Sub-lineage B.1.6 of hMPXV in a global context: Phylogeny and epidemiology.

During the 2022 COVID-19 pandemic, monkeypox emerged as a significant threat to global health. The virus responsible for the disease, the human monkeypox virus (hMPXV), underwent various genetic changes, resulting in the emergence of over a dozen distinct lineages, which could be identified by only a small number of unique mutations. As of January 25, 2023, genomic information of hMPXV generated had reached 4632 accessions in the GISAID database. In this study, we aimed to investigate the epidemiological and phylogenetic characteristics of the B.1.6 sub-lineage of hMPXV in Peru, compared with other circulating sub-lineages during the global outbreak. The B.1.6 sub-lineage, characterized by the 111029G>A mutation, was estimated to have emerged in June 2022 and was found mainly in Peru. Most cases (95.8%) were men with an average age of 33 years, and nearly half of the patients had HIV, of whom only 77.35% received antiretroviral therapy. Our findings revealed that the B.1.6, B.1.4, and B.1.2 sub-lineages were well represented and had a higher number of mutations despite having the lowest media substitution rates per site per year. Moreover, it was estimated that B.1.2 and B.1.4 appeared in February 2022 and were the first two sub-lineages to emerge. A mutation profile was also obtained for each sub-lineage, reflecting that several mutations had a pattern similar to the characteristic mutation. This study provides the first estimation of the substitution rate and ancestry of each monkeypox sub-lineage belonging to the 2022 outbreak. Based on our findings, continued genomic surveillance of monkeypox is necessary to understand better and track the evolution of the virus.

Neutralización del virus SARS-CoV-2 (linaje B.1.1) por suero hiperinmune de llama (Lama glama) en cultivo de células Vero / Neutralization of SARS-CoV-2 (lineage B.1.1) by hyperimmune llama (Lama glama) serum in Vero cell culture

Objetivo. Evaluar la respuesta serológica de anticuerpos de una llama (Lama glama) a la inmunización del virus SARS-CoV-2 (linaje B.1.1) y la capacidad neutralizante del suero de llama hiperinmune frente al virus SARS-CoV-2 (linaje B.1.1) en células Vero. Materiales y métodos. Se inmunizó una llama con el virus SARS-CoV-2 inactivado (Linaje B.1.1) y se analizaron muestras de suero para evaluar el nivel de anticuerpos mediante ELISA, así como la reactividad a antígenos de SARS-CoV-2 mediante Western Blot. Además, se evaluó la neutralización viral en cultivos celulares por la Prueba de Neutralización por Reducción de Placas (PRNT, por sus siglas en inglés). Resultados. Se observó un aumento en la serorreactividad en la llama inmunizada desde la semana 4 en adelante. Los títulos de anticuerpos fueron más elevados en el séptimo refuerzo de inmunización. Los resultados de Western Blot confirmaron los hallazgos positivos del ELISA, y los anticuerpos del suero inmune reconocieron varias proteínas virales. El ensayo de neutralización (PRNT) mostró una neutralización viral visible, concordante con los resultados de ELISA y Western Blot. Conclusiones. Los hallazgos sugieren que el suero hiperinmune de llama podría constituir una fuente de anticuerpos terapéuticos contra las infecciones por el virus SARS-CoV-2 (linaje B.1.1) y que deberá ser evaluado en estudios posteriores.

Objective. To evaluate the serological antibody response of a llama (Lama glama) to SARS-CoV-2 (B.1.1 lineage) immunization and the neutralizing capacity of hyperimmune llama serum against SARS-CoV-2 virus (B.1.1 lineage) in Vero cells. Materials and methods. A llama was immunized with inactivated SARS-CoV-2 (B.1.1 lineage). Serum samples were analyzed to evaluate the level of antibodies by ELISA, as well as reactivity to SARS-CoV-2 antigens by Western Blot. In addition, viral neutralization in cell cultures was assessed by the Plate Reduction Neutralization Test (PRNT). Results . Seroreactivity increased in the immunized llama from week 4 onwards. Antibody titers were the highest after the seventh immunization booster. Western blot results confirmed the positive ELISA findings, and immune serum antibodies recognized several viral proteins. The neutralization assay (PRNT) showed visible viral neutralization, which was in accordance with the ELISA and Western Blot results. Conclusions. The findings suggest that hyperimmune llama serum could constitute a source of therapeutic antibodies against SARS-CoV-2 infections (lineage B.1.1), and should be studied in further research.

Clonación del gen caf1 de yersinia pestis en el centro nacional de productos biológicos del Instituto Nacional de Salud / Cloning of the caf1 gene from yersinia pestis at the National Center for Biological Products of the National Institute of Health

El objetivo del presente trabajo fue amplificar y clonar la secuencia codificante del gen caf1 de Yersinia pestis en el plásmido pET32a (+). Para esta investigación, se empleó una cepa nativa Y.

Emergence of Dengue Virus Serotype 2 Cosmopolitan Genotype, Brazil.

We used nanopore sequencing and phylogenetic analyses to identify a cosmopolitan genotype of dengue virus serotype 2 that was isolated from a 56-year-old male patient from the state of Goiás in Brazil. The emergence of a cosmopolitan genotype in Brazil will require risk assessment and surveillance to reduce epidemic potential.

Características de los primeros casos reportados como sospechosos de Monkeypox en el Perú / Characteristics of the first cases reported as suspected Monkeypox in Peru

RESUMEN Se presenta el caso de 9 pacientes reportados en el contexto de la alerta sanitaria por el aumento de casos de infección por el virus de Monkeypox en el mundo en países no endémicos. Es importante conocer de forma práctica los criterios epidemiológicos y clínicos más importantes para el descarte de viruela símica en el actual contexto de trasmisión en el Perú. Se discute los criterios de los casos confirmados respecto a otras enfermedades que son parte del diagnóstico diferencial como varicela, síndrome mano pie boca, entre otros.

ABSTRACT The case of 9 patients reported in the context of the health alert due to the increase in cases of Monkeypox virus infection in the world in non-endemic countries is presented. It is important to know in a practical way the most important epidemiological and clinical criteria that make us think about ruling out Monkeypox in the current context of transmission in Peru. The characteristics of the confirmed cases are discussed versus those of other diseases that are part of the differential diagnosis such as chickenpox, hand-foot-mouth syndrome, etc.

Cloning, expression and seroreactivity of the recombinant lipopolysaccharide assembly protein - D (LptD) from Bartonella bacilliformis. / Clonamiento, expresión y seroreactividad de la proteína recombinante de ensamblaje de lipopolisacáridos - D (LptD) de Bartonella bacilliformis.

OBJECTIVE.: To evaluate in silico and at the serological level the antigenic potential of the recombinant extracellular domain of the lipopolysaccharide assembly protein - D (LptD) of Bartonella bacilliformis (dexr_LptD). MATERIALS AND METHODS.: Through in silico analysis, we selected a B. bacilliformis protein with antigenic and immunogenic potential. The selected protein gene was cloned into Escherichia coli TOP10 and expressed in Escherichia coli BL21 (DE3) pLysS. Recombinant protein was expressed using isopropyl-ß-D-1-thiogalactopyranoside (IPTG) and induction conditions were optimized. Finally, it was purified with Ni-IDA resin (His60 Ni Superflow) and a Western Blot assay was conducted. RESULTS.: In silico, the selected protein was LptD because it is located in the outer membrane and is antigenic and immunogenic. Optimized conditions for dexr_LptD induction were 0.5 mM IPTG, 16 hours, TB (Terrific Broth) medium, 3% (v/v) ethanol, 28 ºC, OD600: 1-1.5 and 200 rpm. Purification was carried out under denaturating conditions on a small scale and we obtained 2.6 µg/mL of partially purified dexr_LptD. The Western Blot assay showed a positive reaction between the sera from patients with Carrión's Disease and dexr_LptD, which shows the antigenicity of dexr_LptD. CONCLUSIONS.: The dexr_LptD shows antigenicity both in silico and at the serological level, these results are the basis for further studies on vaccine candidates against Carrion's Disease.

OBJETIVO.: Evaluar in silico y a nivel serológico el potencial antigénico del dominio extracelular recombinante de la proteína de ensamblaje de lipopolisacáridos - D (LptD) de Bartonella bacilliformis (dexr_LptD). MATERIALES Y MÉTODOS.: Mediante el análisis in silico se realizó la selección de una proteína de B. bacilliformis con potencial antigénico e inmunogénico. El gen de la proteína seleccionada se clonó en Escherichia coli TOP10 y se expresó en Escherichia coli BL21 (DE3) pLysS. La proteína recombinante fue expresada usando isopropil-ß-D-1-tiogalactopiranósido (IPTG) y se optimizaron las condiciones de inducción. Por último, se purificó con resina Ni-IDA (His60 Ni Superflow) y se realizó un ensayo de Western Blot. RESULTADOS.: In silico, la proteína seleccionada fue LptD por estar localizada en la membrana externa y ser antigénica e inmunogénica. Las condiciones optimizadas para la inducción del dexr_LptD fueron 0,5 mM IPTG, 16 h, medio TB (Terrific Broth), etanol al 3% (v/v), 28 ºC, OD600: 1-1,5 y 200 r.p.m. La purificación se realizó en condiciones denaturantes a pequeña escala y se obtuvo 2,6 µg/mL de dexr_LptD parcialmente purificada. El ensayo de Western Blot mostró una reacción positiva entre los sueros provenientes de pacientes con la enfermedad de Carrión y dexr_LptD, ello evidencia la antigenicidad del dexr_LptD. CONCLUSIONES.: El dexr_LptD muestra antigenicidad in silico y a nivel serológico, estos resultados son base para posteriores estudios sobre candidatos vacunales contra la enfermedad de Carrión.

Genomic surveillance of the Lambda SARS-CoV-2 variant in a global phylogenetic context.

The massive sequencing of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and global genomic surveillance strategies allowed the detection of many variants of concern and interest. The variant of interest Lambda (C.37), which originated in South America, has been the most prevalent in Peru and Chile, but its dispersion in other continents still remains unknown. The current study aims to determine the phylogenetic relationship among C.37 isolates worldwide, focusing on spike mutations to understand the spread of Lambda in pandemics. A total of 7441 sequences identified as C.37 were downloaded from the GISAID database; local analysis was carried out to identify spike mutations and phylogenetic analysis was carried out to determine the rate of spread of the virus. Our results showed some spike mutations of Lambda that allowed us to detect small local outbreaks in different countries that occurred in the past and identify several clades that have not yet been designated. Although the lineage C.37 is not epidemiologically relevant in Europe or North America, the endemic behavior of this variant in Peru had a major impact on the second SARS-CoV-2 wave.

Confirmed Severe Acute Respiratory Syndrome Coronavirus 2 Reinfections After a Second Wave With Predominance of Lambda in Lima and Callao, Peru.

Background: Coronavirus disease 2019 (COVID-19) infection is a major public health problem in the world and reinfections are becoming more frequent. Our main objective was to describe the epidemiological, clinical, and genomic characteristics of the confirmed cases of reinfection by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the capital of Lima and Callao, Peru. Methods: We searched in the Peruvian laboratory information system from April 2020 up to May 2021, looking for cases having 2 positive molecular tests for SARS-CoV-2 with more than 90 days between them. We performed genomic sequencing to the available pairs of samples and described the clinical characteristics, epidemiological impact, and genomic analysis of the confirmed reinfections. Results: There were 1 694 164 people with a positive diagnostic test for SARS-CoV-2 in Lima/Callao during the study period. Of these, 1695 had 2 positive molecular tests with more than 90 days between them. Two hundred eleven had both samples available for genomic analysis according to our selection criteria, and these were retrieved and submitted to sequencing. Thirty cases were confirmed to be SARS-CoV-2 reinfections with 2 different lineages in the 2 episodes. The variant Lambda (C.37) was the most common during the second infection and accounted for 19 (63.3%) of the 30 cases. Conclusions: We report 30 cases of confirmed SARS-CoV-2 reinfections. The Lambda variant was the most common cause of the second infections, in concordance with its predominant circulation during Peru's second wave. This report describes the largest series of confirmed reinfections by SARS-CoV-2 in Latin America.We describe the epidemiological, clinical, and genomic characteristics of the confirmed cases of reinfection by severe acute respiratory syndrome coronavirus 2 in Lima and Callao, durante la segunda ola en Peru. The Lambda variant (C.37) was the most common cause of the second infections.

Reinfecciones por SARS-CoV-2 durante la segunda ola pandémica en Iquitos, Perú / SARS-CoV-2 reinfections during the second pandemic wave in Iquitos, Peru

Señor editor: La infección por SARS-CoV-2 ha ocasionado gran impacto en todo el mundo estimándose en más de 439 millones de casos y más de 5,9 millones de muertes. El Perú ha sido uno de los países en donde la mortalidad de su población ha descrito cifras muy elevadas llegando hasta una tasa de letalidad de 9.14%. Iquitos ha sido una de las ciudades más afectadas desde el inicio de la pandemia en el Perú, en donde se describió una seroprevalencia COVID-19 de 70% una de las más altas reportadas después de la primera ola pandémica de COVID-19. Es de esperar que esta seroprevalencia haya aumentado luego de la segunda ola. La duración de la inmunidad frente al SARS-CoV-2 ya sea por infección previa o por vacunación efectiva continúa siendo una de las interrogantes más importantes, en ese contexto, reportamos 4 casos de reinfecciones confirmadas en Iquitos Perú.

Dear Editor: SARS-CoV-2 infection has caused great impact worldwide, estimated at more than 439 million cases and more than 5.9 million deaths. Peru has been one of the countries where the mortality of its population has described very high figures reaching a case fatality rate of 9.14%. Iquitos has been one of the most affected cities since the beginning of the pandemic in Peru, where a COVID-19 seroprevalence of 70% was described, one of the highest reported after the first COVID-19 pandemic wave. It is to be expected that this seroprevalence has increased after the second wave. The duration of immunity against SARS-CoV-2 either by previous infection or by effective vaccination continues to be one of the most important questions, in that context, we report 4 cases of confirm reinfections in Iquitos Peru.

Clonamiento, expresión y seroreactividad de la proteína recombinante de ensamblaje de lipopolisacáridos - D (LptD) de Bartonella bacilliformis / Cloning, expression and seroreactivity of the recombinant lipopolysaccharide assembly protein - D (LptD) from Bartonella bacilliformis

RESUMEN Objetivo. Evaluar in silico y a nivel serológico el potencial antigénico del dominio extracelular recombinante de la proteína de ensamblaje de lipopolisacáridos - D (LptD) de Bartonella bacilliformis (dexr_LptD). Materiales y métodos. Mediante el análisis in silico se realizó la selección de una proteína de B. bacilliformis con potencial antigénico e inmunogénico. El gen de la proteína seleccionada se clonó en Escherichia coli TOP10 y se expresó en Escherichia coli BL21 (DE3) pLysS. La proteína recombinante fue expresada usando isopropil-β-D-1-tiogalactopiranósido (IPTG) y se optimizaron las condiciones de inducción. Por último, se purificó con resina Ni-IDA (His60 Ni Superflow) y se realizó un ensayo de Western Blot. Resultados. In silico, la proteína seleccionada fue LptD por estar localizada en la membrana externa y ser antigénica e inmunogénica. Las condiciones optimizadas para la inducción del dexr_LptD fueron 0,5 mM IPTG, 16 h, medio TB (Terrific Broth), etanol al 3% (v/v), 28 ºC, OD600: 1-1,5 y 200 r.p.m. La purificación se realizó en condiciones denaturantes a pequeña escala y se obtuvo 2,6 µg/mL de dexr_LptD parcialmente purificada. El ensayo de Western Blot mostró una reacción positiva entre los sueros provenientes de pacientes con la enfermedad de Carrión y dexr_LptD, ello evidencia la antigenicidad del dexr_LptD. Conclusiones. El dexr_LptD muestra antigenicidad in silico y a nivel serológico, estos resultados son base para posteriores estudios sobre candidatos vacunales contra la enfermedad de Carrión.

ABSTRACT Objective. To evaluate in silico and at the serological level the antigenic potential of the recombinant extracellular domain of the lipopolysaccharide assembly protein - D (LptD) of Bartonella bacilliformis (dexr_LptD). Materials and Methods. Through in silico analysis, we selected a B. bacilliformis protein with antigenic and immunogenic potential. The selected protein gene was cloned into Escherichia coli TOP10 and expressed in Escherichia coli BL21 (DE3) pLysS. Recombinant protein was expressed using isopropyl-β-D-1-thiogalactopyranoside (IPTG) and induction conditions were optimized. Finally, it was purified with Ni-IDA resin (His60 Ni Superflow) and a Western Blot assay was conducted. Results. In silico, the selected protein was LptD because it is located in the outer membrane and is antigenic and immunogenic. Optimized conditions for dexr_LptD induction were 0.5 mM IPTG, 16 hours, TB (Terrific Broth) medium, 3% (v/v) ethanol, 28 ºC, OD600: 1-1.5 and 200 rpm. Purification was carried out under denaturating conditions on a small scale and we obtained 2.6 μg/mL of partially purified dexr_LptD. The Western Blot assay showed a positive reaction between the sera from patients with Carrión's Disease and dexr_LptD, which shows the antigenicity of dexr_LptD. Conclusions. The dexr_LptD shows antigenicity both in silico and at the serological level, these results are the basis for further studies on vaccine candidates against Carrion's Disease.

Validación y evaluación de una prueba de RT-PCR en tiempo real in house para la detección de SARS-CoV-2 usando un gen específico RdRp y control endógeno GAPDH / Validation and evaluation of RT-PCR real time in house test to detection of SARS-CoV-2 using specific RdRp gene and GAPDH endogen control

RESUMEN Se validó y evaluó un método de RT-PCR en tiempo real usando cebadores y sondas específicas para los genes RdRP de SARS-CoV-2 y GAPDH de humanos; este último fue usado como control endógeno. Se evaluó la especificidad y sensibilidad; además, se evaluó otros parámetros como la robustez, la repetibilidad, reproducibilidad, comparabilidad y el límite de detección. La sensibilidad, especificidad, los valores predictivos positivo y negativo, la robustez, comparabilidad y la repetibilidad-reproducibilidad de la prueba de RT-PCR en tiempo real dúplex fue de 100%, con un límite de detección de 100 copias/µL, de acuerdo con los criterios de aceptación establecidos para validación del protocolo. Esta prueba estandarizada es una buena alternativa para el diagnóstico de COVID-19; además, la prueba fue aplicada de manera exitosa en personas sospechosas de la enfermedad permitiendo controlar el número de falsos negativos.

ABSTRACT The present work validated and evaluated a duplex real-time RT-PCR using specific primers and probes for genes RdRp from SARS-CoV-2 and GAPDH from humans; the latter was used as an endogenous control in all reactions. We evaluated the specificity, the sensitivity, the robustness, the reproducibility, the repeatability, the comparability, and the limit of detection. The predictive positive and negative values (PPV and PNV, respectively) and all the parameters evaluated using our duplex real-time RT-PCR was 100%. The detection limit was 100 copies/µL according to the acceptance criteria established for the validation of this protocol. Our duplex real-time RT-PCR demonstrated to be a good alternative for the diagnosis of COVID-19; in addition, this PCR was used adequately in suspicion of COVID-19, allowing it to control the number of false-negatives.

Genomic analysis reveals a rapid spread and predominance of lambda (C.37) SARS-COV-2 lineage in Peru despite circulation of variants of concern.

The pandemic generated by SARS-Cov-2 has caused a large number of cases and deaths in the world, but South America has been one of the continents that were most hard hit. The appearance of new variants causes concern because of the possibility that they may evade the protection generated by vaccination campaigns, their greater capacity to be transmitted, or their higher virulence. We analyzed the circulating variants in Peru after improving our Genomic Surveillance program. The results indicate a steep increase of the lambda lineage (C.37) until becoming predominant between January and April 2021, despite the cocirculation of other variants of concern or interest. Lambda lineage deserves close monitoring and could probably become a variant of concern in the near future.

Validation and evaluation of RT-PCR real time in house test to detection of SARS-CoV-2 using specific RdRp gene and GAPDH endogen control. / Validación y evaluación de una prueba de RT-PCR en tiempo real in house para la detección de SARS-CoV-2 usando un gen específico RdRp y control endógeno GAPDH.

The present work validated and evaluated a duplex real-time RT-PCR using specific primers and probes for genes RdRp from SARS-CoV-2 and GAPDH from humans; the latter was used as an endogenous control in all reactions. We evaluated the specificity, the sensitivity, the robustness, the reproducibility, the repeatability, the comparability, and the limit of detection. The predictive positive and negative values (PPV and PNV, respectively) and all the parameters evaluated using our duplex real-time RT-PCR was 100%. The detection limit was 100 copies/µL according to the acceptance criteria established for the validation of this protocol. Our duplex real-time RT-PCR demonstrated to be a good alternative for the diagnosis of COVID-19; in addition, this PCR was used adequately in suspicion of COVID-19, allowing it to control the number of false-negatives.

Se validó y evaluó un método de RT-PCR en tiempo real usando cebadores y sondas específicas para los genes RdRP de SARS-CoV-2 y GAPDH de humanos; este último fue usado como control endógeno. Se evaluó la especificidad y sensibilidad; además, se evaluó otros parámetros como la robustez, la repetibilidad, reproducibilidad, comparabilidad y el límite de detección. La sensibilidad, especificidad, los valores predictivos positivo y negativo, la robustez, comparabilidad y la repetibilidad-reproducibilidad de la prueba de RT-PCR en tiempo real dúplex fue de 100%, con un límite de detección de 100 copias/µL, de acuerdo con los criterios de aceptación establecidos para validación del protocolo. Esta prueba estandarizada es una buena alternativa para el diagnóstico de COVID-19; además, la prueba fue aplicada de manera exitosa en personas sospechosas de la enfermedad permitiendo controlar el número de falsos negativos.

Neutralization of the lethal activity from Bothrops atrox venom by hyperimune llama serum (Lama glama). / Neutralización de la actividad letal del veneno de serpiente Bothrops atrox por suero hiperinmune de llama (Lama glama).

OBJECTIVES: To evaluate the capacity of the hyperimmune llama serum (Lama glama) to neutralize the lethal activity of Bothrops atrox venom in laboratory mice. MATERIALS AND METHODS: Mean lethal dose (LD50) was calculated from a Bothrops atrox venom sample pool from Peru. The antibody titers were measured by ELISA assay; and the immune serum neutralization potency was measured by calculating the mean effective dose (ED50) during the immunization period. RESULTS: The venom's LD50 was 3.96 µg/g; similar to what was found in other studies about Bothrops atrox carried out in Peru. The titers of antibodies against the venom increased rapidly in the llama, demonstrating a fast immune response; however, the neutralization capacity increased slowly and required several doses and immunization reinforcements, obtaining a ED50 of 3.30 µL/g mouse and a neutralization potency of 3.6 mg/mL after 15 immunizations. CONCLUSIONS: The hyperimmune llama serum is able to neutralize the lethality of the Bothrops atrox venom from Peru in laboratory mice.

OBJETIVOS: Evaluar la capacidad del suero hiperinmune de llama (Lama glama) para neutralizar la letalidad del veneno de la serpiente Bothrops atrox en ratones de laboratorio. MATERIALES Y MÉTODOS: Se calculó la dosis letal media (DL50) de un pool de venenos de serpientes de Bothrops atrox de Perú, y se midieron los títulos de anticuerpos por ensayo ELISA; así como la potencia de neutralización del suero inmune por el cálculo de la dosis efectiva media (DE50) durante el periodo de inmunización. RESULTADOS: La DL50 del veneno fue de 3,96 µg/g, similar a otros trabajos realizados en Bothrops atrox en Perú. Los títulos de anticuerpos contra el veneno se incrementan rápidamente en la llama mostrando una rápida respuesta inmune; sin embargo, la capacidad de neutralización se incrementa más lentamente y requiere de varias dosis y refuerzos de las inmunizaciones alcanzado una DE50 de 3,30 µL/g ratón y una potencia de neutralización 3,6 mg/mL después de 15 inmunizaciones. CONCLUSIONES: El suero hiperinmune de llama es capaz de neutralizar la letalidad del veneno de la serpiente Bothrops atrox de Perú en ratones de laboratorio.

Neutralización de la actividad letal del veneno de serpiente Bothrops atrox por suero hiperinmune de llama (Lama glama) / Neutralization of the lethal activity from Bothrops atrox venom by hyperimune llama serum (Lama glama)

RESUMEN Objetivos: Evaluar la capacidad del suero hiperinmune de llama (Lama glama) para neutralizar la letalidad del veneno de la serpiente Bothrops atrox en ratones de laboratorio. Materiales y métodos: Se calculó la dosis letal media (DL50) de un pool de venenos de serpientes de Bothrops atrox de Perú, y se midieron los títulos de anticuerpos por ensayo ELISA; así como la potencia de neutralización del suero inmune por el cálculo de la dosis efectiva media (DE50) durante el periodo de inmunización. Resultados: La DL50 del veneno fue de 3,96 µg/g, similar a otros trabajos realizados en Bothrops atrox en Perú. Los títulos de anticuerpos contra el veneno se incrementan rápidamente en la llama mostrando una rápida respuesta inmune; sin embargo, la capacidad de neutralización se incrementa más lentamente y requiere de varias dosis y refuerzos de las inmunizaciones alcanzado una DE50 de 3,30 µL/g ratón y una potencia de neutralización 3,6 mg/mL después de 15 inmunizaciones. Conclusiones: El suero hiperinmune de llama es capaz de neutralizar la letalidad del veneno de la serpiente Bothrops atrox de Perú en ratones de laboratorio.

ABSTRACT Objectives: To evaluate the capacity of the hyperimmune llama serum (Lama glama) to neutralize the lethal activity of Bothrops atrox venom in laboratory mice. Materials and methods: Mean lethal dose (LD50) was calculated from a Bothrops atrox venom sample pool from Peru. The antibody titers were measured by ELISA assay; and the immune serum neutralization potency was measured by calculating the mean effective dose (ED50) during the immunization period. Results: The venom's LD50 was 3.96 μg/g; similar to what was found in other studies about Bothrops atrox carried out in Peru. The titers of antibodies against the venom increased rapidly in the llama, demonstrating a fast immune response; however, the neutralization capacity increased slowly and required several doses and immunization reinforcements, obtaining a ED50 of 3.30 μL/g mouse and a neutralization potency of 3.6 mg/mL after 15 immunizations. Conclusions: The hyperimmune llama serum is able to neutralize the lethality of the Bothrops atrox venom from Peru in laboratory mice.

Development of Nanobodies Against Hemorrhagic and Myotoxic Components of Bothrops atrox Snake Venom.

Snake envenoming is a globally neglected public health problem. Antivenoms produced using animal hyperimmune plasma remain the standard therapy for snakebites. Although effective against systemic effects, conventional antivenoms have limited efficacy against local tissue damage. In addition, potential hypersensitivity reactions, high costs for animal maintenance, and difficulties in obtaining batch-to-batch homogeneity are some of the factors that have motivated the search for innovative and improved therapeutic products against such envenoming. In this study, we have developed a set of nanobodies (recombinant single-domain antigen-binding fragments from camelid heavy chain-only antibodies) against Bothrops atrox snake venom hemorrhagic and myotoxic components. An immune library was constructed after immunizing a Lama glama with whole venom of B. atrox, from which nanobodies were selected by phage display using partially purified hemorrhagic and myotoxic proteins. Biopanning selections retrieved 18 and eight different nanobodies against the hemorrhagic and the myotoxic proteins, respectively. In vivo assays in mice showed that five nanobodies inhibited the hemorrhagic activity of the proteins; three neutralized the hemorrhagic activity of whole B. atrox venom, while four nanobodies inhibited the myotoxic protein. A mixture of the anti-hemorrhagic and anti-myotoxic nanobodies neutralized the local tissue hemorrhage and myonecrosis induced by the whole venom, although the nanobody mixture failed to prevent the venom lethality. Nevertheless, our results demonstrate the efficacy and usefulness of these nanobodies to neutralize important pathologies of the venom, highlighting their potential as innovative therapeutic agents against envenoming by B. atrox, a viperid species causing many casualties in South America.

Near-Complete Genome Sequence of a 2019 Novel Coronavirus (SARS-CoV-2) Strain Causing a COVID-19 Case in Peru.

A near-complete genome sequence was obtained for a novel coronavirus (SARS-CoV-2) strain obtained from an oropharyngeal swab from a Peruvian patient with coronavirus syndrome (COVID-19) who had contact with an individual who had returned to Peru from travel to Italy.