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Abstract We here describe a monospecific assemblage of giant aquatic snakes from the middle Eocene of Kpogamé, Togo. The material, consisting of large isolated vertebrae, is referred to Palaeophis africanus, an enigmatic palaeophiid species, which was so far otherwise known only from a limited number of vertebrae from the middle Eocene of Nigeria and Angola. Material from the late Eocene of the eastern USA that had been referred to the same species, is here instead considered too fragmentary for species-level determination and Palaeophis africanus is thus so far restricted to Africa. With the aid of micro-CT scanning, we present 3D models of 17 vertebrae, pertaining to different portions of the vertebral column. We provide detailed comparisons of the new material with all named African species of the genus Palaeophis. A tentative diagnosis of Palaeophis africanus is provided. With more than 50 vertebrae, the new Togolese specimens represent the most abundant known material attributed to Palaeophis africanus and significantly enhance our knowledge of the vertebral anatomy and intracolumnar variation for this taxon. Furthermore, this adds to the, as yet, extremely scarce fossil record of squamates from central western Africa, a region where Paleogene herpetofaunas are only rather poorly known.

Background: The 2018 cholera outbreak in Nigeria affected over half of the states in the country, and was characterised by high attack and case fatality rates. The country continues to record cholera cases and related deaths to date. However, there is a dearth of evidence on context-specific drivers and their operational mechanisms in mediating recurrent cholera transmission in Nigeria. This study therefore aimed to fill this important research gap, with a view to informing the design and implementation of appropriate preventive and control measures., Methods: Four bibliographic literature sources (CINAHL (Plus with full text), Web of Science, Google Scholar and PubMed), and one journal (African Journals Online) were searched to retrieve documents relating to cholera transmission in Nigeria. Titles and abstracts of the identified documents were screened according to a predefined study protocol. Data extraction and bibliometric analysis of all eligible documents were conducted, which was followed by thematic and systematic analyses., Results: Forty-five documents met the inclusion criteria and were included in the final analysis. The majority of the documents were peer-reviewed journal articles (89%) and conducted predominantly in the context of cholera epidemics (64%). The narrative analysis indicates that social, biological, environmental and climatic, health systems, and a combination of two or more factors appear to drive cholera transmission in Nigeria. Regarding operational dynamics, a substantial number of the identified drivers appear to be functionally interdependent of each other., Conclusion: The drivers of recurring cholera transmission in Nigeria are diverse but functionally interdependent; thus, underlining the importance of adopting a multi-sectoral approach for cholera prevention and control.

Background: Buruli ulcer (BU), a necrotizing skin infection caused by Mycobacterium ulcerans is the third most important mycobacterial disease globally after tuberculosis and leprosy in immune competent individuals. This study reports on the retrospective analyses of microbiologically confirmed Buruli ulcer (BU) cases in seventy-five health facilities in Ghana., Method/principal Findings: Pathological samples were collected from BU lesions and transported either through courier services or by car directly to the laboratory. Samples were processed and analysed by IS2404 PCR, culture and Ziehl-Neelsen staining for detection of acid-fast bacilli. From 2008 to 2016, we analysed by PCR, 2,287 samples of 2,203 cases from seventy-five health facilities in seven regions of Ghana (Ashanti, Brong Ahafo, Central, Eastern, Greater Accra, Northern and Volta). The mean annual positivity rate was 46.2% and ranged between 14.6% and 76.2%. The yearly positivity rates from 2008 to 2016 were 52.3%, 76.2%, 56.7%, 53.8%, 41.2%, 41.5%, 22.9%, 28.5% and 14.6% respectively. Of the 1,020 confirmed cases, the ratio of female to male was 518 and 502 respectively. Patients who were 15 years of age and below accounted for 39.8% of all cases. The median age was 20 years (IQR = 10-43). Ulcerative lesions were 69.2%, nodule (9.6%), plaque (2.9%), oedema (2.5%), osteomyelitis (1.1%), ulcer/oedema (9.5%) and ulcer/plaque (5.2%). Lesions frequently occurred on the lower limbs (57%) followed by the upper limbs (38%), the neck and head (3%) and the least found on the abdomen (2%)., Conclusions/significance: Our findings show a decline in microbiological confirmed rates over the years and therefore call for intensive education on case recognition to prevent over-diagnosis as BU cases decline., Competing Interests: The authors have declared that no competing interests exist.

Buruli ulcer (BU), caused by Mycobacterium ulcerans is a chronic necrotizing skin disease. It usually starts with a subcutaneous nodule or plaque containing large clusters of extracellular acid-fast bacilli. Surrounding tissue is destroyed by the cytotoxic macrolide toxin mycolactone produced by microcolonies of M. ulcerans. Skin covering the destroyed subcutaneous fat and soft tissue may eventually break down leading to the formation of large ulcers that progress, if untreated, over months and years. Here we have analyzed the bacterial flora of BU lesions of three different groups of patients before, during and after daily treatment with streptomycin and rifampicin for eight weeks (SR8) and determined drug resistance of the bacteria isolated from the lesions. Before SR8 treatment, more than 60% of the examined BU lesions were infected with other bacteria, with Staphylococcus aureus and Pseudomonas aeruginosa being the most prominent ones. During treatment, 65% of all lesions were still infected, mainly with P. aeruginosa. After completion of SR8 treatment, still more than 75% of lesions clinically suspected to be infected were microbiologically confirmed as infected, mainly with P. aeruginosa or Proteus miriabilis. Drug susceptibility tests revealed especially for S. aureus a high frequency of resistance to the first line drugs used in Ghana. Our results show that secondary infection of BU lesions is common. This could lead to delayed healing and should therefore be further investigated.

Background: Previous analyses of sera from a limited number of Ghanaian Buruli ulcer (BU) patients, their household contacts, individuals living in BU non-endemic regions as well as European controls have indicated that antibody responses to the M. ulcerans 18 kDa small heat shock protein (shsp) reflect exposure to this pathogen. Here, we have investigated to what extent inhabitants of regions in Ghana regarded as non-endemic for BU develop anti-18 kDa shsp antibody titers., Methodology/principal Findings: For this purpose we determined anti-18 kDa shsp IgG titers in sera collected from healthy inhabitants of the BU endemic Densu River Valley and the Volta Region, which was so far regarded as BU non-endemic. Significantly more sera from the Densu River Valley contained anti-18 kDa shsp IgG (32% versus 12%, respectively). However, some sera from the Volta Region also showed high titers. When interviewing these sero-responders, it was revealed that the person with the highest titer had a chronic wound, which was clinically diagnosed and laboratory reconfirmed as active BU. After identification of this BU index case, further BU cases were clinically diagnosed by the Volta Region local health authorities and laboratory reconfirmed. Interestingly, there was neither a difference in sero-prevalence nor in IS2404 PCR positivity of environmental samples between BU endemic and non-endemic communities located in the Densu River Valley., Conclusions: These data indicate that the intensity of exposure to M. ulcerans in endemic and non-endemic communities along the Densu River is comparable and that currently unknown host and/or pathogen factors may determine how frequently exposure is leading to clinical disease. While even high serum titers of anti-18 kDa shsp IgG do not indicate active disease, sero-epidemiological studies can be used to identify new BU endemic areas.

Mycobacterium ulcerans (M. ulcerans), the causative agent of the devastating skin disease Buruli ulcer (BU), is characterized by an extremely low level of genetic diversity. Recently, we have reported the first discrimination of closely related M. ulcerans variants in the BU endemic Densu River Valley of Ghana. In the study real-time PCR-based single nucleotide polymorphism (SNP) typing at 89 predefined loci revealed the presence of ten M. ulcerans haplotypes circulating in the BU endemic region. Here we describe the development of temperature-switch PCR (TSP) assays that allow distinguishing these haplotypes by conventional agarose gel-based analysis of the PCR products. After validation of the accuracy of typing results, the TSP assays were successfully established in a reference laboratory in Ghana. Development of the cost-effective and rapid TSP-based genetic fingerprinting method will thus allow investigating the spread of M. ulcerans clones by regular genetic monitoring in BU endemic countries.

The introduction of antibiotic therapy as first-line treatment of Buruli ulcer underlines the importance of laboratory confirmation of clinical diagnosis. Because smear microscopy has very limited sensitivity, the technically demanding and more expensive IS2404 diagnostic polymerase chain reaction (PCR) has become the main method for confirmation. By optimization of the release of mycobacteria from swab specimen and concentration of bacterial suspensions before smearing, we were able to improve the detection rate of acid-fast bacilli by microscopy after Ziehl-Neelsen staining. Compared with IS2404 PCR, which is the gold standard diagnostic method, the sensitivity and specificity of microscopy with 100 concentrated specimens were 58.4% and 95.7%, respectively. We subsequently evaluated a stepwise laboratory confirmation algorithm with detection of AFB as first-line method and IS2404 PCR performed only with those samples that were negative in microscopic analysis. This stepwise approach reduced unit cost by more than 50% to $5.41, and the total costs were reduced from $917 to $433.

For cultivation of Mycobacterium ulcerans from clinical specimens, we optimized the release of bacteria from swabs, as well as decontamination and cultivation on supplemented medium. Nevertheless, the proportions of positive cultures, 41.7% (5/12) for fine-needle-aspiration (FNA) samples and 43.8% (49/112) for swab samples, were lower than those we have previously observed for excised tissue specimens.