Ministry of Health notifies the Agency. The agency immediately responds by conducting a sampling of the vaccines. Vaccines are cold chain products and as such their temperature should be maintained throughout the life cycle of the product. During an inspection by the Agency, the temperature of storage of the vaccines is observed and recorded. The observed temperature should be within the recommended temperature approved by the manufacturer. The product is carefully observed visually for damages, leakage, and expiration. The accompanying documents are collected cited and previewed for completeness and clarity. The test Request Form (TRF) is filled by a representative from the National Primary Health Care Development Agency.

The Federal Ministry of Health issued the mandate for importation of COVID-19 vaccine to the National Primary Health Care Development Agency (NPHCDA). They are responsible for the distribution, storage, traceability (in collaboration with NAFDAC), and vaccination of COVID-19 vaccines. The NAFDAC NCLVB carries out analysis and review of the Summary Lot Protocol before the issuance of Lot Release certificates before they are released for immunization. COVID-19 Vaccine was first imported into the country for use in March 2021, and Emergency Use Authorization (EUA) was issued to the manufacturers for their product. EUA is a mechanism to facilitate the availability and use of medical countermeasures including vaccines during public health emergencies. Acknowledging the risks associated with the EUA process this paper seeks to provide evidence of NAFDAC’s due diligence to ensure the safety of the vaccines imported and used in the country within the COVID pandemic era.

2 Method

This paper presented a case study that provided summaries of laboratory test results carried out on different COVID-19 vaccines imported into Nigeria within 2021–2022. From the study design, temperature data were collected as an indicator of their quality dispositions. This report also provides details on how the COVID samples were handled on receipt and the chain of custody of how they were tested in the laboratory. It also provides abridged results of laboratory quality-control tests and other parameters used to certify the vaccine's safety for use within Nigeria and provides reports from the Ports Inspection directorate. The directorate provided complementary data to ensure the imported vaccines were handled appropriately from the point of entry into the country to the distribution of the products to reach the populace.

The testing of the COVID-19 Vaccines was done following international best practices for vaccine evaluation. The laboratory equipment used to test the vaccines was adequately calibrated by an ISO-accredited measurement body with traceability. The testing was carried out by competent analysts qualified to conduct safety and quality testing of vaccines using applicable pharmacopeia or test methods as recommended by the manufacturer. Approved Standard Operating Procedures (SOPs) were properly followed to ensure the quality of the processes and international good laboratory practices were employed to ensure the reliability of the test results.

3 Results and discussions

Various COVID-19 vaccines were imported into Nigeria during the peak of the pandemic. Table 1 summarizes the recommended storage conditions for these thermolabile products.

Table 1 Recommended storage temperature for the COVID-19 vaccines approved and imported for use in Nigeria
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The sampled vaccine batches were transported in a way not to compromise the integrity of the vaccine (cold box with an ice pack or dry ice) depending on the required storage temperature and a data logger to ascertain the temperature on arrival to the laboratory for analysis. It should be noted that some frozen vaccines can be safely stored between 2 °C and 8 °C for up to 30 days before use and can never be re-frozen after thawing.

The storage temperatures of samples of the COVID-19 vaccines were routinely assessed in NAFDAC laboratories. Table 2 lists some of the verification results for these assessments.

Table 2 Temperature of storage on arrival of some sampled batches of COVID-19 vaccines transported to the laboratory for analysis
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As shown in Table 2 above, the batches of samples of the COVID-19 Vaccines received were stored and received at the recommended temperature approved by the manufacturer.

After the temperature of the vaccines was certified as meeting the recommended ranges, the products were accepted into the laboratory for further analysis.

The Laboratory Information Management System (LIMS), an electronic information management system designed for managing laboratory information, was used to generate the test request identification number and the laboratory reference number. The vaccine samples were assigned to various units for further analysis using the LIMS.

3.1 Quality control testing

The vaccines were analyzed based on the filled test request form using applicable pharmacopeia as recommended by the manufacturer to ensure compatibility of the test result.

The following parameters were conducted on the vaccine pH, Physical appearance, Abnormal toxicity, Bacterial Endotoxin (BET), Sterility, Protein content, DNA to Protein ratio, and extractible volume.

3.1.1 Sterility testing

Sterility testing is a microbiology test that establishes the absence of viable organisms in a sterile product, this test is used to ascertain manufacturers' claim of sterility. It is one of the safety tests conducted on vaccines and other related sterile products to ensure the absence of contaminants that may cause infection. This involves incubating the samples in growth media where the microorganism can be detected. Growth of viable organisms was observed in one of the batches of one brand of the Vaccine tested. The results of testing conducted on the batch show it was not sterile, See Table 3. To maintain confidentiality, the results for tests conducted will be de-identified for the various brands.

Table 3 Summary of sterility test conducted on COVID-19 vaccines imported for use in Nigeria from March 2021 to March 2022
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In line with the SOP for Out of Specification (OOS) for the laboratory, confirmatory replicate tests were performed to confirm the failed batch of the vaccine. Highlights of Table 4 below indicated that the unsatisfactory results were confirmed after 2 other qualified analysts tested the products as per the OOS SOP.

Table 4 Summary of OOS result obtained
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3.1.2 Bacteria endotoxin test (BET)

The bacterial Endotoxin Test is an in-vitro assay for the detection and quantification of bacterial endotoxins present in the vaccine. Endotoxins are heat-stable lipopolysaccharides found in the outer membrane of gram-negative bacteria and are potential contaminants that can be introduced during the manufacturing of vaccines. Parental pharmaceutical products such as COVID-19 vaccines undergo endotoxin tests because endotoxins are pyrogenic in humans and can result in serious physiological reactions. The gel clot method was used in testing BET in COVID-19 Vaccines, this method can show the presence of endotoxins with the presence of gel cloth formed in the sample tube when introduced to Limulus Amebocyte Lysate (LAL). The gel clot assay is carried out in tubes that are placed in a block heater at 37 °C, after an hour incubation period the tubes are flipped at 180°, and a firm clot deposited at the bottom of the tube indicates a positive reaction but if the liquid flows down freely down the side of the tube indicating the absence of clot shows a negative result. Table 5 provides a summary of the BET assessments for the batches of COVID-19 vaccines evaluated in this study. All batches tested met the specifications for BET.

Table 5 Summary of bacteria endotoxin test carried out on COVID-19 vaccines imported for use in Nigeria from March 2021 to March 2022
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The result above shows that all the COVID-19 vaccines imported for use in Nigeria within the specified period were satisfactory for the Bacteria Endotoxin Test.

3.1.3 Abnormal toxicity test

Abnormal Toxicity Test (ATT) is also known as a general safety test. It is an in-vivo method used to detect potential biological contamination lethal to the health of humans. Healthy albino mice were used for this test, ten albino mice weighing 17–22 g were used. Five of the mice served as the test group and five as a control group, the test group was injected with the vaccine sample and the control group was injected with normal saline, all ten mice were injected intraperitoneally, and after injection, all the mice were observed for signs of ill health, death, or weight loss within seven days. The laboratory affirmed the safety of vaccines by conducting any additional QC tests specified in a product’s certificate of analysis (CoA). The ATT test was conducted only on Brand E which listed the tests as a parameter on its CoA and all batches tested had satisfactory results for this parameter, see Table 6

Table 6 Summary of abnormal toxicity test conducted on Brand E COVID-19 vaccines imported for use in Nigeria from March 2021 to March 2022
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3.1.4 pH test

pH tests measure the acidity or alkalinity of a solution on the logarithm scale, it is a physio-chemical test that inversely indicates the concentration of hydrogen ions in the solution. It is an important quality that reflects the chemical composition of a solution. Buffers keep the vaccine at a similar pH within the body's physiological acceptance which is between a pH of 6–8. The pH of a vaccine outside this range of pH indicates instability or degradation of the vaccine which makes it unfit for use. This parameter was conducted using the United States Pharmacopeia method, a calibrated pH meter with a temperature probe was used to measure the pH of the vaccines in triplicate and the average results were recorded. and shown in Table 7. All the COVID-19 vaccines evaluated during this period were found to meet the pH range requirement for the product.

Table 7 Summary of the pH result of COVID-19 vaccines imported for use in Nigeria from March 2021 to March 2022
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3.1.5 Physical appearance

This is a physical parameter that involves the visual observation of the vaccine to ascertain color, clarity, or absence of particles. The vaccines were observed closely under a bright light to ensure the physical appearance was as described by the manufacturer. All the products meet the manufacturer's specifications for physical appearance and are summarized in Table 8.

Table 8 Observed physical appearance vs appearance as described by the manufacturer for all the COVID-19 vaccines imported for use in Nigeria
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3.1.6 Extractible volumes

The test for extractable volume compares the volume on the vaccine label with the volume that can be extracted from the vial. This is done to ensure the vaccine can deliver the precise number of doses that the manufacturer has specified, since the number of doses imported depends on the population of vaccines, it is misleading to the public if the number of doses is less than the claimed dose due to inaccurate filling volume.

Overfilling vials with additional vaccines is a typical practice in the manufacture of vaccines. Manufacturers offer extra vaccines to help medical professionals administer the correct number of doses. Vial-kept volume, vaccine retained in the dead space of the syringe and needle, and vaccine losses during the adjustment of the dose if discharged in the air are all considered by overfilling.

This parameter was performed using the British Pharmacopeia method, which involved thoroughly extracting the vaccine into a calibrated volumetric flask, duplicating the test based on the volume of vaccine tested, and recording the average volume.

The criteria are as follows; less than 3 mL is done in five duplicates, 4–9 mL is done in triplicate and more than 10 mL is done once. The extractable volumes of the vaccines are summarized in Table 9, and all were within the specified range for the parameter.

Table 9 Result of extractible volume obtained for the COVID -19 vaccines imported for use in Nigeria from March 2021 to March 2022
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3.1.7 Protein content

The CoA for the first five batches of Covid-19 vaccines Brand D that were imported listed protein content as a critical test parameter. This parameter is to determine the protein content of the vaccine which induces cellular immunity. This parameter was conducted using the Lowry method and the results obtained for the five batches are detailed in Table 10, one unsatisfactory result was obtained.

Table 10 Result of determination of protein content for vaccines Brand D imported for use in Nigeria from March 2021 to March 2022
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Following the unsatisfactory result for the protein content of one of the batches of vaccines Brand D, an investigation was conducted and the root cause was traced to poor storage by the donor. NAFDAC laboratory followed its SOP for investigationg Out of Specification (OOS) and initiated re-analysis of the failed batch. The lab results obtained confirmed the batch to be unsatisfactory and a rejection notification was issued to disapprove the batch from being circulated in the country.

3.1.8 DNA to protein ratio

This parameter provides information about the virus particles as well as the purity of the sample; it is a direct Ultraviolet spectrophotometer test. The laboratory conducted tests for DNA to Protein ratio if indicated in the vaccines’ CoA. In this study, the first five lots of Brand D were analyzed for protein content as indicated in the CoA. However, the DNA to Protein ratio for the subsequent lots were assessed as a new parameter in the CoA for the products submitted. Similarly, the test was conducted on all lots of Brand A. Table 11 below provides the results obtained for the vaccines.

Table 11 Result of DNA to Protein ratio for vaccine Brand A and D imported for use in Nigeria from March 2021 to March 2022
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3.2 Lot release process

This is the process of evaluating each lot of a licensed product before approving its release into the market.

This is a mechanism that provides the agency with a real-time system to continuously monitor product quality through the review of documents and testing of biological products it regulates. The process of vaccine lot release comprises the following steps: Submission of documents, Review of documents, Quality control testing, and Issuance of lot release certificate.

3.2.1 Submission of documents

The documents required for submission for review are a Summary Lot Protocol (SLP), a Certificate for Analysis (CoA), and a Lot release certificate.

3.2.2 Summary lot protocol (SLP)

The summary lot protocol submitted for each vaccine batch contains at minimum the following details of the finished vaccine product submitted.

  1. 3.2.2.1.

    Composition.

  2. 3.2.2.2.

    Manufacturers and facilities involved in the manufacturing and testing of the vaccine.

  3. 3.2.2.3.

    Manufacturing methods: Detailed description of the manufacturing process.

  4. 3.2.2.4.

    Specifications and methods: description of all test methods selected to assure the identity, purity, and potency as well as lot-to-lot consistency.

  5. 3.2.2.5.

    Container and closure system.

3.3 Certificate of analysis (COA)

Certificate of Analysis is an official document that shows the result of a scientific test carried out on the product, it contains the result obtained and the specification and is required for submission for each vaccine lot. A certificate of analysis and analytical result should be provided and should contain the following information:

  1. 3.3.1.

    The batch number of the vaccine.

  2. 3.3.2.

    The result of the quality test conducted.

  3. 3.3.3.

    Name and signature of the analyst.

  4. 3.3.4.

    Name and address of the quality control laboratory.

  5. 3.3.5.

    Approval by Responsible Personnel.

3.4 Lot release certificate from country of origin

This is a certificate of lot release of vaccine lot/batch by the National Regulatory Authority of the product country of origin, it states where the product was manufactured or processed and shows that the product has been fully released for sale by the National Regulatory Authority of the country of origin. This document is required to ensure that the vaccine has been released for use in the country of origin.

All the above-mentioned documents were reviewed for all the batches of COVID-19 Vaccines imported for use in Nigeria and below is the summary of the result. The results of document reviews of COVID-19 vaccines imported and used in Nigeria from March 2021 to March 2022 were satisfactory as shown in Table 12.

TABLE 12: Result of documents review of COVID-19 vaccines imported and used in Nigeria from March 2021 to March 2022
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3.5 Issuance of lot release /rejection certificate

Table 13 lists the quantity of COVID-19 vaccine batches imported for usage in Nigeria between March 2021 and March 2022 as well as the number of lot release certificates that were issued based on a satisfactory evaluation of the supporting documentation and successful safety and quality control testing of the COVID-19 vaccine batches (refer to Table 14 for the summary of the result of safety and quality testing conducted on COVID -19 Vaccines imported for use in Nigeria from March 2021 to March 2022). However, if the document review or the quality control testing is found unsatisfactory, a rejection certificate is issued for that lot/batch and the lot will not be released for use or vaccination. As observed in the quality control testing of Vaccine brands C and D one batch of each of the two vaccines failed sterility testing and protein content, respectively. Rejection certificates were issued for failed batches of the vaccines. As shown in Table 15, a total of ninety-five batches of COVID-19 vaccines were imported between March 2021 and March 2022. Seventy-eight lot release certificates and 2 rejection certificates were issued.

Table 13 Number of lot release certificates issued for COVID-19 vaccine batches imported for use in Nigeria from March 2021 to March 2022
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Table 14 Summary of safety and quality testing results conducted on COVID-19 vaccines imported for use in Nigeria from March 2021 to March 2022
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Table 15 Summary of the number of lot release and rejection certificates issued for COVID-19 vaccine batches imported for use in Nigeria from March 2021 to March 2022
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3.6 COVID-19 pandemic and pilot of traceability in nigeria by the ports inspection directorate

Upon receipt of the first tranche of COVID-19 vaccines in Nigeria, NAFDAC initiated a pilot traceability which has since progressed to a full track and trace. All the COVID-19 Vaccines arrive through the Nnamdi Azikiwe International Airport and are transported to the NPHCDA national strategic cold store, and the following processes ensue.

3.6.1 COVID-19 vaccines unique product identification

All COVID-19 vaccines received were identified. The Astra Zeneca Covid shield brand was uniquely identified, in compliance with Global Standards 1 (GS1), and data was encoded in a data matrix containing – Global Trade Item Number (GTIN), Expiry date, Batch number, and serial number. For COVID-19 vaccines without unique serial numbers per secondary pack, NAFDAC in collaboration with GS1, carried out In-country serialization for these batches at the National cold store before distribution e.g., Moderna and Pfizer Vaccines. To achieve this, a team of NAFDAC and GS1 staff:

  1. 3.6.1.1.

    Generated serialized unique identifiers (UID) corresponding to the quantities received for each of the batches.

  2. 3.6.1.2.

    Encoded the GTIN, Batch Number, Expiry Date, and Serial Numbers in a GS1 standard Data Matrix and printed the barcode on label stickers.

  3. 3.6.1.3.

    Commissioned the printed barcode stickers by scanning them to capture the data encoded in the Data Matrix into the Traceability Information System.

  4. 3.6.1.4.

    labeled the vaccines with the already commissioned barcode stickers.

3.6.2 Assignment of global location number (GLN)

Over 400 facilities across national, zonal, state, and local government warehouses down to healthcare facilities were assigned GLNs in 36 states and the FCT where these vaccines will be distributed.

3.6.3 Product scanning along the supply chain

After commissioning, as these COVID-19 vaccines were distributed across the different levels of the supply chain, barcodes encoded with data matrix are scanned using the track genic app; an app developed by NAFDAC's technical partner GS1 and downloaded on personal mobile phones, to capture product data and events.

3.6.4 NAFDAC traceability information system (TIS)

NAFDAC TIS provides visibility for the COVID-19 vaccines in distribution by showing the scanning activities by product, location, and type of event for any period specified. This data can be visualized on a map and details of the data are presented in reactive tables. The information on the TIS assures the source, and origin of each secondary pack of the COVID-19 vaccines and can be used to detect potential infiltration of the supply chain with substandard, counterfeit vaccines from data captured during the scanning of the vaccines from the national strategic store to the point at which the secondary pack is broken and decommissioned.

More importantly, the TIS complements other extant regulatory measures such as quality control testing, lot release, and pharmacovigilance monitoring to ensure the safety and quality of the vaccines. Furthermore, this has huge implications for tracking defective batches nationwide and conducting prompt and effective recalls.

3.6.5 Role of track and trace along the supply chain

The regulatory recall of a defective batch of the COVID-19 vaccines (the batch that failed sterility testing) was supported by the data captured from the scanning of COVID-19 vaccines across the State Cold Stores. A list of locations where scanning of the defective batch was detected was extracted from the TIS and respective vaccine accountability officers in those locations were notified to retrieve the defective batch. This was done and the defective vaccines were retrieved for destruction.

4 Conclusion

A total of ninety-five batches of different COVID-19 vaccines were imported for use between March 2021 to March 2022 These batches were analyzed and summarized in Table 13 it is observed from the table that 95.5% of the COVID-19 Vaccine passed sterility testing while 4.5% failed sterility testing. There is a 100% pass of all the vaccines for the Bacteria Endotoxin Test while an 80% pass was observed for protein content conducted on five of the Vaccine brand D. DNA to protein ratio tests conducted on all Vaccine brands A and D were all satisfactory as the value obtained falls within the specification range. It was also observed that all the vaccines fall within the specification for the pH showing there was no degradation or instability of the COVID-19 vaccines imported for use in the country. The extractible volume test conducted on all the COVID-19 Vaccines as shown in Table 7 shows that all batches of the vaccine extractible volume were greater than the actual volume indicated on the label this is in line with WHO guidance on “overfilling” of vaccines which accounts for vial retain volume and vaccine retained in dead space during vaccination. The abnormal toxicity test conducted for Vaccine brand E was satisfactory as there were no signs of ill health or death observed neither was there any loss of weight observed in the animals within seven days of administration of the vaccine showing there is no biological contamination lethal to human health.

The COVID-19 Vaccines imported for use in Nigeria from March 2021 to March 2022 in which lot release certificates were issued, met the stated specifications for parameters analysed as indicated in the Certificate of Analysis and as such are safe for vaccination of the populace.