The global epidemiology of emerging Histoplasma species in recent years.

Histoplasmosis is a serious infectious disease in humans caused by Histoplasma spp. (Onygenales), whose natural reservoirs are thought to be soil enriched with bird and bat guano. The true global burden of histoplasmosis is underestimated and frequently the pulmonary manifestations are misdiagnosed as tuberculosis. Molecular data on epidemiology of Histoplasma are still scarce, even though there is increasing recognition of histoplasmosis in recent years in areas distant from the traditional endemic regions in the Americas. We used multi-locus sequence data from protein coding loci (ADP-ribosylation factor, H antigen precursor, and delta-9 fatty acid desaturase), DNA barcoding (ITS1/2+5.8s), AFLP markers and mating type analysis to determine the genetic diversity, population structure and recognise the existence of different phylogenetic species among 436 isolates of Histoplasma obtained globally. Our study describes new phylogenetic species and the molecular characteristics of Histoplasma lineages causing outbreaks with a high number of severe outcomes in Northeast Brazil between 2011 and 2015. Genetic diversity levels provide evidence for recombination, common ancestry and clustering of Brazilian isolates at different geographic scales with the emergence of LAm C, a new genotype assigned to a separate population cluster in Northeast Brazil that exhibited low diversity indicative of isolation. The global survey revealed that the high genetic variability among Brazilian isolates along with the presence of divergent cryptic species and/or genotypes may support the hypothesis of Brazil being the center of dispersion of Histoplasma in South America, possibly with the contribution of migratory hosts such as birds and bats. Outside Brazil, the predominant species depends on the region. We confirm that histoplasmosis has significantly broadened its area of occurrence, an important feature of emerging pathogens. From a practical point of view, our data point to the emergence of histoplasmosis caused by a plethora of genotypes, and will enable epidemiological analysis focused on understanding the processes that lead to histoplasmosis. Further, the description of this diversity opens avenues for comparative genomic studies, which will allow progress toward a consensus taxonomy, improve understanding of the presence of hybrids in natural populations of medically relevant fungi, test reproductive barriers and to explore the significance of this variation.

Antifolates inhibit Cryptococcus biofilms and enhance susceptibility of planktonic cells to amphotericin B.

The Cryptococcus neoformans species complex contains the most important agents of fungal meningoencephalitis. Therapeutic choices are limited and issues related to toxicity and resistance to antifungals have been described. The present study evaluated the inhibitory effect of the antifolate combinations sulfamethoxazole-trimethoprim (SMX/TMP) and sulfadiazine-pyrimethamine (SDZ/PYR) against planktonic cells and biofilms of C. neoformans and C. gattii. The influence of the antifolate combinations on the amphotericin minimum inhibitory concentration (MIC) of planktonic cells was also investigated. In addition, the effect of these combinations on the cellular ergosterol content of planktonic cells was studied. Strains of C. neoformans (n = 15) and C. gattii (n = 15) obtained from environmental or clinical sources were evaluated by the broth microdilution method. SMX/TMP and SDZ/PYR showed antifungal activity against free living cells and sessile cells of Cryptococcus spp. Moreover, planktonic cells showed increased susceptibility to amphotericin B after pre-incubation with sub-inhibitory concentrations of SMX/TMP or SDZ/PYR. The drug combinations SMX/TMP and SDZ/PYR were able to prevent the biofilm formation and showed inhibitory effect against mature biofilms of both species. Additionally, the study showed that antifolate drugs reduced the ergosterol content in C. neoformans and C. gattii planktonic cells. Our results highlight the antifungal potential of antifolate drugs.

Rapid diagnosis of coccidioidomycosis by nested PCR assay of sputum.

Coccidioidomycosis is a deep infection caused by two dimorphic fungi, Coccidioides immitis and Coccidioides posadasii. Diagnosis of the disease requires culture of suspicious clinical samples on mycological media. However, as these species are virulent pathogens, handling of their cultures is a high-risk activity, and is limited to Biosafety Level 3 laboratories. This study describes the direct detection of C. posadasii DNA in an inappropriate sputum sample by PCR amplification of the highly specific Ag2/PRA antigen gene. The results obtained suggest that direct detection of the Ag2/PRA sequence in sputum is an excellent method for rapid and specific diagnosis of coccidioidomycosis.