A cross-sectional survey was conducted in 265 selected health facilities and eight transport companies in 45 districts across nine regions in Ghana from November to December 2023 by Jhpiego and Ghana Health Service African Field Epidemiologists Network (AFENET).

A multistage sampling technique was used to select regions, districts, and health facilities for the assessment. For the purpose of this study, the country was divided into three geographical zones: Northern, Middle, and Southern (Figure 1). Geographic stratification was employed as a design approach to ensure national-level representation of Ghana's SRS. Simple random sampling using the lottery method was used to select three regions from each zone, adding up to 9 regions. In selected regions, all Regional Health Directorates (RHDs) and teaching hospitals were included in the assessment. RHDs are primarily responsible for the coordination and management of specimen referral at the district and sub-district levels, while teaching hospitals serve as main referral hospitals in the regions.

In the selected regions, districts with high Outpatient Department (OPD) attendance facilities were selected. In all, 45 districts were included in the assessment. All District Health Directorates (DHDs) of selected districts were included in the assessment. This is because DHDs are responsible for the coordination and management of SRS at the district level. Also, OPD attendance was used because it remains a frequently essential indicator in disease surveillance and control programs. Focusing on high-OPD facilities allows the assessment to acquire a more comprehensive picture of the SRS performance.

In addition, hospitals, health centers, and Community-based Health Planning and Services (CHPS) with high OPD attendance were also selected (for hospitals: monthly OPD attendance ≥500; health centers ≥200, and CHPS ≥150). All transport companies transporting specimens in the various sub-districts, districts, and regions were also included in the assessment. Lastly, all reference laboratories in the regions were also included.

Surveillance and disease control officers, health directors, and laboratory scientists at the district, regional, and national levels were purposively selected for interview based on their respective roles in the SRS at each level.

Data was collected over 4 weeks during November–December 2023. A national Technical Working Group (TWG) was formed to review and adapt the African Society for Laboratory Medicine (ASLM) specimen referral tool to fit Ghana's context (14). This tool is semi structured (allowing the capture of both quantitative and qualitive data from participants) and comprised five questionnaires targeting the following areas: (1) MoH or Health Management Team questionnaire captured information on the landscape, laboratory network/infrastructure, referral network structure, policies, and logistics; (2) reference laboratory or Hub questionnaire captured information on testing capacity, laboratory network structure, quality of specimen, standard operating procedures (SOPs), and biosafety and biosecurity; (3) referring facility questionnaire captured information on systems/communication, type of samples referred, and tracking system; (4) disease program questionnaire captured summary of diagnostic capacity for specific programs such as TB and HIV and; (5) transporter questionnaire captured information on specimen transportation and results reported directly from the transporter or the manager.

Eighteen data collectors were recruited for the survey and assigned to teams based on geographical zones. A 3-day residential training was organized for the data collectors to deepen their understanding of the methods and procedures for conducting the survey. The training focused on the effective use of the KoboCollect application (https://kf.kobotoolbox.org), covering its configuration and navigation to ensure reliable digital data capture. Participants were also guided through the content and structure of the data collection tool. Core ethical principles, including data integrity and confidentiality, were emphasized and aligned with global best practices. Furthermore, sessions on effective interviewing techniques were conducted to foster meaningful engagement with respondents. The cadre of staff selected for this training were medical laboratory scientists and disease control officers with in-depth knowledge and experience in specimen management. These participants were drawn from the GHS, Christian Health Association of Ghana, research institutions, and the Veterinary Services Directorate. Each team comprised two data collectors and a team supervisor to conduct face-to-face interviews in their assigned zones using KoboCollect during November–December 2023. Additionally, monthly records (January 2023–November 2023) on SRS performance, such as TAT, were reviewed. In total, 273 respondents were interviewed during the survey. These interviewees comprised a broad array of key management members at the regional, district, and sub-district levels of the healthcare system. Technical officers at the various healthcare facilities were also interviewed. These included medical laboratory scientists, surveillance officers, and program coordinators. In addition, the leadership of teaching hospitals, public health reference laboratories, Allied Health Directorate, Veterinary Services Directorate, and clinical laboratory units were also interviewed. Management officials from courier and public transport services responsible for specimen referral were also interviewed.

Electronic data was exported from KoboCollect into Microsoft Excel 2019 and analyzed using Stata 17/MP (StataCorp, USA). Consistency and appropriateness of responses were verified through follow-ups with regional and district health facilities. Descriptive statistics, including frequencies and proportions, were used to summarize the data. Data from health facilities in the nine regions from the three zones were aggregated for analysis, ensuring a comprehensive national-level analysis of SRS performance.

The study protocol was reviewed and approved by the GHS Ethics Review Committee (GHS-ERC: 015/12/23), Johns Hopkins BSPH (IRB No: 24573/MOD4630), and US CDC (Accession #: CGH-CSIB-7/16/20-95e37). Written approval was also sought from the Director General of GHS, as well as regional and district health directorates of the assessed regions.

Ghana has a well-defined tiered laboratory network consisting of one national and three zonal public health reference laboratories, teaching hospitals, and regional, district, and health center laboratories. The national and zonal public health laboratories fall under the Department of Public Health Laboratories within the Public Health Division of GHS. The GHS is an agency under Ministry of Health responsible for the implementation of national health policies. All other clinical laboratories are coordinated by the Clinical Laboratory Unit within the Institutional Care Division of GHS. Academic research laboratories, such as Noguchi Memorial Institute for Medical Research (NMIMR), affiliated to the University of Ghana and Kumasi Center for Collaborative Research (KCCR) under Kwame Nkrumah University of Science and Technology, support surveillance activities in the country. The national laboratory network has advanced capability to test priority diseases under surveillance and vaccine-preventable diseases at the NPHRL, NMIMR, and KCCR (Figure 2).

Several pathways were utilized for specimen referral depending on the location of the collection point and the type of specimen involved (Figure 3). For priority diseases or diseases under surveillance, including polio, meningitis, yellow fever, and measles, about 53% (105/199) of health facilities (CHPS compounds, Health Centers, and hospitals) sent specimens to the DHD where they were batched for onward submission to reference or specialized laboratories for testing on ad hoc basis. Specimens were labeled with identifiers such as epidemiology number, type of specimen, and address of the laboratory. Motorcycles were typically used to transport specimens from the lower facilities to the next tier(s). The majority of specimens (86%; 171/199) sent to the DHDs were transported by healthcare providers via public vehicles (62%; 82/133). Other facilities (21%; 21/199) sent specimens directly to the reference laboratory for testing or were sent to storage points at the RHD for transport to the testing laboratories (18%; 36/199). Results were tracked via phone calls, referral logs, and the Surveillance Outbreak Response Management and Analysis System (SORMAS). Results were mainly returned electronically (61.8%; 123/199) in addition to using paper-based forms (32.2%; 64/199). The NPHRL also shipped selected biological specimens to WHO-accredited laboratories such as the Pasteur Institute in Dakar for confirmation.

For TB and HIV, all respondents indicated that specimens were transported using a hub and spoke system based on geographic proximity. This model connects facilities with varying levels of diagnostic capacities, where spoke facilities with limited capacity for a particular test refer specimens to “hubs” which are typically well-equipped for testing. TB specimens were collected from the peripheral facilities (spokes) and transported to the “hub” laboratory for testing. Specimens received for tests that could not be performed at the receiving “hub” laboratory were subsequently transferred to the next highest level laboratory that could perform more complex testing, such as testing for extensive drug resistance (XDR) in TB specimens. Results were primarily returned electronically through the TB GxAlert^® (SystemOne, USA) in addition to phone calls and paper-based systems. Specimens were tracked using phone call follow-ups and referral logbooks. For HIV viral load (VL) and early infant diagnosis (EID), specimens were sent to strategically created laboratory hubs for testing. There were 137 GeneXpert sites providing HIV VL and or EID services across the country at the time of the assessment. Two sites, Sunyani and Korle Bu Teaching Hospitals, also provided HIV VL and EID testing using conventional polymerase chain reaction.

Differences were observed with respect to the overall TAT for specimens collected and tested for different pathogens by the system. For example, for testing of a suspected cholera specimen, an average of 12 days was spent from specimen collection to results delivery, with delays occurring at the collection points (up to 4 days). The average TAT for a polio specimen was 20–30 days. An average of 15 days was spent from the collection of an anthrax specimen to the results delivery. COVID-19 and meningitis, had an average of 7 and 12 days between specimen collection to results delivery, respectively (Figure 4). The prolonged TAT for specimen transport was influenced by multiple factors at collection points, including shortages of essential materials like containers, collection kits, and cold chain facilities, which delayed specimen collection and handling (Table 2). Unreliable transportation systems, particularly in rural areas, further contributed to delays, as poor road networks, limited vehicles, and geographic barriers made timely specimen delivery difficult. Additionally, inadequate funding, often dependent on unpredictable sources, hampered transport logistics, affecting vehicle availability and fuel supplies. Unequal and inappropriate distribution of workforce, insufficient training, and weak coordination between collection points and laboratories exacerbated the delays.

An estimated 66% (88/133) of health management teams at the RHD/DHD had the packaging guidelines or SOPs for biological specimens; 59% (79/133) of RHD/DHD health management teams provided training to RHD/DHD workers on proper handling and transportation (Table 2). However, only 28% (55/199) of referring facilities had and were aware of these packaging guidelines or SOPs, and less than half (45%: 90/199) of health workers in these facilities were trained on the proper collection, handling, and packaging of specimens. Triple-packaging materials were not readily available (43%: 57/133) for specimen transport by health management teams for specimen transportation. None of the transport companies had packaging guidelines or SOPs for biological specimens, and transporters were not trained on the proper collection, handling, packaging, or transportation of specimens. Conversely, all reference laboratories had packaging guidelines/SOPs for biological specimens, with 70% (7/10) of the reference laboratories have had at least one health worker adequately trained on how to properly collect, handle and package specimens and 70% (7/10) reference laboratories consistently have triple-packaging materials (Table 3).