The diagnosis of fungal neglected tropical diseases (fungal NTDs) and the role of investigation and laboratory tests: An expert consensus report

The diagnosis of fungal Neglected Tropical Diseases (NTD) is primarily based on initial visual recognition of a suspected case followed by confirmatory laboratory testing, which is often limited to specialized facilities. Although molecular and serodiagnostic tools have advanced, a substantial gap remains between the desirable and the practical in endemic settings. To explore this issue further, we conducted a survey of subject matter experts on the optimal diagnostic methods sufficient to initiate treatment in well-equipped versus basic healthcare settings, as well as optimal sampling methods, for three fungal NTDs: mycetoma, chromoblastomycosis, and sporotrichosis. A survey of 23 centres found consensus on the key role of semi-invasive sampling methods such as biopsy diagnosis as compared with swabs or impression smears, and on the importance of histopathology, direct microscopy, and culture for mycetoma and chromoblastomycosis confirmation in well-equipped laboratories. In basic healthcare settings, direct microscopy combined with clinical signs were reported to be the most useful diagnostic indicators to prompt referral for treatment. The survey identified that the diagnosis of sporotrichosis is the most problematic with poor sensitivity across the most widely available laboratory tests except fungal culture, highlighting the need to improve mycological diagnostic capacity and to develop innovative diagnostic solutions. Fungal microscopy and culture are now recognized as WHO essential diagnostic tests and better training in their application will help improve the situation. For mycetoma and sporotrichosis, in particular, advances in identifying specific marker antigens or genomic sequences may pave the way for new laboratory-based or point-of-care tests, although this is a formidable task given the large number of different organisms that can cause fungal NTDs

Review of influenza-associated pulmonary aspergillosis in ICU patients and proposal for a case definition: an expert opinion

Purpose: Invasive pulmonary aspergillosis is increasingly reported in patients with influenza admitted to the intensive care unit (ICU). Classification of patients with influenza-associated pulmonary aspergillosis (IAPA) using the current definitions for invasive fungal diseases has proven difficult, and our aim was to develop case definitions for IAPA that can facilitate clinical studies. Methods: A group of 29 international experts reviewed current insights into the epidemiology, diagnosis and management of IAPA and proposed a case definition of IAPA through a process of informal consensus. Results: Since IAPA may develop in a wide range of hosts, an entry criterion was proposed and not host factors. The entry criterion was defined as a patient requiring ICU admission for respiratory distress with a positive influenza test temporally related to ICU admission. In addition, proven IAPA required histological evidence of invasive septate hyphae and mycological evidence for Aspergillus. Probable IAPA required the detection of galactomannan or positive Aspergillus culture in bronchoalveolar lavage (BAL) or serum with pulmonary infiltrates or a positive culture in upper respiratory samples with bronchoscopic evidence for tracheobronchitis or cavitating pulmonary infiltrates of recent onset. The IAPA case definitions may be useful to classify patients with COVID-19-associated pulmonary aspergillosis (CAPA), while awaiting further studies that provide more insight into the interaction between Aspergillus and the SARS-CoV-2-infected lung. Conclusion: A consensus case definition of IAPA is proposed, which will facilitate research into the epidemiology, diagnosis and management of this emerging acute and severe Aspergillus disease, and may be of use to study CAPA

Cryptococcal meningitis: A neglected NTD?

Although HIV/AIDS has been anything but neglected over the last decade, opportunistic infections (OIs) are increasingly overlooked as large scale donors shift their focus from acute care to prevention and earlier antiretroviral treatment (ART) initiation. Of these OIs, cryptococcal meningitis, a deadly invasive fungal infection, continues to affect hundreds of thousands of HIV patients with advanced disease each year and is responsible for an estimated 15%-20% of all AIDS-related deaths [1,2]. Yet cryptococcal meningitis ranks amongst the most poorly funded “neglected” diseases in the world, receiving 0.2% of available relevant research and development (RandD) funding according to Policy Cures’ 2016 G-Finder Report [3,4]

A novel environmental azole resistance mutation in Aspergillus fumigatus and a possible role of sexual reproduction in its emergence

This study investigated the dynamics of Aspergillus fumigatus azoleresistant phenotypes in two compost heaps with contrasting azole exposures: azole free and azole exposed. After heat shock, to which sexual but not asexual spores are highly resistant, the azole-free compost yielded 98% (49/50) wild-type and 2% (1/50) azole-resistant isolates, whereas the azole-containing compost yielded 9% (4/45) wild-type and 91% (41/45) resistant isolates. From the latter compost, 80% (36/45) of the isolates contained the TR46/Y121F/T289A genotype, 2% (1/45) harbored the TR46/Y121F/M172I/T289A/G448S genotype, and 9% (4/45) had a novel pan-triazoleresistant mutation (TR46 3/Y121F/M172I/T289A/G448S) with a triple 46-bp promoter repeat. Subsequent screening of a representative set of clinical A. fumigatus isolates showed that the novel TR46 3 mutant was already present in samples from three Dutch medical centers collected since 2012. Furthermore, a second new resistance mutation was found in this set that harbored four TR46 repeats. Importantly, in the laboratory, we recovered the TR46 3 mutation from a sexual cross between two TR46 isolates from the same azole-containing compost, possibly through unequal crossing over between the double tandem repeats (TRs) during meiosis. This possible role of sexual reproduction in the emergence of the mutation was further implicated by the high level of genetic diversity of STR genotypes in the azole-containing compost. Our study confirms that azole resistance mutations continue to emerge in the environment and indicates compost containing azole residues as a possible hot spot. Better insight into the biology of environmental resistance selection is needed to retain the azole class for use in food production and treatment of Aspergillus diseases. IMPORTANCE Composting of organic matter containing azole residues might be important for resistance development and subsequent spread of resistance mutations in Aspergillus fumigatus. In this article, we show the dominance of azoleresistant A. fumigatus in azole-exposed compost and the discovery of a new resistance mutation with clinical relevance. Furthermore, our study indicates that current fungicide application is not sustainable as new resistance mutations continue to emerge, thereby threatening the use of triazoles in medicine. We provide evidence that the sexual part of the fungal life cycle may play a role in the emergence of resistance mutations because under laboratory conditions, we reconstructed the resistance mutation through sexual crossing of two azole-resistant A. fumigatus isolates derived from the same compost heap. Understanding the mechanisms of resistance selection in the environment is needed to design strategies against the accumulation of resistance mutations in order to retain the azole class for crop protection and treatment of Aspergillus diseases.</p

Prevalence of Histoplasmosis among Persons with Advanced HIV Disease, Nigeria

We sought to determine the prevalence of probable disseminated histoplasmosis among advanced HIV disease (AHD) patients in Nigeria. We conducted a cross-sectional study in 10 sites across 5 of 6 geopolitical zones in Nigeria. We identified patients with urinary samples containing CD4 cell counts &lt;200 cells/mm3 or World Health Organization stage 3 or 4 disease who also had &gt;2 clinical features of disseminated histoplasmosis, and we tested them for Histoplasma antigen using a Histoplasma enzyme immune assay. Of 988 participants we recruited, 76 (7.7%) were antigen-positive. The 76 Histoplasma antigen-positive participants had significantly lower (p = 0.03) CD4 counts; 9 (11.8%) were also co-infected with tuberculosis. Most antigen-positive participants (50/76; 65.8%; p = 0.015) had previously received antiretroviral treatment; 26/76 (34.2%) had not. Because histoplasmosis is often a hidden disease among AHD patients in Nigeria, Histoplasma antigen testing should be required in the AHD package of care.</p