Advocating for Coccidioidomycosis to Be a Reportable Disease Nationwide in the United States and Encouraging Disease Surveillance across North and South America.

Coccidioidomycosis (Valley fever) has been a known health threat in the United States (US) since the 1930s, though not all states are currently required to report disease cases. Texas, one of the non-reporting states, is an example of where both historical and contemporary scientific evidence define the region as endemic, but we don't know disease incidence in the state. Mandating coccidioidomycosis as a reportable disease across more US states would increase disease awareness, improve clinical outcomes, and help antifungal drug and vaccine development. It would also increase our understanding of where the disease is endemic and the relationships between environmental conditions and disease cases. This is true for other nations in North and South America that are also likely endemic for coccidioidomycosis, especially Mexico. This commentary advocates for US state and territory epidemiologists to define coccidioidomycosis as a reportable disease and encourages disease surveillance in other endemic regions across North and South America in order to protect human health and reduce disease burden.

Single-tube, dual channel pentaplexing for the identification of Candida strains associated with human infection.

Invasive candidiasis is one of the most common nosocomial fungal infections worldwide. Delayed implementation of effective antifungal treatment caused by inefficient Candida diagnosis contributes to its notoriously high mortality rates. The availability of better Candida diagnostic tools would positively impact patient outcomes. Here, we report on the development of a single-tube, dual channel pentaplex molecular diagnostic assay based on Multiplex Probe Amplification (MPA) technology. It allows simultaneous identification of C. auris, C. glabrata and C. krusei, at species-level as well as of six additional albicans and non-albicans pathogenic Candida at genus level. The assay overcomes the one-channel one-biomarker limitation of qPCR-based assays. Assay specificities are conferred by unique biomarker probe pairs with characteristic melting temperatures; post-amplification melting curve analysis allows simple identification of the infectious agent. Alerting for the presence of C. auris, the well-characterised multi-drug resistant outbreak strain, will facilitate informed therapy decisions and aid antifungal stewardship. The MPA-Candida assay can also be coupled to a pan-Fungal assay when differentiation between fungal and bacterial infections might be desirable. Its multiplexing capacity, detection range, specificity and sensitivity suggest the potential use of this novel MPA-Candida assay in clinical diagnosis and in the control and management of hospital outbreaks.

Detección y genotipificación de Pneumocystis jirovecii en muestras de pacientes mexicanos VIH positivos y negativos.

INTRODUCTION: Pneumocystis jirovecii is an atypical fungus particularly detected in HIV-positive or transplant patients. OBJECTIVE: To detect and genotype Pneumocystis jirovecii in patient samples from two hospitals in Mexico City. METHOD: Eighty-nine respiratory tract samples, corresponding to 53 patients (30 HIV-positive and 23 HIV-negative) with respiratory symptoms and to 11 healthy individuals included as negative control, were processed. DNA was extracted from the ITS region and amplified by nested polymerase chain reaction from the internal transcribed spacer, with one fragment being obtained at each round (693 and 550 bp). Genotypes and their phylogenetic relationship were determined by sequencing the 550 bp fragment. RESULTS: Forty-eight samples from 30 HIV-positive patients were received from a single hospital, out of which 11 (36.6 %) were positive for Pneumocystis jirovecii. No sample was positive in HIV-negative patients or healthy subjects. The most frequently detected haplotypes were Eg and Em. CONCLUSIONS: The frequency of Pneumocystis jirovecii infection was high in the studied Mexican population. The most common genotype was different from those reported in other countries. It is necessary to address this health problem through early detection of this infection.

INTRODUCCIÓN: Pneumocystis jirovecii es un hongo atípico detectado particularmente en pacientes VIH-positivos o con trasplante. OBJETIVO: Detectar y genotipificar Pneumocystis jirovecii en muestras de pacientes de dos hospitales de la ciudad de México. MÉTODO: Fueron procesadas 89 muestras respiratorias, correspondientes a 53 pacientes (30 VIH positivos y 23 VIH negativos) con sintomatología respiratoria y 11 personas sanas incluidas como control negativo. El DNA fue extraído y amplificado por PCR anidada de la región del espaciador transcrito interno, obteniendo un fragmento en cada ronda (de 693 y 550 pb). Los genotipos y su relación filogenética fueron determinados por secuenciación del fragmento de 550 pb. RESULTADOS: Cuarenta y ocho muestras de 30 pacientes VIH-positivos provenían de un solo hospital, de las cuales 11 (36.6 %) fueron positivas a Pneumocystis jirovecii. Ninguna fue positiva en pacientes VIH-negativos o personas sanas. Los haplotipos detectados con mayor frecuencia fueron Eg y Em. CONCLUSIONES: La frecuencia de infección por Pneumocystis jirovecii fue alta en la población mexicana estudiada. El genotipo más frecuente fue diferente a los reportados en otros países. Es necesario encauzar este problema de salud hacia la detección temprana de esta infección.

Detección y genotipificación de Pneumocystis jirovecii en muestras de pacientes mexicanos VIH positivos y negativos / Detection and genotyping of Pneumocystis jirovecii in samples from HIV positive and negative Mexican patients

Resumen Introducción: Pneumocystis jirovecii es un hongo atípico detectado particularmente en pacientes VIH-positivos o con trasplante. Objetivo: Detectar y genotipificar Pneumocystis jirovecii en muestras de pacientes de dos hospitales de la ciudad de México. Método: Fueron procesadas 89 muestras respiratorias, correspondientes a 53 pacientes (30 VIH positivos y 23 VIH negativos) con sintomatología respiratoria y 11 personas sanas incluidas como control negativo. El DNA fue extraído y amplificado por PCR anidada de la región del espaciador transcrito interno, obteniendo un fragmento en cada ronda (de 693 y 550 pb). Los genotipos y su relación filogenética fueron determinados por secuenciación del fragmento de 550 pb. Resultados: Cuarenta y ocho muestras de 30 pacientes VIH-positivos provenían de un solo hospital, de las cuales 11 (36.6 %) fueron positivas a Pneumocystis jirovecii. Ninguna fue positiva en pacientes VIH-negativos o personas sanas. Los haplotipos detectados con mayor frecuencia fueron Eg y Em. Conclusiones: La frecuencia de infección por Pneumocystis jirovecii fue alta en la población mexicana estudiada. El genotipo más frecuente fue diferente a los reportados en otros países. Es necesario encauzar este problema de salud hacia la detección temprana de esta infección.

Abstract Introduction: Pneumocystis jirovecii is an atypical fungus particularly detected in HIV-positive or transplant patients. Objective: To detect and genotype Pneumocystis jirovecii in patient samples from two hospitals in Mexico City. Method: Eighty-nine respiratory tract samples, corresponding to 53 patients (30 HIV-positive and 23 HIV-negative) with respiratory symptoms and to 11 healthy individuals included as negative control, were processed. DNA was extracted from the ITS region and amplified by nested polymerase chain reaction from the internal transcribed spacer, with one fragment being obtained at each round (693 and 550 bp). Genotypes and their phylogenetic relationship were determined by sequencing the 550 bp fragment. Results: Forty-eight samples from 30 HIV-positive patients were received from a single hospital, out of which 11 (36.6 %) were positive for Pneumocystis jirovecii. No sample was positive in HIV-negative patients or healthy subjects. The most frequently detected haplotypes were Eg and Em. Conclusions: The frequency of Pneumocystis jirovecii infection was high in the studied Mexican population. The most common genotype was different from those reported in other countries. It is necessary to address this health problem through early detection of this infection.

The Gα subunit signals through the Ste50 protein during the mating pheromone response in the yeast Kluyveromyces lactis.

Yeast mating signal transduction pathways require a heterotrimeric G protein composed of Gα, Gß, and Gγ subunits connected to a mitogen-activated protein kinase (MAPK) module. While in Saccharomyces cerevisiae elimination of Gα induces constitutive activation of the mating pathway, in Kluyveromyces lactis it produces partial sterility, which indicates that K. lactis Gα (KlGα) is required to positively activate mating. We use physical interaction experiments to determine that KlGα interacts with the adaptor protein KlSte50p. The Ras association (RA) domain of KlSte50p favored interaction with the GDP-bound KlGα subunit, and when the KlGα protein is constitutively activated, the interaction drops significantly. Additionally, KlSte50p strongly associates with the MAPK kinase kinase (MAPKKK) KlSte11p through its sterile alpha motif (SAM) domain. Genetic experiments placed KlSte50p downstream of the G protein α subunit, indicating that KlGα may stimulate the mating pathway via KlSte50p. Fusion of KlSte50p to the KlGß subunit partially eliminated the requirement of KlGα for mating, indicating that one contribution of KlGα to the mating pathway is to facilitate plasma membrane anchoring of KlSte50p.

Protein kinases involved in mating and osmotic stress in the yeast Kluyveromyces lactis.

Systematic disruption of genes encoding kinases and mitogen-activated protein kinases (MAPKs) was performed in Kluyveromyces lactis haploid cells. The mutated strains were assayed by their capacity to mate and to respond to hyperosmotic stress. The K. lactis Ste11p (KlSte11p) MAPK kinase kinase (MAPKKK) was found to act in both mating and osmoresponse pathways while the scaffold KlSte5p and the MAPK KlFus3p appeared to be specific for mating. The p21-activated kinase KlSte20p and the kinase KlSte50p participated in both pathways. Protein association experiments showed interaction of KlSte50p and KlSte20p with Galpha and Gbeta, respectively, the G protein subunits involved in the mating pathway. Both KlSte50p and KlSte20p also showed interaction with KlSte11p. Disruption mutants of the K. lactis PBS2 (KlPBS2) and KlHOG1 genes of the canonical osmotic response pathway resulted in mutations sensitive to high salt and high sorbitol but dispensable for mating. Mutations that eliminate the MAPKK KlSte7p activity had a strong effect on mating and also showed sensitivity to osmotic stress. Finally, we found evidence of physical interaction between KlSte7p and KlHog1p, in addition to diminished Hog1p phosphorylation after a hyperosmotic shock in cells lacking KlSte7p. This study reveals novel roles for components of transduction systems in yeast.

Kluyveromyces lactis sexual pheromones. Gene structures and cellular responses to alpha-factor.

The Kluyveromyces lactis genes for sexual pheromones have been analyzed. The alpha-factor gene encodes a predicted polypeptide of 187 amino acid residues containing four tridecapeptide repeats (WSWITLRPGQPIF). A nucleotide blast search of the entire K. lactis genome sequence allowed the identification of the nonannotated putative a-pheromone gene that encodes a predicted protein of 33 residues containing one copy of the dodecapeptide a-factor (WIIPGFVWVPQC). The role of the K. lactis structural genes KlMFalpha1 and KlMFA1 in mating has been investigated by the construction of disruption mutations that totally eliminate gene functions. Mutants of both alleles showed sex-dependent sterility, indicating that these are single-copy genes and essential for mating. MATalpha, Klsst2 mutants, which, by analogy to Saccharomyces cerevisiae, are defective in Galpha-GTPase activity, showed increased sensitivity to synthetic alpha-factor and increased capacity to mate. Additionally, Klbar1 mutants (putatively defective in alpha-pheromone proteolysis) showed delay in mating but sensitivity to alpha-pheromone. From these results, it can be deduced that the K. lactis MATa cell produces the homolog of the S. cerevisiaealpha-pheromone, whereas the MATalpha cell produces the a-pheromone.

The pheromone response pathway of Kluyveromyces lactis.

The mating pheromone response pathway in Saccharomyces cerevisiae is one of the best understood signalling pathways in eukaryotes. Comparison of this system with pathways in other fungal species has generated surprises and insights. Cloning and targetted disruption of genes encoding components of the pheromone response pathway has allowed the attribution of specific functions to these signal transduction components. In this review we describe current knowledge of the Kluyveromyces lactis mating system, and compare it with the well-understood S. cerevisiae pathway, emphasizing the similarities and differences in the heterotrimeric G protein activity. This mating pathway is controlled positively by both the Galpha and the Gbeta subunits of the heterotrimeric G protein.

Hallazgo de Candida albicans en manos de manejadores de alimentos / Candida albican findings in hands of people handling food

Las levaduras del género Candida frecuentemente son el agente causal de infecciones en sitios corporales expuestos a factores como la humedad y la maceración. En este estudio fueron incluidos 1,059 individuos manipuladores de alimentos de diversos centros comerciales de la ciudad de México y un grupo testigo sano de 139 personas. En todos ellos se tomaron muestras de manos para el aislamiento de hongos potencialmente patógenos. Se encontró que 30.7 por ciento de los trabajadores estudiados son portadores de diversas levaduras; la mayoría de estos aislamientos se obtuvo de los trabajadores del área de panadería. De los 325 aislamientos levaduriformes obtenidos, C. albicans representó 12.4 por ciento y el restante 85.8 por ciento correspondió a Candida no albicans; otras levaduras aisladas fueron Rhodotorula (0.9 por ciento) y Trichosporon (0.9 por ciento). En el grupo testigo solamente se aislaron levaduras de Candida en 11.5 por ciento; de éstas, seis correspondieron a C. albicans (37.5 por ciento) y 10 (62.5 por ciento) a C. no albicans. No se aislaron hongos filamentosos.

Aportaciones al estudio epidemiológico de las dermatofitosis / Epidemiological study of dermaphytoses in Mexico City

Con la finalidad de conocer nuevos datos sobre la frecuencia, agentes etiológicos y localización topográfica de las dermatofitosis, se realizó un estudio en 1734 pacientes que presentaban lesiones sugestivas de estas micosis. De cada paciente se anotaron los datos de localización corporal de la lesión, edad y género. Se tomaron muestras de escamas y/o pelos de las lesiones para realizar examen directo con hidróxido de potasio y cultivo en agar Sabouraud. La frecuencia de la infección por género no presentó diferencia significativa. De los pacientes estudiados, 869 fueron positivos para dermatófitos (50.1 por ciento) y en 446 el diagnóstico se confirmó tanto por examen directo como por cultivo. Las uñas fueron el sitio más afectado (579 casos) y en la mayoría de los cultivos fue aislado Trichophyton rubrum (82.5 por ciento). Del total de pacientes positivos a dermatófitos, 33 fueron niños; 11 de ellos presentaron afectación de uñas por Trichophyton rubrum. Se diagnosticaron tres pacientes con onicomicosis en pies por Microsporum canis. Se discute la importancia de algunas variantes epidemiológicas en las dermatofitosis.