PROGRAMA
DE ABSTRACTS:
Presentaciones
Orales
Carteles
Científicos
XVIII
Simposio Anual de Micología
22 de
abril de 2017
Conferencia Magistral Dr. Carlos Chardón Palacios
Medicinal fungus and the
course of treatment: The role of Ganoderma
lucidum in breast cancerDra. Yaliz Loperena-Álvarez,
Ph.D.
Pontificia Universidad Católica de Puerto Rico, Mayagüez
Natural
products have been the sources of anticancer drugs since the beginning of
treatments against disease. Approximately 36% of the new first-in-class small
molecules medicine approved by US Food and Drug Administration are derived from
natural products. Ganoderma lucidum
is a basidiomycetus fungus of oriental origin, which as a long history of use
for promoting health and longevity in China, Japan, and other Asian countries.
The fungus belongs to the mushroom that grows on the lower trucks of deciduous trees.
Ganoderma sp. have been used in
multiple in vitro and in vivo models due to its
anti-proliferative and growth inhibitory efficacy in cancer cells, especially
breast cancer. Herein, this conference will emphasize in the progress of
alternative and combinatorial therapeutics using G. lucidum extracts in breast cancer.
Presentaciones Orales
Differential sensitivity to QoI fungicide among isolates
of Colletotrichum gloeosporiodes
that cause
anthracnose on yam
Bosques-Martínez, M. and Feliciano-Rivera, M.
Department of Agro-Environmental
Sciences, University of Puerto Rico, Mayagüez
Chemical
control, based on Quinone outside Inhibitor (QoI-a.i. azoxystrobin) fungicides,
has been essential for the management of foliar diseases including yam
anthracnose. Colletotrichum gloeosporioides is the causal agent of yam
anthracnose on cultivars of Dioscorea alata in Puerto Rico. Due to the
excessive use of QoI fungicide and the lack of resistance management practices
at farm level, control failures has been detected more frequently. Therefore,
the objective of this research was to evaluate the sensitivity to azoxystrobin
of two isolates of C. gloeosporioides. One of the isolates was collected
from a farm exposed to QoI fungicide and the other isolate was collected from
the germplasm of the Corozal Agriculture Experimental Station. Sensitivity to
the product was determined through QoI fungicide amended selective media assays
and in vitro colorimetric assays to evaluate mycelial growth and spore
germination. Four concentrations of the fungicide with and without the
alternative oxidase (AOX) inhibitor salicylhydroxamic acid (SHAM) were tested
for sensitivity. Treatment effects were determined by ANOVA and treatment means
separated by Fisher’s LSD using Infostat statistical software. In the assays
evaluated both isolates showed significant differences among the doses tested,
with the combination of azoxystrobin 10 µg/ml and SHAM 100 µg/ml being the most
effective in reducing their growth. Results showed that C. gloesporioides collected
from the farm exposed to QoI fungicide grew in the fungicide amended media in a
manner similar to the non-amended treatment. However, the isolate that was not
exposed to the fungicide grew in a dose-dependent manner indicating a high
sensitivity response to the fungicide. In the absence of SHAM, the isolate
collected from the farm exposed to QoI fungicide, had no significant response
to the tested doses of azoxystrobin, indicating that the isolate was able to
circumvent the effect of the fungicide by the activation of the AOX pathway.
These results suggest that, in Puerto Rico, C. gloesporioides exposed to
QoI fungicide has possibly developed a resistance to azoxystrobin.
Aceites esenciales,
una alternativa segura para el tratamiento de dermatofitosis
Vázquez Alvarado, Y., Torres Caraballo, S., Pagán Medina, C., Morales Miranda, S., y
Arroyo-González, Hna. N.
Departamento
de Biología, Pontificia Universidad Católica de Puerto Rico, Ponce
Los dermatofitos son hongos parásitos
de la queratina, especialmente de las capas externas de uñas, piel y cabello,
que dependen de esta proteína filamentosa para su propagación. Generalmente,
estos hongos son tratados con medicamentos químicos que presentan para el
humano un alto riesgo de daño hepático y un potencial carcinogénico. Estudios
recientes han reportado que aceites esenciales, extraídos de plantas, exhiben
actividad fungicida o fungistática además de un nivel muy bajo de toxicidad. El
objetivo de esta investigación fue determinar el efecto de los aceites
esenciales de Origanum vulgare (orégano), Cinnamomum zeylanicum (hoja
de canela) y Eugenia caryophyllata (clavo) contra los dermatofitos Trichophyton
rubrum, Trichophyton mentagrophytes y Trichophyton tonsurans, que
han tenido una incidencia en la zona sur de Puerto Rico de 45.2%, 25.8% y un
6.5%, respectivamente. La efectividad de los aceites esenciales se evaluó
mediante la formación de halos de inhibición los cuales se midieron a los 7, 14
y 21 días. Los resultados mostraron que los aceites esenciales de orégano, hoja
de canela y de clavo presentaron un efecto fungicida. Sin embargo, el fungicida
de mayor efectividad fue el aceite esencial de orégano que presentó inhibición
total de dermatofitos aun después de 21 días. En conclusión, estos resultados
arrojan una promesa terapéutica inocua para combatir dermatofitos en humanos.
Botryosphaeriaceae as pathogens
of tropical exotic fruits
Soto Bauzó, A.1, Serrato Díaz, L. M. 1, Aviles
Noriega, A. 2, Rivera Vargas, L. I. 3, Goenaga, R. 4,
and Bayman Gupta, P. 1
1Department of Biology, University of Puerto Rico, Rio Piedras
Campus, San Juan
2East University, School of Science and Technology, Carolina
3Department of Crops and Agro-Environmental Sciences, University of Puerto Rico, Mayagüez
2East University, School of Science and Technology, Carolina
3Department of Crops and Agro-Environmental Sciences, University of Puerto Rico, Mayagüez
4USDA-ARS, Tropical Agriculture
Research Station, Mayagüez
Botryosphaeriaceae family fungi have been
identified to cause dieback on tropical fruit crop trees. To identify the
causative agent of dieback in rambutan and longan in Puerto Rico, a disease
survey from 2008 to 2016 was conducted at seven orchards. Diseased tissue of
rambutan, longan, mango, and mandarin showing different symptoms were
surface-sterilized and plated on PDA. Forty-one isolates in the Botryosphaeriaceae
family were isolated and identified using morphological and
molecular characters. For morphology, four different media were used.
Characters such as sporulation, colony color, and shape and size of conidia
were used to identify species. The ITS1-5.8S-ITS2 of rDNA and partial sequences
of β-tubulin and trans-elongation factor 1α (EF1-α) were amplified, sequenced
and compared with sequences of Botrysophaeriaceae in GenBank.
Pathogenicity tests were conducted on 129 rambutan and longan seedlings.
Cross-inoculations were done using 10-day old mycelium disks of 5mm in diameter
from pure cultures grown on PDA. Control seedlings were inoculated with PDA
disks. Symptoms of dieback were evaluated at 8, 14, 30 and 60 days after
inoculations (DAI). Fourteen out of 41 isolates caused dieback in
rambutan at 14 DAI. Five of these were collected from rambutan, four from
longan, two from mango and three from mandarin. Ten isolates caused dieback in
longan, of which nine were collected from rambutan, longan and mandarin (three
each) and one from mango. At 60 DAI pycnidia were observed on the trunk of
rambutan and longan trees. Five species, four Lasiodiplodia spp. and one
Neofusicoccum sp., were identified to cause dieback disease in rambutan
and longan. Cross-inoculations showed that pathogenicity is wide-spectrum,
indicating that different planting alternatives should be considered for better
crop management.
Estudio de la biodiversidad
de hifomicetos acuáticos presentes en la espuma del río Usabón en Barranquitas
Arroyo-Berríos, N.L. 1, Negrón-Martínez, C. 2 y Lozada-Troche, C.
2
1Departamento de Ciencias Naturales, Universidad de Puerto Rico, Cayey
2Departamento de Biología, Universidad de Puerto Rico, Cayey
Los hifomicetos acuáticos, también
conocidos como hongos ingoldianos, son un grupo de hongos microscópicos que se
encuentran en hojas deciduas o cualquier material vegetal que cae en los ríos o
arroyos. En los ecosistemas acuáticos, estos hongos juegan roles de gran
importancia ya que son responsables de degradar la materia vegetal, facilitando
así el reciclaje de nutrientes en niveles tróficos superiores. En el presente
estudio, se realizó un censo de la biodiversidad de los hifomicetos acuáticos
presentes en muestras de espuma obtenidas del río Usabón ubicado en Barranquitas
P.R durante el periodo de agosto 2016 hasta noviembre 2016. Se colectaron
muestras de espuma cada dos semanas las cuales fueron teñidas y observadas bajo
un microscopio para realizar la identificación y cuantificación de los hongos
ingoldianos presentes en las mismas. Cuarenta y cinco especies fueron
identificadas en las muestras de espuma del río Usabón. Los resultados
obtenidos indicaron que las especies dominantes fueron: Campylospora
filicladia, Flagellospora curvula, Campylospora sp, Helicomyces colligatus,
y H.torquatus. También se encontraron cerca de veinte especies
ocasionales. Las variaciones en las frecuencias de los hongos en el río
pudieron ser causadas por los cambios climatológicos, la disponibilidad de
sustratos, la tendencia de prevalecer en las estaciones del año y su
favorabilidad de subsistir.
Afiches de Investigaciones
Afiche
1
Caffeine degradation of Penicillium sp.
Galarza García, A. M.,
and Seda Miró, J.
Biology
Department, Pontifical Catholic University of Puerto Rico, Ponce
Caffeine degradation has been extensively studied in
bacteria, such as Pseudomonas, and produce several non-toxic metabolites
in the process. However, few fungal isolates have been reported to degrade
theobromine, the non-toxic metabolite produced by bacteria in caffeine
degradation. Due to toxic theophylline production in caffeine degradation by
fungi, they have not been considered viable for the decaffeination process. As
a result, few studies on caffeine and theobromine degradation by highly
caffeine-tolerant fungal isolates have been explored. We aim to study caffeine
and theobromine degradation from a fungal isolate (Pencillium sp.) that
was initially isolated from a fresh apple product. Preliminary studies in our
laboratory showed this fungal isolate to be highly tolerant to caffeine (25
mg/mL), which is an alkaloid commonly found in plants and is known for its role
in plant defense. Our objective also includes detection of other metabolites
produced from caffeine degradation. To perform our study, Penicillium
sp. was cultivated in Potato Dextrose Agar (PDA) at 25°C for 5 days.
Approximately 106 spores were then inoculated in a caffeine-sucrose (CS) medium
containing 1 g/l of caffeine. Three biological replicates were performed including
a positive and negative control. After 48 h of growth, 4 g/L of caffeine were
added. Cultures were observed for 7 days and samples were taken every 12 hours.
Caffeine degradation and metabolite production were analyzed using high
performance liquid chromatography (HPLC). At present, results for caffeine
degradation remain in progress and future efforts consist of analyzing
theobromine degradation.
Afiche 2
Captura de micoflora
viable en el interior del Edificio de Biología utilizando el Andersen Sampler
Noguera-Colón, A. J.,
Nazario-Ayala, V. M., Brull-Hayes, P.N. y Maldonado-Ramírez, S. L., PhD.
Departamento de Biología, Universidad de
Puerto Rico, Mayagüez
Es conocido
que los edificios y lugares públicos deben cumplir con ciertas reglas de
limpieza e higiene para poder estar certificados por las agencias pertinentes
dependiendo del uso y actividades en el edificio. En el caso del Edificio de
Biología, donde se llevan a cabo investigaciones y hay una alta frecuencia de
personas, no se pueden ignorar ninguno de los parámetros de higiene, incluyendo
la calidad del aire. Recientemente se colectaron muestras de aire para
determinar si los hongos presentes en el interior del Edificio de Biología de
la Universidad de Puerto Rico, Recinto de Mayagüez, presentan alguna amenaza a
la salud. Aquí se presentan los resultados de un muestreo realizado en enero
2017 exponiendo por 10 minutos platos de Malt Extract Agar suplementado con
cloranfenicol como superficie de colección en un Andersen Sampler de dos
etapas. Cada plato se incubó a 25±2°C por 96 horas y se anotó el crecimiento de
colonias a las 48, 72 y 96 horas. Luego, se identificaron los diferentes
morfotipos en cada plato y se transfirieron a cultivos puros. Se
observó una alta frecuencia de hongos históricamente clasificados como
alérgenos, entre ellos Cladosporium
sp. (16.6%) y Geotrichum sp. (1.49%).
También, se encontró Stachybotrys
sp. (0.186%) en el aire y en una muestra con cinta adhesiva. Además, se
encontraron hongos patógenos de plantas, como Rhizoctonia sp. (0.186%) y Nigrospora
sp. (0.186%). Estos resultados son significativos, ya que hemos encontrados
hongos patógenos para los humanos, los cuales ameritan ser eliminados del aire.
Por lo tanto, muestreos posteriores demostrarán si se tomaron las medidas
necesarias.
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3
Assessment of the abundance of fungi present in three
canopy trimming experiment sites in the Luquillo Experimental Forest
Morales
Pomalaza, B. L.1 and Cantrell Rodríguez, S. A.2
1Departamento de Biología,
Universidad de Puerto Rico, Humacao
2Departamento de Biología,
Universidad del Turabo, Gurabo
Natural phenomena like storms and hurricanes are
known to cause disturbances in forests understory. These disturbances can have
a direct effect on the different communities that inhabit the underbrush by
altering key factors like soil moisture. Fungal communities are a very crucial
part of biomass decomposition in forest understory ecosystems. In Puerto Rico,
the development of a large-scale experiment called the Canopy Trimming
Experiment (CTE), brought up data on how events like canopy trimming and debris
deposition on the forest floor can affect the microbial communities in the
soil, this included responses of fungal populations. The objective of this
study was to assess the diversity of these populations in the different
treatment sites of the CTE, to quantify and characterize it, and determine if
there was any significant difference between treatments. Fungi were sampled
from leaf litter and soil using various methods of isolation and fruiting
bodies were collected and counted from three different blocks with two
treatments, trim and control. DNA sequencing was performed with the purpose of
creating sequencing data of the diversity of the fungal communities within the
different CTE sites for future reference. It was found that there was no
significant difference in the total number of fungi in leaf litter or the
number of fruiting bodies between treatments. In contrast, total fungi in soil
were found to be significantly more abundant in trim plots than in control
plots. A total of thirty-three sampled fungi were sequenced and identified.
Fungi in leaf litter appeared to not be affected by drying in the open canopy
plots, even though the litter moisture has been shown to be reduced. Microfungi
in soil was significantly more abundant in open canopy treatments, as
hypothesized, because the moisture in the soil was apparently conserved while
being reduced on litter, thus the difference in abundance between soils.
Differences in abundance were found but in future studies differences in
species composition between treatments would like to be explored.
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4
Diversity of Thraustochytrids associated with mangroves
in Puerto Rico
Soto-López D.,
Rodríguez E., and Cafaro M.
Departamento de Biología, Universidad de
Puerto Rico, Mayagüez
Omega-3 (ω-3) oil is known for its capability to
reduce and treat heart related diseases. Nowadays the majority of our ω-3 oil
is derived from fish, but some marine microorganisms can produce similar oils.
Thraustochytrids are eukaryotic fungus-like protists known for their
biotechnological potential for ω-3 oil and other novel compounds production via
polyunsaturated fatty acids (PUFAs). This represents an alternative for the
industry to mass produce high-purity oils, meaning that demand for PUFA-rich products
is expected to increase. Thraustochytrids are decomposers usually found in
tropical marine environments, typically mangroves, macroalgae and seagrasses.
In Puerto Rico, Thraustochytrids diversity is unknown. Mangrove and seagrass
leaf-litter samples were collected from Playa Rosada, Lajas, and Playa Buyé,
Cabo Rojo to isolate strains by traditional culture methods in seawater media.
Samples were processed in the lab under sterile conditions and placed in petri
dishes with artificial or aged seawater marine media (0.1% yeast extract, 0.1%
peptone, 0.5% glucose); incubated at 28C for 24-48 hours. Microbial growth was
observed under a compound microscope to determine type of microorganism and
individual strains were purified. Results include the isolation of various
yeasts from both collecting sites and one Thraustochytrid from Lajas so far.
These were morphologically identified and will be further characterized using
molecular techniques. These findings will improve our understanding for
Thraustochytrid’s diversity in Puerto Rico and explore its possibilities for
PUFA production. Moreover, new opportunities for taxonomic and biotechnological
research will be produced.
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5
Identification of Pseudocercospora griseola races in Puerto Rico
Navarro-Monserrat, E. D.,1
Serrato-Diaz, L.,1 Porch, T.,2 and Bayman, P.1
1Department of Biology, University of
Puerto Rico - Rio Piedras, San Juan
2USDA-ARS, Tropical Agricultural
Research Station, Mayagüez
Angular Leaf Spot (ALS), caused by the fungus Pseudocercospora griseola, is a frequent
disease of the common bean (Phaseolus
vulgaris) especially in tropical and subtropical regions. P.griseola and P.vulgaris have co-evolved through time and been classified of
Mesoamerican or Andean groups. In Puerto Rico, isolates of P. griseola have been shown to be Mesoamerican; however, races have
not yet been determined. This study focuses on the isolation and identification
of races of P. griseola in Puerto
Rico. Forty-five isolates were obtained from experimental research stations at
UPR and USDA-ARS in Isabela and Juana Diaz. Races of P. griseola were identified by pathogenicity tests using a set of
twelve differential bean cultivars defined by International Center for Tropical
Agriculture (CIAT). P. griseola
isolates were grown on V8 medium and incubated for 45 days. Conidia suspensions
were used to inoculate 20-day old plants. Twenty days after inoculation
symptoms were observed and the isolates were classified into races according to
compatibility reactions. 21 isolates have been tested (9 from Juana Diaz and 12
from Isabela) and 12 races have been identified. These results suggest that the
pathogen P.griseola may contain
polymorphisms in its genome, possibly in virulence genes. This project can thus
be used as a basis to determine candidate virulence genes and select for
resistant bean varieties.
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6
Yeast associated with demosponges from Puerto Rico
Sánchez-Martínez, E.1, Rosado-Rodríguez, G.2 and Maldonado-Ramírez, S.L.1
1Department of Biology, University of
Puerto Rico, Mayagüez
2Department of Marine Sciences,
University of Puerto Rico, Mayagüez
Sponges are known for their symbiotic associations
with bacteria and to a lesser extent with fungi. It has been argued that the
association between fungi and sponges is not truly symbiotic, and that fungi
found are incidental. Nevertheless, vertically-transmitted endosymbiotic yeast
has been observed by transmission electron microscopy in sponges of the genus Chondrilla. The identity of the symbiont and its presence
in other sponge species is still unknown. This work focused on studying the
presence of yeasts in marine sponges commonly found in Puerto Rico. Samples
were taken from colonies of Ircinia
strobilina, Tedania ignis, and Chondrilla caribensis. Dilution-plate method and high-throughput
culturing by dilution-to-extinction were applied for the isolation of yeasts
from sponge samples. Plates were
incubated for up to 60 days to allow the isolation of slow-growing colonies. A
total of 36 yeast morphotypes were isolated and analyzed by sequencing the ITS
region and D1/D2 domains of the large subunit ribosomal DNA. The brewer’s
yeast, Saccharomyces cerevisiae, was
isolated from all samples, comprising the first report of this organism as a
common inhabitant in marine sponges in Puerto Rico. Other yeasts isolated
include the halophilic black yeast, Hortaea
werneckii, and an unidentified
yeast belonging to the Sporidiobolales (Basidiomycota).
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7
Sensitivity of Colletotrichum
alatae to a quinone outside inhibitor (QoI) fungicide
Escalera-García, I. A.
and Feliciano-Rivera, M.
Department
of Agro-environmental Sciences, University of Puerto Rico, Mayagüez
Yam (Dioscorea
spp.) is a staple crop of economic importance in tropical and sub-tropical
regions worldwide. Yams are very susceptible to diseases caused by a wide
spectrum of pathogens including bacteria, nematodes, virus and fungi that
significantly reduce yield and quality of the commercial product. Anthracnose
disease, caused by Colletotrichum spp.,
is one of the most destructive diseases in yam (Dioscorea alatae) production in Puerto Rico. Chemical control,
based on Quinone outside Inhibitor (QoI) fungicides, has been essential in the
management of anthracnose. However, due to the excessive use of QoI fungicide
control failures has been occurred in recent years. Hence, the purpose of this
research was to measure the sensitivity of an isolate of Colletotrichum alatae collected from commercial fields to a QoI
fungicide. Sensitivity to the product was determined using amended selective
media assays, colorimetric assays, and in vitro spore germination assays with
different fungicide concentrations (0.1, 1.0, 5.0 and 10.0 µg/ml label rate).
Fungicide treatments were evaluated with and without the alternative oxidase
inhibitor salicylhydroxamic acid (SHAM). Treatment effects were determined by
ANOVA and treatment means separated by Fisher’s LSD using InfoStat Statistical
Software. Results showed that C. alatae
grew similar to the non-amended fungicide treatment at concentrations of 0.1,
1.0, 5.0 µg/ml. At the concentration of 10 µg/ml in combination with SHAM C. alatae showed a growth reduction. The
same results were obtained in the colorimetric assays and in the spore
germination assays. These results suggest that, in Puerto Rico, C. alatae have developed a resistance to
QoI fungicides in both the mycelium and spore stage.
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8
Fungal diversity in green litter decomposition in
a hurricane experiment
Velazquez
Cruz, J1, Moreno, I.1, Barberena-Arias, M. F.1
and Cantrell, S. A.1
1Department of Biology, University of
Turabo, Gurabo
Hurricanes generate disturbances in forests such as
canopy opening, fallen trees and leaves which in turn alter physicochemical
characteristics of the habitat, as well as, decomposer activity. Litter
decomposition depends primarily on the interaction among climate, litter
quality and biota; as a consequence any change in habitats will result in
changes in these factors. Identifying the changes in the fungal community
structure in soil and forest floor litter can help understand the factors that
influence ecosystem recovery. This study is part of the Canopy Trimming
Experiment 2 performed by the Luquillo LTER at El Verde Field Station. For
this, three blocks (A, B and C) were selected, each with two plots of 20m x
20m, one plot for control and the other was subject to canopy opening and
debris addition (Trim plus debris). This experimental design represents 3
blocks x 2 plots/block (1 trim+debris/ 1 control) x 3 subplots x 3 litterbag
mesh sizes x 4 collecting times (3, 5, 12 and 24 weeks), for a total of 216
litterbags. A pool sample for each mesh size from the 3 subplots/per plot/time
was obtained for DNA extraction. DNA was extracted using Power Soil DNA
Isolation kit from MoBio. TRFLP technique was used to obtain profiles of fungal
communities using the fungal ITS region. The diversity between samples was
analyzed using NMDS and UPGMA Cluster analysis using the Bray-Curtis similarity
index. Two way Permanova shows significant differences in fungal communities
between blocks, control and trim plus debris, and mesh size. No significant
differences were observed through time. In general, the diversity of fungal
communities increase in the trim plus debris treatment. This increase is mainly
due to the addition of detritus which increases soil moistures. Fungi are the
main decomposers of detritus in the forest floor and the effect of natural
disturbances such as a hurricane on leaf litter microbial communities will help
us understand how litter decomposition might be affected. This one component of
a larger study that is analyzing the interaction between arthropods,
microorganisms, nutrient mineralization and disturbances.
Afiche 9
Tipos de hongos
cultivables transportados por Erythrodiplax
umbrata
(Insecta: Odonata:
Libellulidae)
Ruiz-Arocho J.,
Santos-Flores, C. y Maldonado-Ramírez, S.
Departamento de Biología, Universidad de
Puerto Rico, Mayagüez
El proceso
de dispersión es una de las necesidades básicas que la mayoría de los seres
vivos deben sobrellevar para la supervivencia óptima de su especie. En el reino
Mycota, la dispersión puede verse influenciada no solo por factores abióticos,
sino también por factores bióticos como dispersión en el tracto digestivo de
animales y la dispersión en la superficie de los mismos. Un ejemplo de un
animal con un alto grado de movimiento y en los cuales se han estudiado sus
capacidades motoras, así como sus rangos de dispersión, son las libélulas
(Insecta: Odonata). Por tal razón, en este experimento nos concentramos en
conocer qué tipo de hongos cultivables pueden ser transportados y
subsecuentemente dispersados por Erythrodiplax
umbrata (Odonata: Libellulidae). Para cumplir con este objetivo, se
colectaron nueve machos de la especie en una zona adyacente al Bosque de
Miradero, Mayagüez PR. Se realizó una impresión individual de las áreas
dorsales y ventrales de cada individuo en medio de cultivo de agar de harina de
maíz (CMA) y los hongos encontrados fueron aislados para su identificación en
medio de cultivo de agar de papa y dextrosa (PDA). Hasta el momento se han
recuperado 66 morfotipos de los cuales se han logrado identificar hongos
pertenecientes a los géneros Aspergillus,
Spegazzinia, Alternaria; entre varias levaduras. A pesar de que una cantidad
considerable de especímenes han sido clasificados como micelio carente de
estructuras reproductivas y aún existen otros en proceso de esporulación,
podemos inferir que, si cada morfotipo representa una especie distinta, la
cantidad de especies que pueden ser transportadas por libélulas es
considerablemente alta. Esto nos demuestra la importancia de las interacciones
hongos-invertebrados y el rol que tienen estos insectos para con el movimiento
y dispersión de hongos.
Afiche 10
Hongos fitopatógenos que afectan
al cultivo de Hierbabuena (Mentha spicata L.) en Puerto Rico
Santiago Charneco, J. L.,
Simbaña Carrera, L.L., y Rivera Vargas, L. I.
Departamento de Ciencias Agroambientales,
Universidad de Puerto Rico, Mayagüez
La Mentha spicata L. comúnmente conocida
como hierbabuena, es una especie aromática. En Puerto Rico es valorada por sus
cualidades aromáticas y su uso culinario. Esta planta aunque es producida bajo
condiciones controladas en invernadero, es afectada por microorganismos
patógenos que ocasionan pérdidas en su producción a pequeña escala. Los
síntomas más comunes son: necrosis en hojas y tallos; y manchas cloróticas en
hojas. Es por esto que el objetivo de este estudio fue identificar
microorganismos fitopatógenos asociados al cultivo de hierbabuena. Se
recolectaron plantas sintomáticas en Aguadilla crecidas en condiciones de
invernadero, en Isabela para la venta en un centro comercial y en Mayagüez de una
finca comercial. Los microorganismos se aislaron en medio de acidulado de agar
de papa y dextrosa y se incubaron por 48 h a 28°C. Los hongos fueron
caracterizados morfológicamente a través de microscopia de luz y de fases e
identificados utilizando claves morfológicas. Se obtuvieron ocho aislados de
hongos. Se identificaron los hongos fitopatógenos: Curvularia spp, Colletotrichum
spp., Aspergillus spp. y aislados
pertenecientes a la familia Botryosphaeriaceae. También se identificaron los
hongos saprófitos: Paecilomyces spp.
y Mucor sp. Además, se completaron
los postulados de Koch en ramas sanas colocadas en cámaras húmedas. En un
periodo de una semana, la mayoría de los aislados causaron clorosis en las
hojas inoculadas y luego de dos semanas se observó necrosis intervenal y
pudrición del tejido. Posteriormente se espera realizar pruebas de
patogenicidad en tiestos y caracterizar molecularmente las especies de hongos
patogénicas.
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Dynamics of microbial groups in response to simulated
hurricane at
El Yunque Rain Forest in Puerto Rico
González-Rosario, K. M.1,3,
Cantrell, S. A.1, and Pérez-Jiménez, J.R.,1-3
1School of Natural Science and
Technology
2Interdisciplinary Research
Institute
3Puerto Rico Institute for
Microbial Ecology Research, Universidad del Turabo, Gurabo
Climate variability models predict increase in
incidence and intensity of hurricanes. In Puerto Rico, hurricanes have impacted
the Luquillo Rain Forest in many occasions resulting on canopy debris deposited
in the forest floor. Inhabiting microbes
face input of complex plant biomass, increases in sunlight, and alterations in
redox potential that impact their prevalence and contributions to
biogeochemical cycles. A Canopy Trimming Experiment (CTE), that simulated the
pass of a hurricane, has been done in the Tabonuco forest. Our goal is to
determine temporal heterogeneity of two microbial groups (bacteria, fungi) in
response to detritus deposition of simulated hurricane effect. Soil samples are
being collected from plots, at different times. Two treatments are considered:
with and without detritus deposition trimmed from the local canopy. Bacteria
and fungi are being characterized independently by the molecular analyses of
four distinctive genes (16S rDNA and ITS). Bacteria was homogeneous over time
for the same plot suggesting microbial succession in which rare microbiota
became more prevalent over time. In contrast, diversity increased over time
where detritus was added. Bacterial and fungal communities exhibited spatial variation
regardless the availability of plant debris. In contrast, microbial functional
groups examined demonstrated greater heterogeneity in response to plant debris
addition (treatments). In the future, characterization of specific taxa, qPCR
and Next Generation Sequencing.
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Detecting antimycotic properties of gold and
silver nanoparticles
against Aspergillus
niger
Morales-Rodríguez, L.M.
and Colón-Mena, Y.
Department
of Natural Sciences, University of Puerto Rico, Aguadilla
Gold and silver nanoparticles (Au/Ag-NPs) have been
shown to have inhibitory effects and antibacterial activity. Over the years,
resistance to antibiotics and antimycotic agents by pathogenic mushrooms has
become a major health problem. In this study, we evaluate the inhibitory
effects of gold and silver nanoparticles against pathogenic fungus Aspergillus
niger – causative agent of
aspergillosis (lung disease) and otomycosis (fungal ear infections) in humans.
To test the effects of metal nanoparticles (Me-NPs), we performed
the Kirby-Bauer Disk Susceptibility Test on Mueller-Hinton agar plates. The
plates (n=10) were inoculated with the test strain once the desired absorbance
reading was achieved. First round, all standard paper disks (D=6mm) in five
plates were impregnated with Au-NPs only. The second round of experiments
followed the procedure previously described, but with Ag-NPs instead. Measurement of the inhibition halos was performed
after 48 hours of incubation. Our results show that silver
nanoparticles have a significantly higher inhibitory effect on the test strain
-when compared to gold nanoparticles. Therefore, silver nanoparticles have a
potent inhibitory effect against A. niger.
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Isolation and characterization of mycological bioprospects
from the
Arecibo Observatory Radio Telescope
Rivera-De Jesús, M. A.
and Ríos-Velázquez, C.
Department
of Biology, University of Puerto Rico, Mayagüez
The Arecibo Observatory is a single-dish radio
telescope, once the largest in the world, situated within the vegetation of
Arecibo, Puerto Rico that can send and receive powerful radio waves. Despite
its unique environment, the radio telescope’s microbiological flora has seldom
been studied, in particular its mycological flora. This study, in addition to
describing the mycological flora of the area of the telescope, also focuses on
exploring mycological bioprospects that could have or have acquired novel attributes
with Biotechnological or Biomedical applications. The first stage involves the
isolation of mycoflora present at the four radar support columns located at the
periphery of the radar dish. Samples
were collected using sterile swabs, serially diluted, and dispersed on Potato
Dextrose Agar (PDA) media. The samples were incubated at 25°C and the presence
of colonies was monitored daily. The mycoprospects were isolated and analyzed
macroscopic and microscopically. A total of 136 individuals have been isolated.
The most mycological samples were collected on the northeastern and
northwestern columns, with the northeastern column containing the highest
amount of yeasts of all. Some of the genera found belong to Penicillium sp., Paecilomyces sp. and Aspergillus
sp., in addition to a variety of yeasts that were also identified. Other
macroscopic colony features indicate to more genera to be identified,
demonstrating the mycological variety found in the area. Functionally, some of
the mycological bioprospects isolated have shown antibiosis activity to other
fungi and are to be further analyzed to discover potential antibiosis activity
against bacteria.
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Ubiquity of Aspergillus
spp. and other fungal Genera in green coffee beans
Rodríguez-Soto, M. M.,
Simbaña Carrera L, L. L., Dumas, J., and Rivera Vargas, L. I.
Department
of Agro-Environmental Sciences, University of Puerto Rico, Mayagüez
Different species of Aspergillus produce diverse mycotoxins such as: aflatoxins and
Ochratoxin A, known for their adverse effects on human health. These fungi
colonize tissues, epiphytically or endophytically, of economic important crops
including coffee, one of the most consumed drinks globally and a very important
agricultural commodity in Puerto Rico. Studies have reported Aspergillus
carbonarius as an important source of Ochratoxin A in coffee. Our study
aimed to elucidate the occurrence of different fungi in green coffee beans,
especially those characterized for their mycotoxin production, emphasizing Aspergillus spp. For this purpose,
two-year-old cold preserved samples of green beans from six coffee cultivars
(i.e. Frontón, Typica, Borbón, Catuaí, Catuaí honey and Limaní) harvested in
Adjuntas, Puerto Rico were evaluated. To isolate fungi, green coffee samples
were surface sterilized and placed on potato dextrose agar media amended with
antibiotics. Fungal colonies were counted, purified and identified
morphologically using light and phase microscopy. A total of 68 fungal colonies
were isolated. Aspergillus spp. (44%) was the most ubiquitous
endophytic fungal genera identified, followed by Chaetomium spp. (21%). Five different Aspergillus spp. morpho-groups were observed. Other fungal genera
identified were: Penicillium spp. (13%) and
Rhizopus sp. (4%). Eighteen percent of the isolates did not produce
reproductive structures and were not identified. Coffee cultivar Borbón showed
the highest amount of fungal colonies (n=20) while cultivar Limaní showed the
lowest (n=3). We demonstrate that a diverse group of fungal species colonize
green coffee beans and survive cold storage conditions. These findings are
important because of its known mycotoxins production and their implications in
human health.
Afiche 15
Bioprospección y biocontrol
de la Broca del Café con Beauveria
bassiana
García Rodríguez, N. M.,
Mariño Cardenas, Y. A., y Bayman-Gupta, P.
Departamento de Biología, Universidad de
Puerto Rico, Río Piedras
Reportada
por primera vez en Puerto Rico en el 2007, la broca del café (Hypothenemus hampei) es el insecto plaga
más dañino al café en Puerto Rico y el mundo. Actualmente, el manejo integrado
de la broca suele usar el hongo entomopatógeno Beauveria bassiana (Bb) como control biológico. En Puerto Rico,
este hongo es aplicado usando el producto con cepa comercial GHA (de Mycotrol®
y BotaniGard®). Nuestro objetivo es evaluar el potencial uso de cepas locales
de Bb y compararlas con la cepa comercial para el biocontrol de la broca del
café. Cepas locales de Bb fueron colectadas de brocas parasitadas en cafetales
de Puerto Rico. Se escogieron nueve de éstas y la cepa GHA, para utilizarse
pruebas de virulencia in vitro
aplicando una suspensión de 4 x 106 conidias/mL a brocas hembras en
placas petri y documentando mortandad durante ocho días. Se transfirieron las
brocas muertas a placas Petri con un papel filtro húmedo y se evaluó
diariamente durante 10 días la presencia del micelio, para luego cuantificar
las conidias producidas por broca. En el campo se comparó la persistencia de la
cepa local más virulenta y la cepa GHA, donde se asperjaron árboles con cada
una de éstas, cada dos semanas se contabilizaron los frutos con colonias
esporuladas de Bb. Se observaron diferencias significativas en la virulencia de
cepas locales y la cepa GHA, sin embargo, dos de las locales produjeron
porcentajes de mortandad similares a los registrados por la cepa GHA. Además,
una de éstas superó la cantidad de esporas producidas de la cepa GHA. En el
campo, el tratamiento con la cepa local mostró mayor persistencia de Bb en
comparación con el tratamiento con la cepa comercial. Estos resultados sugieren
que las cepas locales de B. bassiana
son un recurso genético de alto potencial para el biocontrol de la broca del
café y otros insectos plagas.
Afiche 16
Isolation of Fusarium
spp. inhibiting and Fumonisin B1
degrading microbes
from corn (Zea
mays L.) and sorghum (Sorghum bicolor L.)
Rivera Hernández, O.1,
Rivera-Vargas, L. I.1 and Kuldau, G.2
1Department of Agro-Environmental
Sciences, University of Puerto Rico, Mayagüez
2Department of Plant Pathology and
Environmental Microbiology, Pennsylvania State University
Fumonisins are secondary metabolites produced by
some species of the genus Fusarium.
These are hazardous and prevalent mycotoxins in crops such as corn and sorghum.
Fumonisins usually accumulate in grain affected by Fusarium Ear Rot caused by Fusarium verticillioides. The
possibility of reducing harmful levels of these toxins by using
fumonisin-degrading microbes seems promising. In the same way, using biological
control to inhibit the growth of Fusarium
species appears to be an efficient way of managing the disease. As a first
step towards identifying possible biological controls, we isolated two bacteria
from corn and sorghum with potential fungal inhibitory properties. Screenings
of the samples as well as co-cultures were done in order to evaluate inhibiting
organisms. Inhibition tests were conducted by co-culture of Fusarium spp. isolates M-8773 and M-8778, with potential inhibitory microbes on
Waksman’s glucose agar. Lisogeny broth was used as a negative control. For
microbes capable of degrading the mycotoxin, a suspension of corn and sorghum
leaves was incubated in a mycotoxin enriched liquid mineral medium for four
weeks. Medium changes were done between weeks to prevent organisms from taking
carbon out of dead organisms. After incubation, four Gram-negative bacterial
strains were isolated as potential degraders. Although this opens the
possibility of finding degrading organisms in host plants for F. verticillioides is not certain that
all the bacterial strains isolated are fumonisin decomposers. This can be
explained through cooperative catabolism where some bacterial species degrade
the products of other reactions that occurred first, obtaining carbon out of
other compounds in the process. HPLC and PCR are contemplated to characterize
true fumonisin degrading bacteria.
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Inventory of cultivable fungi from fresh casts of
the
earthworm Yuisia
olgae (Annelida, Benhamiidae): Preliminary results
Carrera-Martínez, R.,1,2, Rosado-Rodríguez, G.,1,3
and Maldonado-Ramírez, S.L.1
1Department of Biology, University of
Puerto Rico, Mayagüez
2Warnell School of Forestry and Natural
Resources, University of Georgia, Georgia
3Department of Marine Sciences,
University of Puerto Rico, Mayagüez
Earthworm and fungi interaction have been the focus
of many studies, as earthworms can disturb fungal diversity and species
composition. However, few studies focused on the fungal communities associated
with earthworms, including their fresh casts. Here, we propose a novel
methodology to study fungal diversity associated with earthworm casts. Two
specimens of the earthworm Yuisia olgae
were collected together and maintained alive on a container with wet soil for
about 12 hours. At the laboratory, the specimens were separated from the soil
and externally cleaned with sterilized water until no remnants of soil
particles were observed on the specimens. The earthworms were placed on two
zip-lock bags previously sterilized with ethanol that contained a piece of
sterilized paper towel soaked in sterilized water. One additional identical
system was placed without earthworms to discard fungi that could be associated
to the zip-lock bag system. The zip-lock bags were then incubated at room temperature
in the dark for 24 hours. A sample of approximately 1mg of earthworm casts was
extracted with a sterile swab. The serial dilution method was used to process
the samples that were then cultivated on Potato Dextrose Agar at room
temperature. The control system was sampled with a sterile swab and then
cultivated directly on PDA. Fungal cultures from the earthworm surface were
obtained with a sterile swab, which were then discarded as casts-associated
morphotypes. Fungi were identified using morphological characters and DNA
sequences of the ITS region. A total of 13 fungal species were isolated from
the earthworm casts. Fusarium was the
most abundant genus (frequency of 0.77), followed by Penincillium (0.13). Fusarium
lichenicola, Thrichosporon sp., Clonostachys sp. and Aphanoascus sp. were isolated for the
first time from earthworm casts. Additionally, these results contrasts with
previous publications that suggests that earthworm feeding behavior is
antagonistic to Fusarium.
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Bio-remediation of oil pollutants by fungi
González-Ferrer, S.M.1, Hernández-Lamberty, M.1 , Peterson,
K.1 and Cafaro, M.1
1Department of Biology, University of
Puerto Rico, Mayagüez
Pollution in marine environments comes from certain
components in the motor oil that indefinitely stay in these environments,
especially in the sediments. Among the chemical components of diesel, motor
oil, and other petroleum-based fuels are partially combusted polycyclic
aromatic hydrocarbons (PAHs). These have stable ring structures difficult to
degrade, hence harmful to the environment and even carcinogenic. Previous
studies of lignin degradation by fungi have shown potential towards degrading
PAHs due to the non-specific fungal secreted enzymes. Acknowledging this
possibility, we designed an experiment to use fungi isolated from coastal
mangroves to degrade waste motor oil, Purpureocillium
lillacinum, Fusarium solani, Aspergillius niger, and Aspergillius caelatus.
The experiment consisted of a base culture medium, containing aged sea water,
NO3 solution, and used motor oil as the sole carbon source. Along with the
motor oil, we tested other treatments consisting of lignin, cellulose, and a
mixture of these on the fungal samples. The purpose was to use minimal sources
of nutrients possible to fully measure fungal growth based on motor oil
degradation. Our preliminary results indicate that all four fungi have the
capacity to grow with motor oil as the only carbon source, as well as in
cellulose, lignin, and the mixture of treatments. We found that a consortium
consisting of all four fungal species was more efficient at using the waste
motor oil as a carbon source than in the treatments containing single species
isolates. We expect to analyze more fungal strains and other organisms to add
to our synergistic consortium. We will also incorporate gas chromatography-mass
spectrophotometry to measure actual degradation of the carbon compounds and
characterize the enzymes that are responsible of the degradation process.
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19
Occurrence of Cryptococcus
neoformans/gatti in dry forest areas
of the southwestern zone of Puerto Rico
Fumero Nazario, S.,
Velez Justiniano, Y., Rodriguez Golderos, G.M. and Ruiz Acevedo, A.
Department
of Biology, University of Puerto Rico, Mayagüez
Cryptococcus
neoformans/gatti are opportunistic and encapsulated yeasts that
adversely affect primarily immunosuppressed patients. Although infections of C. neoformans/gatti are relatively rare
in humans with a healthy immune system, its effects on wildlife and ecology are
poorly understood. The yeasts live freely in the environment, mainly associated
to pigeon droppings and to eucalyptus, or other tree detritus. Suspected
carriers of C. neoformans/gatti were
sampled to determine the presence of the yeast on succulent plant sources.
Surface samples were collected during dry season from Pilosocereus royenii cacti at sites affected by an unknown insect
caused disease at the southwest of Puerto Rico. The prospective C. neo/gatti isolates were preliminarily
identified by melanin production on Niger Seed Agar and then confirmed by
biochemical characterization. Two out of
thirteen samples showed yeast-like growth with non-capsulation. The presence of
C. neo/gatti or other fungi helps to
prove if the colonization process in lesions is initiated by an insect. In
addition, the lesions in the cacti provide favorable environmental conditions
for fungal community growth and development. Further studies to determine
fungi-insect relationship are needed.
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Antibiotic potential of endophytic fungi
associated with the Black Mangrove Avicennia
germinans
in Cabo Rojo, Puerto
Rico
Figueroa-Negrón, V.1 and Cafaro, M.J.1
1Department of Biology, University of
Puerto Rico, Mayagüez
The search for new antimicrobial compounds has been of great interest in recent years. These compounds are naturally produced by microorganisms including endophytic fungi isolated from mangroves. This study focuses on endophytic fungi isolated from leaves and seeds of the black mangrove (Avicennia germinans) from Bahia Salinas, Cabo Rojo, Puerto Rico. We isolated 42 fungal endophytes from which nine identified strains were tested for antimicrobial secondary metabolite production. We selected fungal strains from Penicillium, Aspergillus flavus, Aspergillus clavatus, Stereum, Nigrospora, Purpureocillium, Engyodontium, Hortaea and Bionectria for the analysis. Four bacterial strains: Escherichia coli, Pseudomonas sp., Serratia sp. and Staphylococcus sp. and two yeast strains of Candida albicans and C. tropicalis were used to perform bioassays. Growth curves were prepared for yeast and bacteria. Fungal extracts were added after lag, exponential and stationary growth phase of each bacteria and yeast. Our results showed that Aspergillus flavus and Aspergillus clavatus extracts strongly decreased the growth of C. albicans and E. coli, while Nigrospora only affected the growth of E. coli. There was no significant change in the growth of C. tropicalis or the other bacterial strains. These results could open a way for future discoveries of new antimicrobial o antifungal compounds.
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Phytophthora spp. y especies de hongos
asociadas a daños en la producción de
piña (Ananas comosus) en
Puerto Rico
Veléz Negrón, Y. I., Simbaña Carrera, L. L. y Rivera Vargas, L. I.
Departamento de Ciencias Agroambientales, Universidad de Puerto Rico,
Mayagüez
La piña (Ananas comosus) es un cultivo de
importancia en Puerto Rico. En los últimos años la producción ha alcanzado
5,332 toneladas, valorándose en $5 millones para el periodo 2014-2015. Este
cultivo se ve afectado por microorganismos como hongos y oomicetos que ocasionan
graves pérdidas en la producción, incrementando la preocupación entre los
productores. Phytophthora cinammomi,
es uno de los patógenos más importantes que ocasiona una pudrición en la base
del cogollo de la fruta, las hojas centrales presentan una coloración amarilla
y las mismas pueden ser desprendidas con facilidad. Debido, a esta preocupación
se esbozaron dos objetivos: determinar la presencia de Phytophthora spp. en zonas productoras de piña de la isla e
identificar hongos asociados a otros daños observados en el campo. Las muestras
de tejido vegetal enfermo fueron recolectadas en las localidades de Lajas,
Guánica, Comerío y Santa Isabel. En el laboratorio se prepararon cámaras
húmedas y material vegetal enfermo. A partir del micelio obtenido, se sembraron
muestras directamente en medios sintéticos (i.e. PDA, V8 y PARPH). Las placas
se incubaron de 48 horas a 28°C. Los microorganismos patógenos fueron
identificados mediante microscopía de luz y de fases. Los géneros de
microorganismos fueron identificados utilizando claves taxonómicas. Se han
identificado los siguientes microorganismos: Phytophthora spp., Fusarium
spp., Curvularia y tres aislados de
hongos pertenecientes a la Familia Botryosphaeriaceae Hasta el momento el
patrón de distribución sintomatológica propia de Phytophthora spp, está presente en todas las zonas muestreadas. Se
estarán realizando pruebas de patogenicidad y caracterizándolos mediante
análisis molecular con diferentes regiones genéticas (ITS del rDNA y el gen
COI).