Fungi involved in rhinosinusitis in arid regions: insights from molecular identification and antifungal susceptibility.

Fungal rhinosinusitis (FRS) is a common problem worldwide, with an increasing burden in arid climate regions. Aspergillus species are the most common causative agents involved. In the present study, we investigated the prevalence, molecular characterization, and antifungal susceptibility of opportunists causing FRS in Sudan on the basis of strains collected over a period of 5 years. ß-Tubulin and calmodulin sequencing were used for species identification, and antifungal susceptibility profiles were evaluated by the protocol of the European Committee on Antimicrobial Susceptibility Testing (EUCAST). Predominant species belonged to the Aspergillus flavus complex (n = 244), A. terreus complex (n = 16), A. fumigatus complex (n = 7), and other fungi (n = 17). Molecular identification of 94 strains of Aspergillus revealed the following species: A. flavus (n = 88), A. terreus (n = 1), A. citrinoterreus (n = 2), A. fumigatus (n = 1), A. caespitosus (n = 1), and A. sydowii (n = 1). Several A. flavus and an A. fumigatus isolates showed reduced susceptibility to azoles (minimum inhibitory concentrations above the clinical breakpoints or epidemiological cutoff values). Despite several mutations revealed in cyp51A of these isolates, none could be directly linked to azole resistance. Molecular identification of fungi causing FRS is useful to identify cryptic species and for epidemiologic studies. IMPORTANCE Fungal rhinosinusitis (FRS) is a significant clinical problem in arid regions. This study provides new insights into the prevalence, etiology, and antifungal susceptibility of FRS pathogens in Sudan, where the disease burden is high. Aspergillus species, particularly the A. flavus complex, were identified as the primary FRS pathogens in the region, with some evidence of antifungal resistance. The molecular identification of fungal species causing FRS is useful for detecting antifungal resistance, identifying cryptic species, and characterizing the epidemiology of the disease. The emergence of Azole resistance Aspergilli in Sudan highlights the need for continued surveillance and appropriate use of antifungal agents. These findings have important implications for clinical management, public health policy, and future research on FRS. Publishing this study in Microbiology Spectrum would enable other researchers and clinicians to build on these findings, ultimately improving the diagnosis, treatment, and prevention of FRS.

Mycetoma: a unique neglected tropical disease.

Mycetoma can be caused by bacteria (actinomycetoma) or fungi (eumycetoma) and typically affects poor communities in remote areas. It is an infection of subcutaneous tissues resulting in mass and sinus formation and a discharge that contains grains. The lesion is usually on the foot but all parts of the body can be affected. The causative microorganisms probably enter the body by a thorn prick or other lesions of the skin. Mycetoma has a worldwide distribution but is restricted to specific climate zones. Microbiological diagnosis and characterisation of the exact organism causing mycetoma is difficult; no reliable serological test exists but molecular techniques to identify relevant antigens have shown promise. Actinomycetoma is treated with courses of antibiotics, which usually include co-trimoxazole and amikacin. Eumycetoma has no acceptable treatment at present; antifungals such as ketoconazole and itraconazole have been used but are unable to eradicate the fungus, need to be given for long periods, and are expensive. Amputations and recurrences in patients with eumycetoma are common.

Seventeen years of subcutaneous infection by Aspergillus flavus; eumycetoma confirmed by immunohistochemistry.

Chronic subcutaneous infections caused by Aspergillus species are considered to be extremely rare. Because these fungi are among the most common laboratory contaminants, their role as eumycetoma causative agents is difficult to ascertain. Here, we report the first case of A. flavus eumycetoma confirmed by isolation, molecular identification and immunohistochemical analysis. Patient was a 55-year-old male from Sudan suffering from eumycetoma on his left foot for a period of 17 years. He developed swelling, sinuses and white grain discharge was observed. He has been operated nine times and was treated with several regimens of ketoconazole and itraconazole without improvement. Initial diagnosis based on histology and radiology was Scedosporium eumycetoma. However, examination of the biopsy revealed A. flavus, which was identified by molecular analysis and MALDI-TOF MS. Immunohistochemistry using antibody directed against Aspergillus species was positive. Because of the earlier treatment failures with ketoconazole and itraconazole, therapy with voriconazole was initiated. However, in vitro susceptibility testing yielded a lower Minimum Inhibitory Concentration (MIC) value for itraconazole (0.25 µg ml(-1) ) than for voriconazole (1 µg ml(-1) ). Based on the presented results, A. flavus can be considered as one of the agents of white-grain eumycetoma.

Head and neck mycetoma: the mycetoma research centre experience.

Mycetoma is a unique neglected tropical disease which is endemic in what is known as the "mycetoma belt". The disease has many devastating impacts on patients and communities in endemic area and is characterised by massive deformity, destruction and disability. Mycetoma is commonly seen in the foot and hand and less frequent in other parts of the body. Mycetoma of the head and neck is a rarity and is associated with high morbidity and even mortality if not treated early. In this communication we report on 49 patients with head and neck mycetoma followed up at the Mycetoma Research Centre in Khartoum. Most of the reported patients had actinomycetoma and the majority were young adult males from mycetoma endemic areas in the Sudan. Most of them were students, farmers and workers. Prior to presentation the majority had long disease duration and the cause was multifactorial. Advanced disease with massive lesion, deformity and disability was the common presentation. There was no obvious history of local trauma, familial tendency or other predisposing factor identified in this group of patients. MRI and CT scan were the most accurate diagnostic tools to determine the disease extent. The treatment outcome was rather poor and characterised by low cure rate, poor outcome and high follows-up dropout. Such a gloomy outcome calls for structured and objective health education programs.

Mycetoma in the Sudan: an update from the Mycetoma Research Centre, University of Khartoum, Sudan.

This communication reports on the Mycetoma Research Centre of the University of Khartoum, Sudan experience on 6,792 patients seen during the period 1991-2014.The patients were predominately young (64% under 30 years old) males (76%). The majority (68%) were from the Sudan mycetoma belt and 28% were students. Madurella mycetomatis eumycetoma was the most common type (70%). In 66% of the patients the duration of the disease was less than five years, and 81% gave a history of sinuses discharging mostly black grains (78%). History of trauma at the mycetoma site was reported in 20%. Local pain was reported in 27% of the patients, and only 12% had a family history of mycetoma. The study showed that 57% of the patients had previous surgical excisions and recurrence, and only 4% received previous medical treatment for mycetoma. Other concomitant medical diseases were reported in 4% of the patients. The foot (76%) and hand (8%) were the most commonly affected sites. Less frequently affected sites were the leg and knee (7%), thigh (2%), buttock (2%) and arm and forearm (1%). Rare sites included the chest wall, head and neck, back, abdominal wall, perineum, oral cavity, tongue and eye. Multiple sites mycetoma was recorded in 135 (2%) of cases. At presentation, 37% of patients had massive lesions, 79% had sinuses, 8% had local hyper-hydrosis at the mycetoma lesion, 11% had regional lymphadenopathy, while 6% had dilated tortuous veins proximal to the mycetoma lesions. The diagnosis of mycetoma was established by combined imaging techniques and cytological, histopathological, serological tests and grain culture. Patients with actinomycetoma received a combination of antimicrobial agents, while eumycetoma patients received antifungal agents combined with various surgical excisions. Surgical excisions in the form of wide local excision, debridement or amputation were done in 807 patients, and of them 248 patients (30.7%) had postoperative recurrence. Different types of amputations were done in 120 patients (1.7%).

A new model for management of mycetoma in the Sudan.

Patients with mycetoma usually present late with advanced disease, which is attributed to lack of medical and health facilities in endemic areas, poor health education and low socio-economic status. With this background, an integrated patient management model at the village level was designed to address the various problems associated with mycetoma. The model was launched in an endemic village in the Sudan, between 2010 and 2013. This model is described in a prospective, descriptive, community-based study, aimed to collect epidemiological, ecological, and clinical data and to assess knowledge, attitude and practice (KAP) in order to design effective and efficient management measures. In this study, the prevalence of mycetoma was 14.5 per 1,000 inhabitants. The patients were farmers, housewives and children of low socio-economic status, and no obvious risk group was detected. All had surgery performed in a mobile surgical unit in the village which encouraged patients to present early with small early lesion leading to a good clinical outcome. The close contact with the Acacia tree thorns, animals and animal dung, walking bare footed and practising poor hygiene may all have contributed to the development of mycetoma in the village. Knowledge of mycetoma was poor in 96.3% of the study population, 70% had appropriate attitudes and beliefs towards interaction with mycetoma patients and treatment methods, and 49% used satisfactory or good practices in the management of mycetoma. Knowledge and practices on mycetoma were found to be significantly associated with age. Based on the KAP and epidemiological data, several health education sessions were conducted in the village for different target groups. The integrated management approach adopted in this study is unique and appeared successful and seems suitable as an immediate intervention. While for the longer term, establishment of local health facilities with trained health staff remains a priority.

Merits and pitfalls of currently used diagnostic tools in mycetoma.

Treatment of mycetoma depends on the causative organism and since many organisms, both actinomycetes (actinomycetoma) and fungi (eumycetoma), are capable of producing mycetoma, an accurate diagnosis is crucial. Currently, multiple diagnostic tools are used to determine the extent of infections and to identify the causative agents of mycetoma. These include various imaging, cytological, histopathological, serological, and culture techniques; phenotypic characterisation; and molecular diagnostics. In this review, we summarize these techniques and identify their merits and pitfalls in the identification of the causative agents of mycetoma and the extent of the disease. We also emphasize the fact that there is no ideal diagnostic tool available to identify the causative agents and that future research should focus on the development of new and reliable diagnostic tools.

The combination of amoxicillin-clavulanic acid and ketoconazole in the treatment of Madurella mycetomatis eumycetoma and Staphylococcus aureus co-infection.

Eumycetoma is a chronic progressive disabling and destructive inflammatory disease which is commonly caused by the fungus Madurella mycetomatis. It is characterized by the formation of multiple discharging sinuses. It is usually treated by antifungal agents but it is assumed that the therapeutic efficiency of these agents is reduced by the co-existence of Staphylococcus aureus co-infection developing in these sinuses. This prospective study was conducted to investigate the safety, efficacy and clinical outcome of combined antibiotic and antifungal therapy in eumycetoma patients with superimposed Staphylococcus aureus infection. The study enrolled 337 patients with confirmed M. mycetomatis eumycetoma and S. aureus co-infection. Patients were allocated into three groups; 142 patients received amoxicillin-clavulanic acid and ketoconazole, 93 patients received ciprofloxacin and ketoconazole and 102 patients received ketoconazole only. The study showed that, patients who received amoxicillin-clavulanic acid and ketoconazole treatment had an overall better clinical outcome compared to those who had combined ciprofloxacin and ketoconazole or to those who received ketoconazole only. In this study, 60.6% of the combined amoxicillin-clavulanic acid/ketoconazole group showed complete or partial clinical response to treatment compared to 30.1% in the ciprofloxacin/ketoconazole group and 36.3% in the ketoconazole only group. The study also showed that 64.5% of the patients in the ciprofloxacin/ketoconazole group and 59.8% in the ketoconazole only group had progressive disease and poor outcome. This study showed that the combination of amoxicillin-clavulanic acid and ketoconazole treatment is safe and offers good clinical outcome and it is therefore recommended to treat eumycetoma patients with Staphylococcus aureus co-infection.

New species of Madurella, causative agents of black-grain mycetoma.

A new species of nonsporulating fungus, isolated in a case of black-grain mycetoma in Sudan, is described as Madurella fahalii. The species is characterized by phenotypic and molecular criteria. Multigene phylogenies based on the ribosomal DNA (rDNA) internal transcribed spacer (ITS), the partial ß-tubulin gene (BT2), and the RNA polymerase II subunit 2 gene (RPB2) indicate that M. fahalii is closely related to Madurella mycetomatis and M. pseudomycetomatis; the latter name is validated according to the rules of botanical nomenclature. Madurella ikedae was found to be synonymous with M. mycetomatis. An isolate from Indonesia was found to be different from all known species based on multilocus analysis and is described as Madurella tropicana. Madurella is nested within the order Sordariales, with Chaetomium as its nearest neighbor. Madurella fahalii has a relatively low optimum growth temperature (30°C) and is less susceptible to the azoles than other Madurella species, with voriconazole and posaconazole MICs of 1 µg/ml, a ketoconazole MIC of 2 µg/ml, and an itraconazole MIC of >16 µg/ml. Since eumycetoma is still treated only with azoles, correct species identification is important for the optimal choice of antifungal therapy.

Blindness due to Acanthamoeba: first case report from Sudan.

Acanthamoeba is a free-living protozoan widely distributed in soil, water, and air. It can cause corneal infection (keratitis) in man, and may lead to blindness of the affected eye in 15% of untreated cases. The main risk factors are eye trauma and contact lens wearing. Early presentation of Acanthamoeba keratitis patients usually includes ocular pain, photophobia, and a unilateral red eye. The story of a female Sudanese housewife who suffered a blind left eye due to Acanthamoeba is described in this report.