Improved detection of microbiological pathogens: role of partner and non-governmental organizations

Globally, the fight against infectious diseases is still a great force to reckon with. The ability of disease-causing organisms to spread beyond national and international borders means an infectious disease threat anywhere is a threat everywhere [1]. Thus, every country has a role to play in making the world safer from epidemics by strengthening its capacity to prevent, detect in timely manner and respond effectively to current and emerging health threats. Key health threats that could likely pose danger to human lives include increasing trend of antimicrobial resistance, zoonotic diseases, biosafety and biosecurity, weak laboratory and surveillance systems and poor work force development.

Addressing the threats of zoonosis, antimicrobial resistance, biosafety and biosecurity begins with detection of aetiological agents involved in disease outbreaks and infections. Timely and accurate detection and reporting of infectious disease outbreaks and events are critical to controlling the course of outbreaks and avoiding large-scale epidemics. Detection of microbial pathogens also enables the performance, reporting and surveillance of antimicrobial resistant microbial organisms.

Antimicrobial resistance is one of the biggest threats to global health [2]. According to WHO, there are 12 families of resistant bacteria which pose the greatest threat to human health, and these are termed priority pathogens [3]. These bacteria are further categorized into Priorities 1 (critical), 2 (high) and 3 (medium). Examples of priority 1 pathogens include Carbapenem-resistant Acinetobacter baumannii, Carbapenem-resistant Pseudomonas aeruginosa and Carbapenem-resistant, ESBL-producing Enterobacteriaceae. Priority 2 pathogens include Vancomycin-resistant Enterococcus faecium, Methicillin and Vancomycin-resistant Staphylococcus aureus, Clarithromycin-resistant Helicobacter pylori, Fluoroquinolone-resistant Salmonella spp., Fluoroquinolone-resistant Campylobacter spp., and Cephalosporin-resistant, fluoroquinolone-resistant Neisseria gonorrhoeae. Other bacteria such as Penincillin-non-susceptible Streptococcus pneumoniae, Ampicillin-resistant Hemophilus influenzae, and Fluoroquinolone-resistant Shigella spp. are referred to as Priority 3 pathogens [3]. These resistant pathogens are a threat to global health security because of their potential to cause significant economic and public health problems [4, 5].

One of the effective ways to maximize global health security and preparedness for infectious disease threat is to invest in Global Health and address global health security challenges [6]. Since June 2007, various countries have been putting in place efforts to strengthen their International Health Regulations (IHR) core capacities through the Global Health Security expanded activities. As a way of achieving these objectives, WHO conducted a Joint External Evaluation (JEE) of the IHR core capacities in 2017 in Ghana [7]. Among the key findings identified was the need to strengthen laboratory capacities to improve detection of epidemic prone infectious diseases, improve logistics for surveillance, standardize methods for antimicrobial resistance susceptibility testing and improve collaboration between disease surveillance officers and laboratory scientists.

However, availability of human and material resources to address these gaps has been difficult because of funding limitations. Bacterial isolation and identification from clinical specimens such as blood, stool and urine involve the use of sophisticated devices and specialized skills which are expensive [8]. Interventions from partner organizations and non-governmental organizations (NGOs) and/or corporate institutions are needed in the quest to address global health security challenges especially in resource poor settings such as Ghana.

As a way of addressing these challenges, Ghana received funding as one of the high-risk non-Ebola affected countries to strengthen the public health infrastructures and improve detection of epidemic prone infectious pathogens such as Salmonella, Shigella, Vibrio and diarrheagenic E. coli. The U. S Centers of Disease Control and Prevention engaged the Centre for Health Systems Strengthening (CfHSS) through the Association of Public Health Laboratories (APHL) to design and implement strategies for strengthening Public Health Laboratories (PHLs) in Ghana. This report presents a series of activities leading to improved detection of bacterial pathogens in the laboratory.

The role microbiological laboratories play in the detection and surveillance of pathogenic bacteria is important in addressing the global health security threats posed by infectious agents. Resourcing of the PHLs with equipment, reagents and human resource capacity building have enabled the increased and accurate detection of bacterial pathogens from clinical specimens of blood, stool, urine and cerebro-spinal fluid at three different PHLs in Ghana. It is instructive to note that prior to this programme, TPHL and KPHL for instance had stopped blood cultures due to unavailability of logistics and non-functional BACTEC equipment. SPHL as well was not well-versed in the use of CLSI standards for isolation of bacteria. Adequate measures including provision of distilled water plants, establishments of sheep farms, training in microbiological media preparation and identification of microbial pathogens were put in place. Training and adequate resourcing are therefore essential in strengthening the capacity and functional role of PHLs in developing countries.

In sub-Saharan Africa (sSA), about 12 million people die each year [12], with the causes of deaths largely due to undiagnosed infectious diseases such as HIV, malaria and tuberculosis [13]. A study in Kenya found that bacterial bloodstream infection diagnosed only by blood culture accounted for 26% of deaths among children [14], which gives credence to the fact that laboratory diagnosis of bacterial infections needs to be strengthened in sub-Sahara Africa. In line with previous studies in Africa [15, 16], Gram-negative bacteria predominated in our study. Infections from Gram-negative bacteria pose significant public health problems and this is mainly due to high resistance to antimicrobial agents [17]. Prior to implementation of the project, all three PHLs had serious challenges with regards to microbiological detection of bacterial pathogens from clinical specimens. At TPHL, identification was solely done by observing morphological characteristics of the colonies which grew on various culture media. There was absence of biochemical testing and automated blood culture system due to logistical and technical constraints. Both KPHL and SPHL performed very limited biochemical tests, and only SPHL performed automated blood culture analysis. Across the three PHLs, identification of suspected bacterial pathogens was performed up to the Genus level. All these hindered the identification process and proper administration of antimicrobials, which might directly or indirectly contribute to increasing rate of antimicrobial resistance globally. Procurement and maintenance of automated blood culture machines for the PHLs and training of staffs on its use resulted in an increased rate of blood samples received for blood culture. Reagents necessary for biochemical tests were also procured for all the PHLs and staffs were trained on the use and interpretation of tests such as triple sugar iron (TSI), citrate, indole, catalase, urease, coagulase and catalase tests. Analytical profile index and serotyping were also introduced to all PHLs which served as confirmatory tests for identification of some microbial pathogens. All these led to a significant upsurge in bacterial detection (Table 3) which hitherto could have easily been overlooked and missed.

Staphylococcus aureus was the most prevalent bacteria found in the blood. Naber reported that S. aureus is a major cause of bacteremia, and it is associated with higher morbidity and mortality, compared with bacteremia caused by other pathogens [18]. Again, life-threatening complications from S. aureus bloodstream infections such as infective endocarditis and metastatic infections could occur [19, 20], and these complications place high resource burden on health-care systems [21]. Without quality laboratory testing, detection of this medically relevant organism could not be done, and this would have a devastating clinical outcome on the patients. However, in this crisis time, laboratory and healthcare infrastructures are woefully inadequate to meet the pressing needs and/or perhaps have been ignored in several areas of sSA [22]. More often than not, a lot of financial resources from funding organizations are channeled to prevention of diseases and patients’ care, whereas building of laboratory capacity receives relatively little financial support [23].

E. coli and V. parahaemolyticus were commonly isolated pathogens from urine and stool cultures, respectively. Other common pathogens detected in urine included Klebsiella spp., Citrobacter spp., Staphylococcus saprophyticus and Staphylococcus aureus. UTI is among the leading causes of morbidity and mortality and inappropriate diagnosis could lead to treatment failures and additional complications [24]. This study showed that females were more likely to contract UTI than males, consistent with findings from rural Nigeria [25] and other parts of the world [26, 27]. In women, UTIs are one of the most frequent clinical bacterial infections, constituting about 25% of all infections [28]. This is mainly due to women having extremely short urethra, very close to the anal region, where several enterobacteria are shed frequently in stool. Other factors thought to predispose women to recurrent UTIs include voiding patterns pre- and post-coitus, wiping techniques, wearing tight undergarments, and vaginal douching; however, there is no proven association [29]. Also, medical conditions such as pregnancy, diabetes mellitus and immunosuppression increase risk of women having UTI [30]. Cases of V. parahaemolyticus infection are few globally; however, it is a common cause of bacterial gastroenteritis in Asia, especially in Japan [31]. Transmission of V. parahaemolyticus is mainly through the consumption of infected seafood causing acute gastroenteritis [32]. The organism can also make its way into an open wound during exposure to salt water [31]. All V. parahaemolyticus cases recorded in this study were from SPHL; located in the only coastal and southernmost region in this study where consumption of raw/undercooked seafoods is high. SPHL received the most specimens, because of its strategic location within the enclave of the South-Western part of Ghana. As a result, clinicians could easily refer patients to the laboratory for culture analysis. Inhabitants living along the coast principally engage in numerous outdoor activities such as fish farming and swimming in the deep ocean which predispose them to some waterborne infections, especially urinary tract infections (UTIs), and this could also account for the high proportions of urine samples collected in SPHL compared to the other PHLs.

Most detections of S. pneumoniae, H. influenzae and N. meningitidis in this study were achieved by multiplex real-time PCR. This platform was however only available at the TPHL, which also doubles as a reference testing centre for meningitis in Ghana. It is possible the other laboratories had low detections because of the use of only culture methods which is less sensitive as compared to multiplex PCR. PCR is a fast, sensitive and reliable technique for simultaneous detection of different molecular targets in one reaction. Training and logistical support provided by the CDC to the TPHL enabled them to utilize this molecular approach to detect common etiological agents of meningitis from CSF in Ghana. It would be important for the other PHLs to be adequately resourced with molecular testing capacities to help them to appropriately detect and respond to infectious agents and outbreaks.

The high number of meningitis pathogens identified at the TPHL could also be due to the geographical location and catchment populations targeted which lies within the sSA meningitis belt. Cases of meningitis are frequently reported from the Northern part of Ghana, where TPHL is situated, especially during the hot dry seasons (December – March, and August), leading to several meningitis outbreaks. Unlike CSF samples, other specimens such as urine, stool and blood did not show such seasonal trend. It is an established fact that cerebrospinal meningitis is an infectious disease which is commonly impacted by climate, specifically hot climate [10]. The five Northern regions (Upper East, Upper West, Savannah, North-East and Northern regions) are the most hottest regions in Ghana and the weather worsens during the period between December and June, and this results in a number of CSM outbreaks in Northern Ghana, as previously reported [10, 33]. Several studies have indicated that climatic conditions characterized by dry winds, dust storms, low humidity and cold nights considerably diminish the local immunity of the pharynx thereby increasing the risk of meningitis [34‐36]. These climatic conditions are typically found in the Northern Ghana during the dry season, and this likely explains why CSF samples were largely collected during the peak dry seasons compared to the other sample types.

Almost all CSF pathogens (145/148; 98%) recorded in this study emanated from TPHL. Streptococcus pneumoniae was the most isolated pathogen from CSF, consistent with previous data from Brong Ahafo region [37], about 210 km away from Tamale. This Global Health Security pathogen is the leading cause of bacterial meningitis, with an average mortality rate of 25%, despite effective antibiotic therapy and improved intensive care facilities [38]. Bacterial meningitis outbreaks are common in countries located in the Africa’s meningitis belt [39]. Rapid detection of the etiology of these outbreaks can lead to targeted public health interventions. Building and sustaining laboratory capacity in countries where meningitis outbreaks are common will be critical to ensure rapid and effective response to these outbreaks.

Global emergence and spread of antibiotic resistant strains of bacteria is still a major health problem. CfHSS provided training in the performance and interpretation of antimicrobial susceptibility tests to all the PHLs. Guidelines and protocols by Clinical and Laboratory Standards Institute (CLSI) were made available to the PHLs and all staff were adequately trained in the use of these documents. Antimicrobial susceptibility tests from blood cultures in this study revealed high resistance (35–55%) of S. aureus, K. pneumoniae and E. coli to gentamicin. This drug is commonly used to treat wide range of Gram-negative and some Gram-positive infections due to its antimicrobial efficacy, widespread availability and low cost [40]. However, the present report shows increase in resistance to this vital drug, consistent with data reported by Ababneh and colleagues [41]. Escherichia coli in urine showed high resistance to drugs such as ciprofloxacin (39.2%) and ampicillin (34%). These are historically useful antibiotics for the treatment of UTIs [42]. Also, high rate of Klebsiella spp. resistance (39.7%) to the 3rd generation cephalosporin cefotaxime is of great concern. Resistance to 3rd generation cephalosporins such as cefotaxime, ceftazidime and ceftriaxone serves as surrogate marker for detection of extended-spectrum beta-lactamases (ESBL) [43]. There have been reported cases of increasing trend of Klebsiella resistance to these 3rd generation cephalosporins [44, 45]. Third generation cephalosporin resistance leaves clinicians with limited options for treating patients with gram-negative infections, and as a result, relatively expensive drugs within the carbapenem class are usually considered the treatment of choice [45].

A limitation of this study was the inability to examine for the presence of ESBL phenotypes in the bacterial isolates which were resistant to the 3rd generation cephalosporins. A suggested approach would be to conduct double-disc diffusion synergy test for phenotypic confirmation of organisms possibly encoding ESBL. Another limitation was the exclusion of laboratory detection of viral and parasitic infections. This was due to restricted CDC-Ghana PHL budget for the project. Further support is therefore needed for capacity building in the detection of viruses and parasites in the three zonal PHLs.

Isolation and proper identification of aetiological agents in bacterial infection are of great importance. Partner and NGOs are encouraged to contribute their quota to help strengthen PHLs in sub-Saharan Africa. Outcome of this report clearly indicates that with routine and effective laboratory trainings, equipment and reagents support, detection of bacteria pathogens could be greatly enhanced, and the right antimicrobial therapy administered.