O2 Saturation Predicted the ICU Stay of COVID-19 Patients in a Hospital at Altitude: A Low-Cost Tool for Post-Pandemic.

Background and Objectives: Patients at high altitudes with COVID-19 may experience a decrease in their partial oxygen saturation (PO2S) levels. The objective was to assess the association between PO2S and intensive care unit (ICU) stay in patients at high altitudes with COVID-19. Materials and Methods: Clinical records of 69 COVID-19 patients (36% women) admitted to the ICU were analyzed. Median values were considered for intra-group categories ("≤11 days" and ">11 days" in the ICU) and for PO2S height categories ("<90%" and "≥90%"). Logistic regression and linear regression models adjusted for confounding variables were used. Results: Patients with >11 days in the ICU had 84% lower odds of having a PO2S ≥ 90% (OR: 0.16 [CI: 0.02, 0.69], p = 0.005) compared to those with ≤11 days in the ICU. An increase in PO2S by 1% reduced ICU stay by 0.22 days (ß: -0.22 [CI: -0.33, -0.11], p < 0.001), potentially leading to a reduction of up to 1.44 days. Conclusions: PO2S is a crucial factor in estimating ICU stays for COVID-19 patients at high altitudes and serves as an accessible and cost-effective measure. It should be used in infected patients to complement the prognosis of post-pandemic ICU stay.

Proteome of the Triatomine Digestive Tract: From Catalytic to Immune Pathways; Focusing on Annexin Expression.

Rhodnius prolixus, Panstrongylus megistus, Triatoma infestans, and Dipetalogaster maxima are all triatomines and potential vectors of the protozoan Trypanosoma cruzi responsible for human Chagas' disease. Considering that the T. cruzi's cycle occurs inside the triatomine digestive tract (TDT), the analysis of the TDT protein profile is an essential step to understand TDT physiology during T. cruzi infection. To characterize the protein profile of TDT of D. maxima, P. megistus, R. prolixus, and T. infestans, a shotgun liquid chromatography-tandem mass spectrometry (LC-MS/MS) approach was applied in this report. Most proteins were found to be closely related to metabolic pathways such as gluconeogenesis/glycolysis, citrate cycle, fatty acid metabolism, oxidative phosphorylation, but also to the immune system. We annotated this new proteome contribution gathering it with those previously published in accordance with Gene Ontology and KEGG. Enzymes were classified in terms of class, acceptor, and function, while the proteins from the immune system were annotated by reference to the pathways of humoral response, cell cycle regulation, Toll, IMD, JNK, Jak-STAT, and MAPK, as available from the Insect Innate Immunity Database (IIID). These pathways were further subclassified in recognition, signaling, response, coagulation, melanization and none. Finally, phylogenetic affinities and gene expression of annexins were investigated for understanding their role in the protection and homeostasis of intestinal epithelial cells against the inflammation.

In vitro Trypanocidal Activity, Genomic Analysis of Isolates, and in vivo Transcription of Type VI Secretion System of Serratia marcescens Belonging to the Microbiota of Rhodnius prolixus Digestive Tract.

Serratia marcescens is a bacterium with the ability to colonize several niches, including some eukaryotic hosts. S. marcescens have been recently found in the gut of hematophagous insects that act as parasite vectors, such as Anopheles, Rhodnius, and Triatoma. While some S. marcescens strains have been reported as symbiotic or pathogenic to other insects, the role of S. marcescens populations from the gut microbiota of Rhodnius prolixus, a vector of Chagas' disease, remains unknown. Bacterial colonies from R. prolixus gut were isolated on BHI agar. After BOX-PCR fingerprinting, the genomic sequences of two isolates RPA1 and RPH1 were compared to others S. marcescens from the NCBI database in other to estimate their evolutionary divergence. The in vitro trypanolytic activity of these two bacterial isolates against Trypanosoma cruzi (DM28c clone and Y strain) was assessed by microscopy. In addition, the gene expression of type VI secretion system (T6SS) was detected in vivo by RT-PCR. Comparative genomics of RPA1 and RPH1 revealed, besides plasmid presence and genomic islands, genes related to motility, attachment, and quorum sensing in both genomes while genes for urea hydrolysis and type II secretion system (T2SS) were found only in the RPA1 genome. The in vitro trypanolytic activity of both S. marcescens strains was stronger in their stationary phases of growth than in their exponential ones, with 65-70 and 85-90% of epimastigotes (Dm28c clone and Y strain, respectively) being lysed after incubation with RPA1 or RPH1 in stationary phase. Although T6SS transcripts were detected in guts up to 40 days after feeding (DAF), R. prolixus morbidity or mortality did not appear to be affected. In this report, we made available two trypanolytic S. marcescens strains from R. prolixus gut to the scientific community together with their genomic sequences. Here, we describe their genomic features with the purpose of bringing new insights into the S. marcescens adaptations for colonization of the specific niche of triatomine guts. This study provides the basis for a better understanding of the role of S. marcescens in the microbiota of R. prolixus gut as a potential antagonist of T. cruzi in this complex system.

Biology of Triatoma sherlocki (Hemiptera: Reduviidae) Under Laboratory Conditions: Biological Cycle and Resistance to Starvation.

Triatoma sherlocki Papa, Jurberg, Carcavallo, Cerqueira & Barata was described in 2002, based on specimens caught in the wild in the municipality of Gentio do Ouro, Bahia, Brazil. In 2009, nymphs and adults were detected inside homes and sylvatic specimens were positive for Trypanosoma cruzi (Chagas). No information on the bionomics of T. sherlocki exists, although such data are considered essential to estimate its vector and colonization potential in domestic environments. Herein, the biological cycle of T. sherlocki was studied based on 123 eggs, with nymphs and adults fed on Mus musculus (Linnaeus). Nymphal development time phases, number of meals consumed, and stage-specific mortality rates were analyzed. Survival time under starvation conditions was measured between ecdysis and death among 50 nymphs (first to fifth instar) and 50 male and female adults. The median development time from egg to adult was 621.0 (CI: 489-656) d. The number of meals consumed ranged from 1 to 20 for nymphs of the first to fifth instar. The fifth instar showed the greatest resistance to starvation, with a mean of 156.5 d. The high number of meals consumed by T. sherlocki favored infection with and transmission of T. cruzi. The full development of this species under laboratory conditions with a low mortality rate indicates that this vector presents biological characteristics that may contribute to its adaptation to artificial human ecotopes. Its high resistance to starvation emphasizes the importance of entomological surveillance for this species.

Everybody loves sugar: first report of plant feeding in triatomines.

BACKGROUND: Triatomines, which are the vectors of Trypanosoma cruzi, have been considered to be exclusive blood feeders for more than 100 years, since the discovery of Chagas disease. METHODS: We offered artificial sugar meals to the laboratory model-insect Rhodnius prolixus, which is considered a strict haematophagous insect. We registered feeding by adding colorant to sugar meals. To assess putative phytophagy, fruits of the tomato Solanum lycopersicum were offered to R. prolixus and the presence of tomato DNA was assessed in the insects using PCR. We also assessed longevity, blood feeding and urine production of fruit-exposed triatomines and control insects. RESULTS: All instars of R. prolixus ingested sugar from artificial sugar meals in laboratory conditions. First instar R. prolixus ingested plant tissue from S. lycopersicum fruits, and this increased the amount of blood ingested and urine excreted. Decreased mortality was also observed after blood feeding. Exposure to S. lycopersicum increased longevity and reduced weight loss caused by desiccation. CONCLUSIONS: We describe here the first report of sugar feeding and phytophagy in a species that was considered to be a strict blood-feeder for over a century. We suggest that local plants might be not merely shelters for insects and vertebrate hosts as previously described, but may have a nutritional role for the maintenance of the triatomine vectors. The description of sugar and plant meals in triatomines opens new perspectives for the study and control of Chagas Disease.

Phenotypic variability confirmed by nuclear ribosomal DNA suggests a possible natural hybrid zone of Triatoma brasiliensis species complex.

Triatoma brasiliensis macromelasoma occurs in Pernambuco state, Brazil, which is situated between the distribution areas of Triatoma brasiliensis brasiliensis (north) and Triatoma juazeirensis (south). T. b. macromelasoma displays greater variations in its chromatic phenotype than either T. b. brasiliensis or T. juazeirensis, and patterns reminiscent of one or the other. Experimental crosses from each of these members of the T. brasiliensis species complex generated fertile offspring suggesting that viable hybrids could be present in nature, despite their significant genetic distances. Considering the geographical position of occurrence of the T. b. macromelasoma (in Pernambuco) it was proposed to be an area capable of supporting natural hybridization between T. b. brasiliensis and T. juazeirensis. Since phenotypic variability is expected, this study investigated the existence of intermediate chromatic phenotypes for T. b. macromelasoma in various locations in areas between the T. b. brasiliensis and T. juazeirensis occurrences. Thirteen different color patterns were for the first time characterized and nine of those displayed intermediate phenotypes. Molecular analysis performed using ribosomal DNA intergenic region, grouped all within the T. brasiliensis complex. The intermediate chromatic phenotypes, molecular analysis and experimental crosses all support the distinction of a zone of hybridization that gave rise to the T. b. macromelasoma through homoploidal evolution.

Characterization of the microbiota in the guts of Triatoma brasiliensis and Triatoma pseudomaculata infected by Trypanosoma cruzi in natural conditions using culture independent methods.

BACKGROUND: Chagas disease is caused by Trypanosoma cruzi, which is transmitted by triatomine vectors. The northeastern region of Brazil is endemic for Chagas disease and has the largest diversity of triatomine species. T. cruzi development in its triatomine vector depends on diverse factors, including the composition of bacterial gut microbiota. METHODS: We characterized the triatomines captured in the municipality of Russas (Ceará) by sequencing the cytochrome c oxidase subunit I (COI) gene. The composition of the bacterial community in the gut of peridomestic Triatoma brasiliensis and Triatoma pseudomaculata was investigated using culture independent methods based on the amplification of the 16S rRNA gene by polymerase chain reaction (PCR), denaturing gradient gel electrophoresis (DGGE), DNA fragment cloning, Sanger sequencing and 454 pyrosequencing. Additionally, we identified TcI and TcII types of T. cruzi by sequencing amplicons from the gut metagenomic DNA with primers for the mini-exon gene. RESULTS: Triatomines collected in the peridomestic ecotopes were diagnosed as T. pseudomaculata and T. brasiliensis by comparing their COI sequence with GenBank. The rate of infection by T. cruzi in adult triatomines reached 80% for T. pseudomaculata and 90% for T. brasiliensis. According to the DNA sequences from the DGGE bands, the triatomine gut microbiota was primarily composed of Proteobacteria and Actinobacteria. However, Firmicutes and Bacteroidetes were also detected, although in much lower proportions. Serratia was the main genus, as it was encountered in all samples analyzed by DGGE and 454 pyrosequencing. Members of Corynebacterinae, a suborder of the Actinomycetales, formed the next most important group. The cloning and sequencing of full-length 16S rRNA genes confirmed the presence of Serratia marcescens, Dietzia sp., Gordonia terrae, Corynebacterium stationis and Corynebacterium glutamicum. CONCLUSIONS: The study of the bacterial microbiota in the triatomine gut has gained increased attention because of the possible role it may play in the epidemiology of Chagas disease by competing with T. cruzi. Culture independent methods have shown that the bacterial composition of the microbiota in the guts of peridomestic triatomines is made up by only few bacterial species.

Rhodnius prolixus interaction with Trypanosoma rangeli: modulation of the immune system and microbiota population.

BACKGROUND: Trypanosoma rangeli is a protozoan that infects a variety of mammalian hosts, including humans. Its main insect vector is Rhodnius prolixus and is found in several Latin American countries. The R. prolixus vector competence depends on the T. rangeli strain and the molecular interactions, as well as the insect's immune responses in the gut and haemocoel. This work focuses on the modulation of the humoral immune responses of the midgut of R. prolixus infected with T. rangeli Macias strain, considering the influence of the parasite on the intestinal microbiota. METHODS: The population density of T. rangeli Macias strain was analysed in different R. prolixus midgut compartments in long and short-term experiments. Cultivable and non-cultivable midgut bacteria were investigated by colony forming unit (CFU) assays and by 454 pyrosequencing of the 16S rRNA gene, respectively. The modulation of R. prolixus immune responses was studied by analysis of the antimicrobial activity in vitro against different bacteria using turbidimetric tests, the abundance of mRNAs encoding antimicrobial peptides (AMPs) defensin (DefA, DefB, DefC), prolixicin (Prol) and lysozymes (LysA, LysB) by RT-PCR and analysis of the phenoloxidase (PO) activity. RESULTS: Our results showed that T. rangeli successfully colonized R. prolixus midgut altering the microbiota population and the immune responses as follows: 1 - reduced cultivable midgut bacteria; 2 - decreased the number of sequences of the Enterococcaceae but increased those of the Burkholderiaceae family; the families Nocardiaceae, Enterobacteriaceae and Mycobacteriaceae encountered in control and infected insects remained the same; 3 - enhanced midgut antibacterial activities against Serratia marcescens and Staphylococcus aureus; 4 - down-regulated LysB and Prol mRNA levels; altered DefB, DefC and LysA depending on the infection (short and long-term); 5 - decreased PO activity. CONCLUSION: Our findings suggest that T. rangeli Macias strain modulates R. prolixus immune system and modifies the natural microbiota composition.

Cross-mating experiments detect reproductive compatibility between Triatoma sherlocki and other members of the Triatoma brasiliensis species complex.

Phylogenetic approaches based on mitochondrial DNA variation (fragments of Cyt B and 16S ribosomal RNA) have revealed Triatoma sherlocki as the most recent species addition to the Triatoma brasiliensis species complex; a monophyletic group which includes T. brasiliensis, Triatoma melanica, and Triatoma juazeirensis. T. sherlocki is the most differentiated among all species of this complex: it is unable to fly, possesses longer legs than the other members, and has reddish tonality in some parts of its exochorion. We question whether these species are reproductively compatible because of this pronounced morphological differentiation, and therefore, we present a series of cross breeding experiments that test compatibility between T. sherlocki and other members of the T. brasiliensis complex. We extended our analyses to include crosses between T. sherlocki and Triatoma lenti, because the latter has been suggested as a possible member of this complex. T. sherlocki male×T. lenti female pairs failed to produce hybrids. All other crosses of T. sherlocki and members of T. brasiliensis species complex, as well as backcrosses, produced viable offspring through the third generation. This study stresses the importance of searching for the features that may isolate members of the T. brasiliensis species complex.

Dispersion capacity of Triatoma sherlocki, Triatoma juazeirensis and laboratory-bred hybrids.

Flight dispersion is recognized as one of the most important mechanisms for triatomine house infestation. Triatoma sherlocki and T. juazeirensis are closely related species that occur within the same ecotope and their possible reproductive boundaries are unknown. T. sherlocki has shorter wings than T. juazeirensis; a characteristic that possibly implies in reduced flight dispersion, however, this species has been found to invade and colonize homes in Bahia, Brazil. Here, we tested the flight potential of T. sherlocki, compared to that of T. juazeirensis and laboratory-bred hybrids. Insects were kept in an apparatus designed to distinguish flyers from nonflyers. Fifty-one and 53% of T. juazeirensis and hybrids were flyers respectively, whereas no T. sherlocki were recorded to fly. Morphometric analysis of the main structures associated with the locomotor abilities showed that hybrids exhibited intermediate size for most of characters. The width of pronotum of both hybrids and T. juazeirensis was significantly larger than T. sherlocki. We suggested that lack of flight ability of T. sherlocki is possibly a result of reduced wing size and distinct shape, combined with undeveloped flight muscles in a shorter thoracic box. The mobility of T. sherlocki might be compensated by its significantly longer legs, and may possibly increase its ability to invade human dwellings by active dispersion. What is more, this study showed that hybrids between T. sherloki and T. juazeirensis have intermediate morphological characters that may give them higher fitness than their parents, and thus may advance the process of house infestation by either fight or walking in case of an eventual natural hybridization.

Phylogenetic and phenotypic relationships among Triatoma carcavalloi (Hemiptera: Reduviidae: Triatominae) and related species collected in domiciles in Rio Grande do Sul State, Brazil.

Triatoma carcavalloi is considered a rare Chagas disease vector often collected inside domiciles in Rio Grande do Sul State. In this Brazilian state, T. carcavalloi has been collected in the same ecotope (rock piles) with two other species (T. rubrovaria and T. circummaculata), with which it also shares morphological characteristics. Previous morphological studies placed T. carcavalloi in the same species complex ("infestans complex") and subcomplex ("rubrovaria subcomplex") as T. rubrovaria, whereas T. circummaculata was placed in the "circummaculata complex." The phylogeny of a group composed of 16 species of triatomines was reevaluated with the inclusion of T. carcavalloi by Bayesian analysis using mtDNA sequences of subunits 12S and 16S of the ribosomal RNA, and the cytochrome oxidase I (COI) genes. The phenotypic relationship among T. carcavalloi and related triatomines was also inferred from morphometrics. Phylogenetic results indicate that T. carcavalloi is a sister species of T. rubrovaria, and both were recovered as closely related to T. circummaculata. Morphometric studies confirmed the closeness among T. carcavalloi, T. rubrovaria, and T. circummaculata, prompting the placement of the latter species in the "infestans complex" and "rubrovaria subcomplex."

Phylogenetic relationships within the oliveirai complex (Hemiptera:Reduviidae:Triatominae).

The oliveirai complex is actually composed of nine related species of Triatominae that occur in the Pantanal ecosystem of Mato Grosso, Brazil. In order to confirm their specific status and infer their phylogenetic relationships, we performed a genetic and morphometric comparison concerning seven of the nine species of the complex. Isoenzyme analysis provided broadly similar phylogenetic information to that derived from discriminant analysis of size-free variables. The derived trees reveal two main species groups, one composed of three closed species (T. jurbergi, T. matogrossensis and T. vandae), and another one subdivided into two different pairs, the T. guazu-T. williami pair and the T. klugi-T. oliveirai pair. The results call the specific status of T. guazu and T. williami into question, support the possibility that the other analyzed species represent distinct taxa and query the existence of the oliveirai complex.