Biogenic production of silver, zinc oxide, and cuprous oxide nanoparticles, and their impregnation into textiles with antiviral activity against SARS-CoV-2.

Nanotechnology is being used to fight off infections caused by viruses, and one of the most outstanding nanotechnological uses is the design of protective barriers made of textiles functionalized with antimicrobial agents, with the challenge of combating the SARS-CoV-2 virus, the causal agent of COVID-19. This research is framed within two fundamental aspects: the first one is linked to the proposal of new methods of biogenic synthesis of silver, cuprous oxide, and zinc oxide nanoparticles using organic extracts as reducing agents. The second one is the application of nanomaterials in the impregnation (functionalization) of textiles based on methods called "in situ" (within the synthesis), and "post-synthesis" (after the synthesis), with subsequent evaluation of their effectiveness in reducing the viral load of SARS-CoV-2. The results show that stable, monodisperse nanoparticles with defined geometry can be obtained. Likewise, the "in situ" impregnation method emerges as the best way to adhere nanoparticles. The results of viral load reduction show that 'in situ' textiles with Cu2O NP achieved a 99.79% load reduction of the SARS-CoV-2 virus.

Characterization of the bacterial microbiome of Amblyomma scalpturatum and Amblyomma ovale collected from Tapirus terrestris and Amblyomma sabanerae collected from Chelonoidis denticulata, Madre de Dios- Peru.

BACKGROUND: Ticks are arthropods that can host and transmit pathogens to wild animals, domestic animals, and even humans. The microbiome in ticks is an endosymbiotic, pathogenic and is yet to be fully understood. RESULTS: Adult male Amblyomma scalpturatum (A. scalpturatum) and Amblyomma ovale (A. ovale) ticks were collected from Tapirus terrestris (T. terrestris) captured in the rural area of San Lorenzo Village, and males Amblyomma sabanerae were collected from Chelonoidis denticulate (C. denticulate) of the Gamita Farm in the Amazon region of Madre de Dios, Peru. The Chao1 and Shannon-Weaver analyses indicated a greater bacterial richness and diversity in male A. sabanerae (Amblyomma sabanerae; 613.65-2.03) compared to male A. scalpturatum and A. ovale (A. scalpturatum and A. ovale; 102.17-0.40). Taxonomic analyses identified 478 operational taxonomic units representing 220 bacterial genera in A. sabanerae and 86 operational taxonomic units representing 28 bacterial genera in A. scalpturatum and A. ovale. Of the most prevalent genera was Francisella (73.2%) in A. sabanerae, and Acinetobacter (96.8%) in A. scalpturatum and A. ovale to be considered as the core microbiome of A. sabanerae and A. scalpturatum/A. ovale respectively. CONCLUSIONS: We found a high bacterial diversity in male of A. sabanerae collected from C. denticulata showed prevalence of Francisella and prevalence of Acinetobacter in male A. scalpturatum and A. ovale collected from T. terrestris. The greatest bacterial diversity and richness was found in males A. sabanerae. This is the first bacterial metagenomic study performed in A. scalpturatum/A. ovale and A. sabanerae collected from T. terrestris and C. denticulata in the Peruvian jungle.

Characterization of the bacterial microbiome of Rhipicephalus (Boophilus) microplus collected from Pecari tajacu "Sajino" Madre de Dios, Peru.

Ticks are arthropods that can host and transmit pathogens to wild animals, domestic animals, and even humans. The bacterial microbiome of adult (males and females) and nymph Rhipicephalus microplus ticks collected from a collared peccary, Pecari tajacu, captured in the rural area of Botijón Village in the Amazon region of Madre de Dios, Peru, was evaluated using metagenomics. The Chao1 and Shannon-Weaver analyses indicated greater bacterial richness and diversity in female ticks (GARH; 375-4.15) and nymph ticks (GARN; 332-4.75) compared to that in male ticks (GARM; 215-3.20). Taxonomic analyses identified 185 operational taxonomic units representing 147 bacterial genera. Of the 25 most prevalent genera, Salmonella (17.5%) and Vibrio (15.0%) showed the highest relative abundance followed by several other potentially pathogenic genera, such as Paracoccus (7.8%), Staphylococcus (6.8%), Pseudomonas (6.6%), Corynebacterium (5.0%), Cloacibacterium (3.6%), and Acinetobacter (2.5%). In total, 19.7% of the detected genera are shared by GARH, GARM, and GARN, and they can be considered as the core microbiome of R. microplus. To the best of our knowledge, this study is the first to characterize the microbiome of ticks collected from P. tajacu and to report the presence of Salmonella and Vibrio in R. microplus. The pathogenic potential and the role of these bacteria in the physiology of R. microplus should be further investigated due to the possible implications for public health and animal health in populations neighboring the habitat of P. tajacu.

Meteorological impact on the COVID-19 pandemic: A study across eight severely affected regions in South America.

The role of meteorological factors in the transmission of the COVID-19 still needs to be determined. In this study, the daily new cases of the eight severely affected regions in four countries of South America and their corresponding meteorological data (average temperature, maximum temperature, minimum temperature, average wind speed, visibility, absolute humidity) were collected. Daily number of confirmed and incubative cases, as well as time-dependent reproductive number (Rt) was calculated to indicate the transmission of the diseases in the population. Spearman's correlation coefficients were assessed to show the correlation between meteorological factors and daily confirmed cases, daily incubative cases, as well as Rt. In particular, the results showed that there was a highly significant correlation between daily incubative cases and absolute humidity throughout the selected regions. Multiple linear regression model further confirmed the negative correlation between absolute humidity and incubative cases. The absolute humidity is predicted to show a decreasing trend in the coming months from the meteorological data of recent three years. Our results suggest the necessity of continuous controlling policy in these areas and some other complementary strategies to mitigate the contagious rate of the COVID-19.

Diseño y evaluación de la expresión de una potencial vacuna de ADN contra el virus de la Tilapia de Lago (TiLV)

El Virus de la Tilapia del Lago (TiLV), es un patógeno causante de mortalidades masivas tanto en poblaciones de tilapias cultivadas y silvestres alrededor del mundo. El desarrollo de una vacuna efectiva contra este patógeno emergente es imperativo para prevenir pérdidas económicas. En este trabajo se diseñó y evaluó un vector de expresión como una potencial vacuna de ADN contra este virus. Inicialmente, se realizó un análisis de enhebramiento para predecir las estructuras tridimensionales y las funciones de las proteínas del TiLV. Se encontraron homologías estructurales entre las proteínas correspondientes al segmento genómico 1 y al segmento genómico 4 del TiLV, con las proteínas de ARN polimerasa dependiente de ARN del virus de la influenza B (56%) y la proteína neuraminidasa que pertenece a la cápside del virus de la influenza A (12%), respectivamente. Se insertó el producto de PCR del gen neuraminidasa viral en el vector plasmídico de expresión pCMV. Finalmente, se inyectó el constructo plasmídico en juveniles de la tilapia del Nilo Oreochromis niloticus y se midió su expresión mediante RT-PCR en tiempo real a las 8h, 16h, 24h, 72h después de la segunda inyección inmunizante. Se logró detectar expresión génica en los cuatro tiempos evaluados, con mayor expresión a las 16 horas post inyección. Estos resultados constituyen el primer paso para el desarrollo de una vacuna efectiva para la protección de los stocks de tilapias alrededor del mundo.

Tilapia Lake Virus (TiLV) is a pathogen that causes massive mortalities in both cultured and wild tilapia populations around the world. The development of an effective vaccine against this emerging pathogen is imperative to prevent economic losses. In this work an expression vector was designed and evaluated as a potential DNA vaccine against this virus. Initially, a threading analysis was done to predict the threedimensional structures and functions of the TiLV proteins. Structural homologies were found between the TiLV proteins corresponding to the genomic segment 1 and the genomic segment 4, with the RNA-dependent RNA polymerase proteins of the influenza B virus (56%) and the neuraminidase protein belonging to the influenza A virus capsid (12%), respectively. The PCR product of the viral neuraminidase gene was inserted into the expression plasmid vector pCMV. Finally, the plasmid construct was injected into juveniles of the Nile tilapia Oreochromis niloticus and its expression was measured by real time RT-PCR at 8h, 16h, 24h, and 72h after the second immunizing injection. It was possible to detect gene expression in the four evaluated times and greater expression at 16 hours post injection. These results are the first step in the development of an effective vaccine for the protection of tilapia stocks around the world.

Genetic structure, phylogeography, and demography of Anadara tuberculosa (Bivalvia) from East Pacific as revealed by mtDNA: Implications to conservation.

Wild populations of the pustulose ark, Anadara tuberculosa (Bivalvia), an emblematic species of the East Pacific mangrove ecosystem declined in South American countries (Colombia, Ecuador, and Peru) mainly due to overharvesting and habitat loss or degradation. Understanding the genetic aspects of geographic variations and population structure of A. tuberculosa, currently unknown, appears as a priority to fishery authorities in order to elaborate integrated and collaborative conservation policies for fishery management, aquaculture, and stock enhancement programs. We used mtDNA sequence data to investigate haplotype diversity, genetic structure, and demography of A. tuberculosa. Results indicate genetic homogeneity of populations distributed north and south of the equator, respectively. However, statistically significant differentiation emerged between northern and southern populations with pairwise ф ST values ranging between 0.036 and 0.092. The oceanic current system acting in the area (Panama Current and Humboldt Current) might play a role in limiting the larval dispersal of the species, still poorly understood. Demography reconstruction supported recent population expansion, possibly started after last glacial maximum. Our results would suggest separate and independent management of populations north and south of the equator.

[Molecular characterization of pathogenic bacteria of the respiratory tract in peruvian patients with cystic fibrosis]. / Caracterización molecular de bacterias patógenas de las vías respiratorias de pacientes peruanos con fibrosis quística.

OBJECTIVES.: To molecularly characterize the pathogenic bacteria of the respiratory tract isolated from patients with cystic fibrosis (CF) in Peru. MATERIALS AND METHODS.: Bacterial communities cultured from sputum samples of pediatric and adult patients with CF admitted to the Edgardo Rebagliati Martins National Hospital and the National Institute of Child Health were characterized. Standard microbiological techniques were used for bacterial culture, and gene sequencing of 16S rRNA and matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry and tandem MALDI-TOF mass spectrometry (MALDI TOF/TOF) were used for molecular characterization. RESULTS.: Seventeen bacterial strains were characterized by 16S rRNA sequencing, and the identified pathogenic bacteria were Pseudomonas aeruginosa (31.5%), Staphylococcus aureus (12.6%), Pseudomonas spp. (11.8%), and Klebsiella oxytoca (3.1%). MALDI-TOF analysis generated a series of spectra representative of each isolated bacterial species, whereas MALDI TOF/TOF analysis identified the peptides and proteins of the most common strains and provided data on pathogenicity and sensitivity to antibiotics. CONCLUSIONS.: The primary pathogenic microorganisms found in the respiratory tract of patients with CF in Peru were the same as those found in other countries. This study is the first to perform 16S rRNA sequencing as well as MALDI-TOF and MALDI-TOF/TOF analysis of the bacterial pathogens circulating in Peru. The inclusion of proteomic analysis further allowed for the identification of native microorganisms involved in CF.

OBJETIVOS.: Caracterizar a nivel molecular las bacterias patógenas de las vías respiratorias de pacientes peruanos con fibrosis quística (FQ). MATERIALES Y MÉTODOS.: Se caracterizaron las comunidades bacterianas cultivables a partir de muestras de esputo de pacientes pediátricos y adultos con FQ registrados en el Hospital Nacional Edgardo Rebagliati Martins y el Instituto Nacional de Salud del Niño (INSN). Para el cultivo bacteriano se utilizaron técnicas microbiológicas estándares, y para la caracterización molecular la secuenciación del gen ARNr 16S y espectrometría de masas de tipo desorción/ionización con láser asistido por matriz con tiempo de vuelo (MALDI TOF) y MALDI TOF/TOF. RESULTADOS.: Por secuenciación del ARNr 16S se identificaron 127 cepas, encontrando las bacterias patógenas Pseudomonas aeruginosa (31,5%), Staphylococcus aureus (12,6%), Pseudomonas spp. (11,8%), Klebsiella oxytoca (3,1%), otras especies mostraron baja prevalencia. El análisis por MALDI TOF permitió obtener una serie de espectros representativos de cada especie aislada, mientras que el análisis por MALDI TOF/TOF reveló péptidos y proteínas de las especies más comunes con informaciones complementarias que revelarían datos sobre su patogenicidad o sensibilidad a antibióticos. CONCLUSIONES.: Los principales microorganismos patógenos encontrados en las vías respiratorias son similares a los reportados en otros países. Estos son los primeros hallazgos en Perú que muestran la caracterización bacteriana por secuenciación del ARNr 16S, por MALDI TOF y MALDI TOF TOF. Los hallazgos permiten la identificación bacteriana de microorganismos nativos relacionados con la FQ basada en el análisis de su proteoma.

Respuesta fisiológica y molecular de Anadara tuberculosa (Arcoida: Arcidae) al estrés de salinidad / Physiological and molecular response of Anadara tuberculosa (Arcoida: Arcidae) to salinity stress

Resumen: La concha negra Anadara tuberculosa es una especie emblemática del ecosistema manglar que está actualmente en condición vulnerable. El desarrollo de su acuicultura requiere identificar biomarcadores moleculares, en particular asociados al estrés por salinidad en mira al inicio de programas de mejoramiento genético. Se recolectaron ejemplares de Anadara tuberculosa del manglar colindante a la Bahía de Puerto Pizarro (Tumbes, Perú) entre enero 2015 y febrero 2016. Estos individuos fueron sometidos a condiciones de estrés hipo-osmótico (extremo: 5 y 10 ppt); (moderado: 15 y 25 ppt) y sin estrés (grupo control: 33 ppt) por 16 días después de haber sido separados en grupos de diez animales y por triplicado. La presencia de biomarcadores del estrés por salinidad fue evaluada a nivel genético con la detección por PCR de 19 genes reportados como actores claves de la osmorregulación en bivalvos como ostras y mejillones y a nivel proteomico con la secuenciación de péptidos expresados en tejidos de animales expuestos a diferentes salinidades por espectrometría de doble masa. Ninguno de los marcadores genéticos probados pudo ser amplificado por PCR lo que sugiere que A. tuberculosa presente diferencias genéticas significativas en comparación con otros moluscos. El análisis proteómico realizado por MALDI TOF/TOF a nivel de tejido branquial de A. tuberculosa permitió identificar 26 péptidos expresados de formas presenciales y diferenciales a las diferentes salinidades evaluadas, resaltando posibles marcadores como la HSP70 y una proteína transmembrana de transporte de cloruro que están relacionadas con la adaptación a la salinidad. Estas secuencias aminoacídicas permitirán diseñar iniciadores nucleotidicos específicos a A. tuberculosa para la puesta en marcha de futuras investigaciones en ecofisiología de este importante recurso.

Abstract: The pustulose ark A. tuberculosa is an emblematic species of mangrove ecosystem that is currently in a vulnerable condition. The development of its aquaculture, to begin with genetic breeding programs, requires the identification of molecular biomarkers, particularly those associated with salinity stress. With this purpose, specimens of A. tuberculosa were collected from the adjacent mangroves of Puerto Pizarro bay (Tumbes, Perú), from January 2015 to February 2016. Different assays (groups of ten animals in triplicate) were undertaken in separated periods of 16 days: hypo-osmotic stress (extreme: 5, 10 ppt); (Moderate: 15, 25 ppt) and no stress (control group: 33 ppt). The presence of salinity stress biomarkers was assessed at the genetic level throughout PCR detection of 19 genes reported to be key actors in osmoregulation, and at the proteomic level with the sequencing of peptides (tandem mass spectrometry MALDI TOF/TOF), expressed in ark tissues exposed to different salinities. None of the tested genetic markers could be amplified by PCR, suggesting that A. tuberculosa has significant genetic differences compared to other mollusks. Proteomic analysis by mass spectrometry on A. tuberculosa gill tissue, allowed to identify 26 peptides expressed in presential and differential forms at different salinities, highlighting possible markers such as HSP70 and trans-membrane chloride channel transportation protein, to be related with salinity adaptation. These amino acid sequences will allow the design of target specific primers for A. tuberculosa, to implement future research in ecophysiology of this important fishery resource. Rev. Biol. Trop. 65 (3): 1142-1151. Epub 2017 September 01.

Caracterización molecular de bacterias patógenas de las vías respiratorias de pacientes peruanos con fibrosis quística / Molecular characterization of pathogenic bacteria of the respiratory tract in peruvian patients with cystic fibrosis

RESUMEN Objetivos. Caracterizar a nivel molecular las bacterias patógenas de las vías respiratorias de pacientes peruanos con fibrosis quística (FQ). Materiales y métodos. Se caracterizaron las comunidades bacterianas cultivables a partir de muestras de esputo de pacientes pediátricos y adultos con FQ registrados en el Hospital Nacional Edgardo Rebagliati Martins y el Instituto Nacional de Salud del Niño (INSN). Para el cultivo bacteriano se utilizaron técnicas microbiológicas estándares, y para la caracterización molecular la secuenciación del gen ARNr 16S y espectrometría de masas de tipo desorción/ionización con láser asistido por matriz con tiempo de vuelo (MALDI TOF) y MALDI TOF/TOF. Resultados. Por secuenciación del ARNr 16S se identificaron 127 cepas, encontrando las bacterias patógenas Pseudomonas aeruginosa (31,5%), Staphylococcus aureus (12,6%), Pseudomonas spp. (11,8%), Klebsiella oxytoca (3,1%), otras especies mostraron baja prevalencia. El análisis por MALDI TOF permitió obtener una serie de espectros representativos de cada especie aislada, mientras que el análisis por MALDI TOF/TOF reveló péptidos y proteínas de las especies más comunes con informaciones complementarias que revelarían datos sobre su patogenicidad o sensibilidad a antibióticos. Conclusiones. Los principales microorganismos patógenos encontrados en las vías respiratorias son similares a los reportados en otros países. Estos son los primeros hallazgos en Perú que muestran la caracterización bacteriana por secuenciación del ARNr 16S, por MALDI TOF y MALDI TOF TOF. Los hallazgos permiten la identificación bacteriana de microorganismos nativos relacionados con la FQ basada en el análisis de su proteoma.

ABSTRACT Objectives. To molecularly characterize the pathogenic bacteria of the respiratory tract isolated from patients with cystic fibrosis (CF) in Peru. Materials and methods. Bacterial communities cultured from sputum samples of pediatric and adult patients with CF admitted to the Edgardo Rebagliati Martins National Hospital and the National Institute of Child Health were characterized. Standard microbiological techniques were used for bacterial culture, and gene sequencing of 16S rRNA and matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry and tandem MALDI-TOF mass spectrometry (MALDI TOF/TOF) were used for molecular characterization. Results. Seventeen bacterial strains were characterized by 16S rRNA sequencing, and the identified pathogenic bacteria were Pseudomonas aeruginosa (31.5%), Staphylococcus aureus (12.6%), Pseudomonas spp. (11.8%), and Klebsiella oxytoca (3.1%). MALDI-TOF analysis generated a series of spectra representative of each isolated bacterial species, whereas MALDI TOF/TOF analysis identified the peptides and proteins of the most common strains and provided data on pathogenicity and sensitivity to antibiotics. Conclusions. The primary pathogenic microorganisms found in the respiratory tract of patients with CF in Peru were the same as those found in other countries. This study is the first to perform 16S rRNA sequencing as well as MALDI-TOF and MALDI-TOF/TOF analysis of the bacterial pathogens circulating in Peru. The inclusion of proteomic analysis further allowed for the identification of native microorganisms involved in CF.

[Frequency of the most common mutations of the CFTR gene in peruvian patients with cystic fibrosis using the ARMS-PCR technique]. / Frecuencia de las mutaciones más comunes del gen CFTR en pacientes peruanos con fibrosis quística mediante la técnica ARMS-PCR.

OBJECTIVES.: To determine the frequency of the ten most common mutations of the CFTR gene reported in Latin Americausing amplification-refractory mutation system-polymerase chain reaction (ARMS-PCR) in patients with cystic fibrosis (CF) in two referral hospitals in Peru during the year 2014. MATERIALS AND METHODS.: The frequency of the ten most common mutations of the CFTR gene was assessed in patients of the Hospital Nacional Edgardo Rebagliati Martins and the Instituto Nacional de Salud del Niño, both located in Lima, Peru. Blood samples were collected from 36 patients with CF, and the ARMS-PCR technique was used to determine the presence of these mutations. RESULTS.: The study group included 73.5% of patients with a known diagnosis of CF in the country when the study was carried out. ARMS-PCR allowed three of the mutations to be identified in a combined 30.6% of the alleles from patients with CF, and 64.9% of the mutated alleles were not identified. The mutations found were p.Phe508del (22,2%), p.Gly542* (6,9%), and p.Arg1162* (1,4%). CONCLUSIONS.: There is significant variability in both the frequency and type of mutations present in our study population and in what has been reported in other Latin American countries. It is necessary to perform studies that use complete sequencing technology for the CFTR gene to identify other mutations present in our population.

Frecuencia de las mutaciones más comunes del gen CFTR en pacientes peruanos con fibrosis quística mediante la técnica ARMS-PCR / Frequency of the most common mutations of the CFTR gene in peruvian patients with cystic fibrosis using the ARMS-PCR technique

RESUMEN Objetivos. Determinar la frecuencia de las diez mutaciones más comúnmente reportadas en América Latina del gen CFTR mediante Sistema de Mutación Refractario a la amplificación por PCR (ARMS-PCR) en los pacientes con fibrosis quística (FQ) de dos instituciones hospitalarias de referencia en el Perú durante el año 2014. Materiales y métodos. Se evaluó la frecuencia de las diez comúnmente reportadas más comúnmente reportadas del gen CFTR en los pacientes del Hospital Nacional Edgardo Rebagliati Martins y el Instituto Nacional de Salud del Niño, ambos ubicados en Lima, Perú. Se recogieron muestras de sangre de 36 pacientes con FQ y se utilizó la técnica de ARMS-PCR para determinar la presencia de tales mutaciones. Resultados. Se incluyó al 73,5% de los pacientes con diagnóstico conocido de FQ en el país al momento en que se realizó el estudio. El diagnóstico por ARMS-PCR permitió identificar las mutaciones en 30,6% de los alelos de los pacientes con FQ, el 64,9% de los alelos mutados no fue identificado. Las mutaciones encontradas fueron p.Phe508del (22,2%), p.Gly542* (6,9%) y p.Arg1162* (1,4%). Conclusiones. Existe una variabilidad significativa de las mutaciones presentes en nuestra población de estudio en comparación con lo reportado en otros países de Latinoamérica, tanto en la frecuencia como en el tipo. Es necesario realizar estudios que usen la tecnología de secuenciación completa del gen CFTR para identificar otras mutaciones presentes en nuestra población.

ABSTRACT Objectives. To determine the frequency of the ten most common mutations of the CFTR gene reported in Latin Americausing amplification-refractory mutation system-polymerase chain reaction (ARMS-PCR) in patients with cystic fibrosis (CF) in two referral hospitals in Peru during the year 2014. Materials and Methods. The frequency of the ten most common mutations of the CFTR gene was assessed in patients of the Hospital Nacional Edgardo Rebagliati Martins and the Instituto Nacional de Salud del Niño, both located in Lima, Peru. Blood samples were collected from 36 patients with CF, and the ARMS-PCR technique was used to determine the presence of these mutations. Results. The study group included 73.5% of patients with a known diagnosis of CF in the country when the study was carried out. ARMS-PCR allowed three of the mutations to be identified in a combined 30.6% of the alleles from patients with CF, and 64.9% of the mutated alleles were not identified. The mutations found were p.Phe508del (22,2%), p.Gly542* (6,9%), and p.Arg1162* (1,4%). Conclusions. There is significant variability in both the frequency and type of mutations present in our study population and in what has been reported in other Latin American countries. It is necessary to perform studies that use complete sequencing technology for the CFTR gene to identify other mutations present in our population.