Bacteriology Unit

One ​​of the CPC's main missions is to contribute to strengthening disease surveillance within the Ministry of Health and the Ministry of Public Health (MINEPIA) through two actions: supporting health monitoring operations and analyzing surveillance data.

This mission is achieved through the CPC's participation in major national and international disease control programs, particularly those with epidemic risks for Cameroon and the Central African subregion.

Although it serves as a national laboratory for various diseases, the CPC's expertise often gives it a subregional dimension. It therefore receives and analyzes samples from other countries. All its analyses are subject to external quality control in accordance with WHO recommendations. The databases are shared weekly with the Ministry of Health and all other organizations involved. By Ministerial Note E2107/L/MINSANTE/SG/DPML/SDLTS/SL of 18/08/2015 designating the official list of reference laboratories for the various diseases under epidemiological surveillance, the CPC was designated as the reference laboratory for 28 diseases. Surveillance was extended to viruses related to the smallpox virus.

Malaria

The Malaria Research Service of the Centre Pasteur du Cameroun aims to improve knowledge of malaria transmission in Cameroon in order to develop more effective control strategies. Thus, for several years, basic and applied research activities on the pandemic have been conducted. The goal is not only to better understand the dynamics of disease transmission, but also to evaluate the impact of the control measures implemented. This approach will help improve national policy on disease control. The topics of interest addressed include the surveillance of antimalarial drug resistance, studies on pathogenicity and the mechanisms of the host's immune response to the parasite, the identification of new vaccine candidates and antimalarial molecules, and the development of new diagnostic tests. Researchers in this Department conduct research projects in collaboration with numerous national and international partners, and actively participate in strengthening local capacities through training courses and student supervision. This Department is recognized as a National Reference Laboratory for the diagnosis and monitoring of antimalarial drug resistance in Cameroon. It houses a large collection of blood samples, representing the different clinical forms of malaria that can be found in Cameroon.

Main thematic areas

The Department comprises three major research teams with the following thematic areas:

1-Molecular epidemiology and pathogenicity of malaria (Team leader, Prof. EBOUMBOU Else Carole): The team conducts studies on the distribution and dynamics of malaria transmission in Cameroon, and on the identification of associated risk factors. In addition, it is responsible for monitoring drug resistance and confirming the diagnosis of malaria infections.

2-Vector-parasite interaction (Team leader, Prof. NSANGO Sandrine Evéline): This team's activities focus on studying the mosquito's immune response following Plasmodium infection. It is also involved in the identification and development of vaccines that block transmission.

3-Applied research (Team leader, Dr. AYONG Lawrence): The team is involved in the development of new, inexpensive diagnostic tests for malaria. It is also working on the identification of new antimalarial molecules in collaboration with numerous national and international research institutes.

Tuberculosis and Mycobacterial Infections

Evaluation of new diagnostic methods for intrathoracic tuberculosis in children in three sub-Saharan African cities: Abidjan (Côte d’Ivoire), Yaoundé (Cameroon), and Antananarivo (Madagascar) – TB Kids Project (Dr. Eyangoh / Dr. Tejiokem)

Objectives

To identify optimal algorithms for the diagnosis of intrathoracic TB in children according to different environments and levels of healthcare resources; to evaluate the new diagnostic tool Xpert MTB/RIF and alternative bacteriological sampling methods (nasopharyngeal aspiration, stool samples) for pediatric intrathoracic TB diagnosis; to identify the determinants of false positives and false negatives for each diagnostic tool; to assess the performance of the pediatric scoring system used by pediatricians.

Progress Status

- Enrollment began in October 2016 at the Mother and Child Center of the Chantal Biya Foundation. The Essos Hospital Center began five months later due to delays in preparing the partnership agreements.

- By the end of 2016, 26 children had been enrolled in the study, representing 30% of the expected number for the period. Hence, it became necessary to start the study at the CHE site and to raise awareness among staff at study sites and pediatricians consulting in other healthcare facilities in Yaoundé.

Study of the epidemiological, entomological, and immuno-genetic aspects related to the transmission mode of Mycobacterium ulcerans to humans (Dr. Eyangoh)

Main National and International Collaborations: Epidemiology and Public Health Laboratory of the CPC, hospitals in endemic sites, University of Angers (Inserm Avenir ATOMycA team), Emerging Diseases Epidemiology Unit, Institut Pasteur, Paris, IRD Cameroon, and IRD Montpellier (UMR MIVEGEC).

Role of Sara EYANGOH: Southern Coordinator for ANR, Principal Investigator for Sanofi Fondation Espoir, and Head of the JEAI team.

All funding ended in 2015. During 2016, we continued to enhance the value of samples collected during the large-scale sampling campaign for the characterization of indoor arthropod fauna, in order to create a reference collection for the scientific community. Student Solange Meyin, who defended her PhD thesis on this project in December 2015, is continuing with a postdoctoral fellowship until March 2017 to conduct a SWOT analysis of 10 years of activities and assist in securing funding for future work.

A total of 309,510 arthropod specimens belonging to 213 families were collected from pre-selected houses in the endemic site of Akonolinga. Detection of the molecular signature of Mycobacterium ulcerans showed a positivity rate of 7.30%. Two manuscripts, presenting respectively the diversity of arthropods and the detection of Mycobacterium ulcerans, are in progress.

Evaluation of Rapid Diagnostic Tools for Tuberculosis

The mycobacteriology service conducts research projects on the implementation of the most recent diagnostic algorithms. Thanks to funding from Expand-TB UNITAID (Expanding Access for New Diagnostic Tools) and OFID/RIIP (OPEC Fund for International Development), the CPC has a technical platform equipped with all WHO-approved diagnostic tools for tuberculosis, facilitating surveillance of multidrug-resistant TB: microscopy, liquid culture (MGIT), solid culture, molecular identification and resistance tests (GenXpert, Hain), spoligotyping, and RFLP. Cultures are performed under a microbiological safety cabinet within the CPC’s BSL-3 laboratory. This expertise is used to evaluate diagnostic tools and propose to the National Tuberculosis Control Program diagnostic algorithms adapted to different patient types.

Thanks to the EXPAND-TB project, a new database developed on the Ubuntu web application called “EXTBCAM,” based on the standardized examination request form, was implemented to enable rapid reporting of detected multidrug-resistant patients. A new algorithm was adopted for retreatment patients suspected of carrying MDR strains.

Tuberculosis Quality Assurance and Training Programs

Surveillance of multidrug-resistant tuberculosis and microscopy quality control: The CPC, as the National Reference Laboratory for tuberculosis, coordinates the laboratory network for TB diagnosis and multidrug resistance surveillance. Quality control of the microscopy laboratory network is ensured through blind re-reading of slides and supervision missions.

Annual report: Two supervision missions were conducted in each of the 36 Tuberculosis Diagnostic and Treatment Centers in the Central Region (a total of 72 supervisions), and one mission in each of the East, South, West, and North regions. Approximately 210 slides were rechecked. A total of 468 drug susceptibility tests were carried out, including 109 MDR-TB cases reported for treatment. The data file for rifampicin-resistant tuberculosis detected by GeneXpert MTB/RIF was standardized for rapid notification to the NTP upon detection. 2,652 GeneXpert analyses were performed, 765 were positive, including 84 resistant to rifampicin and reported.

Surveillance of Multidrug-Resistant Tuberculosis

The National Reference Laboratory for tuberculosis coordinates the laboratory network and resistance surveillance. In 2012, 523 drug susceptibility tests were conducted on retreatment patients suspected of carrying MDR strains (relapse, treatment failure, or return after default). Among these, 221 strains showed resistance to at least one antibiotic, including 182 MDR (resistant to both Isoniazid and Rifampicin).

No extensively drug-resistant (XDR) strains—MDR strains resistant to a fluoroquinolone and at least one of the three second-line injectable drugs (amikacin, kanamycin, or capreomycin)—were isolated, although five patients already showed fluoroquinolone resistance.

Hepatitis Viruses and Other Oncogenic Viruses

Liver Cancer in Equatorial Africa: Characterization of New Serum Biomarkers of Primary Liver Cancer in Patients in Equatorial Africa (Prof. Njouom)

Objective: This study aims to identify its infectious and non-infectious etiological factors; to evaluate the influence of different viral genotypes and variants on its development; and to describe the somatic genetics of liver tumors.

Progress Status

To date, 88 cases of hepatocellular carcinoma (HCC), 85 control group 1, and 140 control group 2 (100 for HBV and 40 for HCV) have been included in this study. Serological analyses of HBV and HCV among cases and controls revealed that both viruses are strongly associated with the development of HCC in Cameroon, with respective prevalences of (64% and 26%) among HCC subjects versus (10% and 4.3%) among controls without HCC. Phylogenetic analyses of HBV revealed the circulation of two genotypes,

A and E, in our study population, with 35/42 genotype A (83.3%) and 7/42 genotype E (16.7%) among cases, compared to 13/44 genotype A (29.5%) and 31/44 genotype E (70.5%) among controls. Phylogenetic analyses of HCV revealed the circulation of three genotypes—1, 2, and 4—in our study population, with 7/17 genotype 4 (41.2%), 5/17 genotype 2 (29.4%), and 5/17 genotype 1 (29.4%) among cases, versus 2/8 genotype 1 (25%), 4/8 genotype 2 (50%), and 2/8 genotype 4 (25%) among controls. Evaluation of HDV serology among all patients positive for HBsAg showed that among 54 HCC cases tested, 21 (38.9%) were positive, while among healthy controls without cancer, only 1/6 (16.7%) tested positive for anti-HDV antibodies.

Mutations in the Pre-S and C genes have already been identified. The mutations C1766T, T1768A, and G1862T/A in the HBV C gene were found only in cases, whereas mutations C2875A, G2951C, and C3116T in the HBV pre-S fragment were significantly more frequent among cases compared to HBV-positive controls. These mutations may therefore be strongly involved in the development of HCC among HBV-infected patients.

Epidemiology of Viral Hepatitis B, C, and Delta in Cameroon: Analysis of Samples from the 2011 Demographic and Health Survey (DHS Hepatitis). ANRS 12289 Project (Prof. Njouom)

Objectives: Given the availability of data and the biobank from the 2011 Cameroon National Demographic and Health Survey (DHS), this project was initiated to study the seroprevalence, distribution, and factors associated with HBV, HCV, and HDV infections in a representative sample of the general population, and to determine the contribution of historical iatrogenic HCV transmission to the current epidemic in Cameroon.

Activities Conducted and Conclusions

Analyses yielded the following national prevalence rates: Hepatitis B Virus: 11.9% (95% CI: 11.0%–12.8%); Hepatitis C Virus: 1.03% (95% CI: 0.8%–1.6%); Hepatitis D Virus: 10.5% (95% CI: 8.7%–12.7%).

These results show that Cameroon may be considered a low-endemic area for hepatitis C and a high-endemic area for hepatitis B and D. Prevalence varies widely across the country, indicating distinct epidemiological patterns in certain administrative regions.

HBV prevalence differs significantly by age group: between 11–13% among individuals under 45 years, and between 8–9% among those over 45 (p<0.05). Rural populations are more affected than urban ones (13.3% vs. 10.7%, p<0.05). Prevalence also differs significantly by administrative region, with the Far North and North regions most affected (17.7% and 17.5%, respectively).

Among HBV-infected individuals, HDV prevalence also varies significantly by age, with those over 45 years being the most affected (16.8%). HDV prevalence also differs by region, with the East and South regions showing the highest rates (53.7% and 48.5%, respectively).

HCV prevalence increases significantly with age—from about 1% in those under 45, to over 3% among those aged 45–55, and about 7% among those over 55. A recent study in Ebolowa among individuals over 60 found a prevalence of about 50% in this population group (Pépin et al., 2010). Rural populations are more affected than urban ones (1.2% vs. 0.9%, p<0.05). Prevalence also varies by region, with the Centre region being the most affected (2.4%).

In summary, at-risk populations vary depending on the type of hepatitis virus: Hepatitis B among young people under 45 years (prevalence above 10%), and Hepatitis C among older adults over 55 years (prevalence around 7%).

Strategies to combat each of these viral hepatitis infections should consider these at-risk populations: Hepatitis B among the young, and Hepatitis C among the elderly. Observational study on the effectiveness of hepatitis B (HB) vaccination initiated at birth and follow-up of the expanded vaccination program including the EPI-HB component among children born to HBsAg-positive mothers in the Tokombéré district, Cameroon. ANRS 12303 (Prof. Richard Njouom)

Objectives:

To evaluate the effectiveness of “targeted prevention of mother-to-child transmission (PMTCT) of HBV,” initiated since 2009 in Tokombéré with GEMHEP support, involving systematic screening during pregnancy for HIV and HBsAg, followed by HB vaccination at birth for infants born to HBsAg-positive mothers (without anti-HBs immunoglobulin), and subsequent administration of the three EPI-HB doses at 6, 10, and 14 weeks.

The primary objective is to evaluate vaccination effectiveness in the target population by estimating the mother-to-child transmission rate among infants vaccinated at birth from HBsAg-positive mothers (HB birth vaccination failures).

The secondary objective is to study the risk factors associated with these failures (e.g., vaccination delay).

Project Progress Summary

Recruitment of mother-child pairs is ongoing in Tokombéré. Blood samples are collected and will be sent to CPC and the University Hospital of Angers, France, for HBV serological and molecular analyses. However, security concerns in the Far North of Cameroon have slowed recruitment.

Evaluation of the Performance of Three Rapid Diagnostic Tests for the Detection of Anti-Hepatitis C Virus Antibodies in Cameroon.

Acronym: (ANRS HEPATITE 12336) (Prof. Richard Njouom)

Research Objective

– To evaluate the performance of three rapid diagnostic tests (Multisure HCV Antibody Assay, First Response® HCV Card Test, and Toyo® Anti-HCV Test) for detecting anti-HCV antibodies in Cameroon.

– To determine sensitivity, specificity, predictive values (positive and negative), likelihood ratios (positive and negative), and the area under the ROC curve for these three RDTs using plasma samples.

– To determine the same performance indicators for the Multisure test using whole blood dried on filter paper (DBS).

Progress Status:

A retrospective cross-sectional study was conducted at the Centre Pasteur du Cameroun (CPC) over six months (July–December 2016). Using the ABBOTT Diagnostic Architect anti-HCV immunoassay as reference, a panel of 200 plasma samples (100 positive and 100 negative) was assembled from the CPC plasmathèque. The three RDTs (Multisure, First Response®, and Toyo®) were evaluated on plasma. DBS were reconstituted from 200 whole blood samples (100 negative and 100 positive) from patients who came to CPC for HCV serology. Only the Multisure test was evaluated on DBS. Test performance was expressed as sensitivity (Se), specificity (Sp), positive predictive value (PPV), negative predictive value (NPV), positive likelihood ratio (LR+), negative likelihood ratio (LR–), and area under the ROC curve (AUC).

Evaluation on plasma yielded: Se (99%, 96%, 96%), Sp (83%, 90%, 78%), PPV (85.3%, 96%, 89.7%), NPV (98.8%, 90%, 90.7%), LR+ (5.82, 9.6, 4.3), LR– (0.01, 0.04, 0.05), and AUC (0.96, 0.93, 0.88) for Multisure, First Response®, and Toyo®, respectively. On DBS, Multisure yielded: Se (71%), Sp (100%), PPV (100%), NPV (78.7%), LR+ (∞), LR– (0.27), and AUC (0.68).

Diagnosis and Monitoring of Viral Hepatitis

Epidemiology of Viral Hepatitis B, C, and Delta in Cameroon: Analysis of Samples from the 2011 Demographic and Health Survey (DHS) ANRS 12289-Hepatitis C (Prof. Njouom)

Objectives: Using data and biobank resources from the 2011 Cameroon DHS, we initiated this project to study seroprevalence, distribution, and factors associated with HBV, HCV, and HDV infections in a representative population sample and to determine the contribution of historical iatrogenic HCV transmission to the current epidemic.

Progress: All biological analyses (HIV, HCV, HBV, and HDV serology) were completed on over 14,000 samples collected during DHS-4. Statistical analysis is ongoing.

Diagnosis and Management of Chronic Viral Hepatitis

The high prevalence of chronic HBV and HCV infection in Northern Cameroon makes patient management a daily challenge for clinicians. While diagnostic tests for viral carriage are accessible, access to tests confirming viral replication, genotype, and disease stage is limited. These essential tests for appropriate therapy are expensive and must be prescribed judiciously. Dr. Richard Njouom, Head of the Virology Service at the Centre Pasteur du Cameroun in Yaoundé, organized a meeting in Garoua, Cameroon, on November 3, 2015, to address practical issues such as: How to diagnose chronic viral hepatitis? What assessments should follow diagnosis? How to initiate antiviral treatment? What biological follow-up should be offered to patients?

Prevalence and Genetic Diversity of Hepatitis Viruses

Prevalence and Genetic Diversity of Hepatitis E Virus in Porcine and Human Populations in Cameroon: Evaluation of interspecies transmission. This project, coordinated by Prof. Richard Njouom, is funded by the University of California, Los Angeles.

Epidemiology of Viral Hepatitis B, C, and Delta in Cameroon: Analysis of samples from the 2011 Demographic and Health Survey. ANRS Project.

This project is a collaboration with the Institut Pasteur (Prof. Arnaud Fontanet) and the Universities of Sherbrooke, Canada (Dr. J. Pépin) and Paris Diderot (G. Lachenal).

The demonstration of effective therapeutic protocols and the introduction of affordable generic or biosimilar drugs are expected to revolutionize hepatitis B and C management in developing countries. It is now urgent for the most affected countries to obtain reliable estimates of the number of infected individuals who will require treatment in the coming years.

Cameroon is one of the Central African countries most affected by chronic viral hepatitis, especially hepatitis C. The data and biobank from the 2011 Cameroon DHS provide a unique opportunity to study the seroprevalence, distribution, and factors associated with HBV, HCV, and HDV infections in a representative general population sample. The project also aims to identify factors associated with HCV infection and assess the contribution of historical iatrogenic transmission to the current epidemic.

HBV, HCV, and HDV serology as well as HBV DNA quantification will be performed on 14,700 dried blood spot (DBS) samples collected during the 2011 DHS. Historical iatrogenic transmission of HCV will be analyzed through geo-referenced DHS data and spatial correlation analyses between HCV infection and historical maps of medical interventions.

Molecular Diagnosis of Human Papillomavirus (HPV)

HPVs (Human Papillomaviruses) are viruses that can infect the skin and mucous membranes. There are more than 150 types of HPV, about 40 of which can infect male and female genital organs. Around 20 types can cause moderate to severe cellular abnormalities, precancerous lesions, and cancers. In collaboration with the Department of Anatomopathology, the Virology Service has implemented molecular diagnosis of HPV to confirm suspected cases detected by cervical-vaginal smears during cervical cancer screening. In 2012, 89 molecular diagnostic tests (by viral DNA detection on cervical smears) were performed.