Mariana Leguia

• Ph.D.
• Managing Director at Pontifical Catholic University of Peru

Given the local and systemic adverse reactions associated with whole cell pertussis vaccines combined with diphtheria and tetanus toxoids (DTP), acellular pertussis vaccines combined with the same toxoids (DTaP) were developed in the 1990s. In comparison to DTP, DTaP vaccines demonstrated reduced reactogenicity and equivalent or improved immunogeni...

Dengue virus (DENV) is currently causing epidemics of unprecedented scope in endemic settings and expanding to new geographical areas. It is therefore critical to track this virus using genomic surveillance. However, the complex patterns of viral genomic diversity make it challenging to use the existing genotype classification system. Here, we prop...

Dengue virus (DENV) is currently causing epidemics of unprecedented scope in endemic settings and expanding to new geographical areas. It is therefore critical to track this virus using genomic surveillance. However, the complex patterns of viral genomic diversity make it challenging to use the existing genotype classification system. Here we propo...

Highly pathogenic avian influenza (HPAI) A/H5N1 viruses (lineage 2.3.4.4b) are rapidly invading the Americas, threatening wildlife, poultry, and potentially evolving into the next global pandemic. In November 2022 HPAI arrived in Peru, triggering massive pelican and sea lion die-offs. We report genomic characterization of HPAI/H5N1 in five species...

Dengue virus (DENV) transmission from humans to mosquitoes is a poorly documented, but critical component of DENV epidemiology. Magnitude of viremia is the primary determinant of successful human-to-mosquito DENV transmission. People with the same level of viremia, however, can vary in their infectiousness to mosquitoes as a function of other facto...

RNA purification and cDNA synthesis represents the starting point for molecular analyses of snake venom proteins-enzymes. Usually, the sacrifice of snakes is necessary for venom gland extraction to identify protein-coding transcripts; however, the venom can be used as a source of transcripts. Although there are methods for obtaining RNA from venom,...

Highly pathogenic avian influenza (HPAI) A/H5N1 viruses (lineage 2.3.4.4b) are rapidly invading the Americas, threatening wildlife, poultry, and potentially evolving into the next global pandemic. In November 2022, HPAI arrived in Peru, where massive pelican and sea lion die-offs are still underway. We report complete genomic characterization of HP...

Current knowledge of dengue virus (DENV) transmission provides only a partial understanding of a complex and dynamic system yielding a public health track record that has more failures than successes. An important part of the problem is that the foundation for contemporary interventions includes a series of longstanding, but untested, assumptions b...

Over half the world’s population is at risk for viruses transmitted by Aedes mosquitoes, such as dengue and Zika. The primary vector, Aedes aegypti , thrives in urban environments. Despite decades of effort, cases and geographic range of Aedes -borne viruses (ABVs) continue to expand. Rigorously proven vector control interventions that measure prot...

Peru celebrates 200 years of independence in 2021. Over this period of independent life, and despite the turbulent socio-political scenarios, from internal armed conflict to economic crisis to political instability over the last 40 years, Peru has experienced major changes on its epidemiological and population health profile. Major advancements in...

Despite several reports worldwide documenting the presence of Rickettsia asembonensis in samples derived from ectoparasites, animals and more recently humans, genomic information of these specimens remains scarce, and when available, is usually limited to small genomic fragments of limited value. We generated complete sequences for two conserved (1...

Background: Over half the world's population is at risk for viruses transmitted by Aedes mosquitoes, such as, dengue and Zika. The primary vector, Aedes aegypti, thrives in urban environments. Despite decades of effort, cases and the geographic range of Aedes-borne viruses (ABV) continue to expand. To date, there are no rigorously proven vector con...

Minority Gene Expression Profiling (MGEP) refers to a scenario where the expression profiles of specific genes of interest are concentrated in a small cellular pool that is embedded within a larger, non-expressive pool. An example of this is the analysis of disease-related genes within sub-populations of blood or biopsied tissues. These systems are...

This brief report serves as an introduction to a supplement of the Journal of Infectious Diseases entitled "Next-Generation Sequencing (NGS) Technologies to Advance Global Infectious Disease Research." We briefly discuss the history of NGS technologies and describe how the techniques developed during the past 40 years have impacted our understandin...

Rickettsia and Leptospira spp. are under-recognized causes of acute febrile disease worldwide. Rickettsia species are often placed into the spotted fever group rickettsiae (SFGR) and typhus group rickettsiae (TGR). We explored the antibody prevalence among humans for these two groups of rickettsiae in four regions of Peru (Lima, Cusco, Puerto Maldo...

While studying respiratory infections in Peru, we identified Venezuelan equine encephalitis virus (VEEV) in a nasopharyngeal swab, indicating that this alphavirus can be present in human respiratory secretions. Because VEEV may be infectious when aerosolized, our finding is relevant for the management of VEEV-infected patients and for VEEV transmis...

While studying rickettsial infections in Peru, we detected Rickettsia asembonensis in fleas from domestic animals. We characterized 5 complete genomic regions (17kDa, gltA, ompA, ompB, and sca4) and conducted multilocus sequence typing and phylogenetic analyses. The molecular isolate from Peru is distinct from the original R. asembonensis strain fr...

Reference sequences used for phylogenetic analysis of Peru Rickettsia asembonensis molecular isolate.

Background: The transmission dynamics of human metapneumovirus (HMPV) in tropical countries remain unclear. Further understanding of the genetic diversity of the virus could aid in HMPV vaccine design and improve our understanding of respiratory virus transmission dynamics in low- and middle-income countries. Materials & methods: We examined the...

Using a large, passive, febrile surveillance program in Iquitos, Peru, we retrospectively tested human blood specimens for scrub typhus group orientiae by ELISA, immu-nofluorescence assay, and PCR. Of 1,124 participants, 60 (5.3%) were seropositive, and 1 showed evidence of recent active infection. Our serologic data indicate that scrub typhus is p...

We have developed methods for full-genome sequencing of Zika viruses (ZIKVs) based on a targeted amplification approach. We used alignments of publicly available complete genome data to design a primer set that selectively amplifies ZIKVs. The approach includes amplification strategies for templates present at both high- and low-copy number, and PC...

The genus Bartonella contains >40 species, and an increasing number of these Bartonella species are being implicated in human disease. One such pathogen is Bartonella ancashensis, which was isolated in blood samples from 2 patients living in Caraz, Peru, during a clinical trial of treatment for bartonellosis. Three B. ancashensis strains were analy...

Using a large, passive, clinic-based surveillance program in Iquitos, Peru, we characterized the prevalence of rickettsial infections among undifferentiated febrile cases and obtained evidence of pathogen transmission in potential domestic reservoir contacts and their ectoparasites. Blood specimens from humans and animals were assayed for spotted f...

Univariate analysis comparing participants with co-infections to those without (DOCX)

We report optimized workflows for full-genome sequencing of dengue viruses (DENVs) 1-4. Based on alignments of publicly available complete genomes we modified and expanded existing primers sets to amplify DENV genotypes that were previously difficult or impossible to sequence. We also report improvements to streamline laboratory handling, including...

While studying respiratory infections of unknown etiology we detected Saffold virus in an oropharyngeal swab collected from a two-year-old female suffering from diarrhea and respiratory illness. The full viral genome recovered by deep sequencing showed 98% identity to a previously described Saffold strain isolated in Japan. Phylogenetic analysis co...

Here we present the complete genome sequence of Bartonella ancashensis strain 20.00, isolated from the blood of a Peruvian patient with verruga peruana, known as Carrion’s disease. Bartonella ancashensis is a Gram-negative bacillus, phylogenetically most similar to Bartonella bacilliformis , the causative agent of Oroya fever and verruga peruana.

There is an increasing role for bioinformatic and phylogenetic analysis in tropical medicine research. However, scientists working in low- and middle-income regions may lack access to training opportunities in these methods. To help address this gap, we offered a 5-day intensive bioinformatics workshop in Lima, Peru and present the syllabus here fo...

Three Bartonella isolates were recovered from the blood of two patients enrolled in a clinical trial for the treatment of chronic stage Bartonella bacilliformis infection (verruga peruana) in Caraz, Ancash, Peru. The isolates were initially characterized by sequencing a fragment of the gltA gene, and found to be disparate from B. bacilliformis. The...

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A recently described clinical isolate, “Candidatus Bartonella ancashi,” was obtained from a blood sample of a patient presenting with verruga peruana in the Ancash region of Peru. This sample and a second isolate obtained 60 days later from the same patient were molecularly typed using multilocus sequence typing (MLST) and multispacer sequence typi...

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There is growing demand for robust DNA assembly strategies to quickly and accurately fabricate genetic circuits for synthetic biology. One application of this technology is reconstitution of multi-gene assemblies. Here, we integrate a new software tool chain with 2ab assembly and show that it is robust enough to generate 528 distinct composite part...

Basic parts used for BglBricks-based 2ab assembly of bi-cistronic operons of Mediator complex.

Heat maps of protein concentration values obtained via indirect ELISAs. Maps include all possible tags and tagging combinations in order to visualize expression patterns for each epitope-tagged ORF in all 528 bi-cistronic operons. The top map illustrates patterns of expression for Med7 located in the first ORF position and should be read from left...

Antibodies used in ELISA assays.

Complete data set of protein concentration values obtained via indirect ELISAs from all 528 bi-cistronic operons.

We describe discovery of two enteroviruses detected in nasopharyngeal samples obtained from subjects with respiratory disease in Peru. Phylogenetic analysis indicated that both viruses belong to a clade within the human enterovirus C species, which includes the recently characterized human enteroviruses 109 and 104. Members of this clade have under...

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A metamorphic life-history is present in the majority of animal phyla. This developmental mode is particularly prominent among marine invertebrates with a bentho-planktonic life cycle, where a pelagic larval form transforms into a benthic adult. Metamorphic competence (the stage at which a larva is capable to undergo the metamorphic transformation...

Appendix 1 We identified and cloned the full length cDNA from the sea urchin histidine decarboxylase (HDC) gene based on the annotation present in NCBI (XP_789367). We then used the protein sequence for phylogenetic analysis and comparison with other animal HDCs as well as three dopa decarboxylase genes (Ddc). HDC from S. purpuratus clearly falls w...

Appendix 2 Neighbor joining phylogenetic analysis of sea urchin histamine receptor predictions in comparison to human H1, H2, H3 and H4 receptor. We identified all histamine-receptor like genes from SpBase (http://sugp.caltech.edu/SpBase/) and aligned them with human H1, H2 and H3 receptor. Phylogenetic analysis suggests that all three receptor typ...

The histamine H1 receptor is a seven transmembrane G protein-coupled receptor thought to participate in the release of calcium from intracellular stores during egg activation. Prior to fertilization, the receptor is present exclusively at the surface of the egg. Shortly after fertilization, however, the receptor is endocytosed. To follow this trans...

The primary bottleneck in synthetic biology research today is the construction of physical DNAs, a process that is often expensive, time-consuming, and riddled with cloning difficulties associated with the uniqueness of each DNA sequence. We have developed a series of biological and computational tools that lower existing barriers to automation and...

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Synthetic biological systems are currently created by an ad-hoc, iterative process of specification, design, and assembly. These systems would greatly benefit from a more formalized and rigorous specification of the desired system components as well as constraints on their composition. Therefore, the creation of robust and efficient design flows an...

Standard biological parts, such as BioBricks parts, provide the foundation for a new engineering discipline that enables the design and construction of synthetic biological systems with a variety of applications in bioenergy, new materials, therapeutics, and environmental remediation. Although the original BioBricks assembly standard has found wide...

The BglBricks standard has been developed as an alternative to the original XbaI/SpeI standard primarily to provide a solution to the issue of generating translational-fusion parts. When joined together in a standard assembly reaction, BglBricks parts contain a scar sequence of GGATCT between the two parts. This sequence conveniently translates as...

The signaling networks controlling calcium release and cortical granule exocytosis at fertilization are complex and multilayered, providing various points for regulatory input and quality control. Though it is clear that many of the mechanisms leading to both calcium release and cortical granule exocytosis are conserved, a great deal of variability...

Sperm fusion with the egg initiates a signaling cascade that releases intracellular calcium (Ca(i) (2+)) from the endoplasmic reticulum (ER). In sea urchins, Ca(2+) is released as a single, large transient via two distinct pathways. The first depends on inositol 1,4,5-triphosphate (IP(3)) production and triggers the initial phase of Ca(2+) release,...

Cortical granules are stimulus-dependent secretory vesicles found in the egg cortex of most vertebrates and many invertebrates. Upon fertilization, an increase in intracellular calcium levels triggers cortical granules to exocytose enzymes and structural proteins that permanently modify the extracellular surface of the egg to prevent polyspermy. Sy...

Regulated secretion is mediated by SNAREs (soluble NSF attachment receptors) and their regulators and effectors, which include the SM (sec1/munc18) family of proteins. Homologs of the SNAREs have been identified in sea urchins, associated with cortical granule exocytosis at fertilization, with membranes of the cleavage furrow, and in secretory cell...

The human Ku86 gene and an isoform, KARP-1 (Ku86 autoantigen related protein-1), encode overlapping, but differentially regulated, transcripts. Ku86 is constitutively transcribed at high levels and, although it plays a seminal role in DNA double-strand break repair, its expression is not induced by DNA damage. KARP-1, in contrast, is expressed cons...