New occurrences of mosquitoes (Diptera: Culicidae) in the Atlantic Forest biome of the Brazilian Northeast.

BACKGROUND: Information regarding the distribution of Culicidae species in the northeastern region of Brazil is scarce. METHODS: Immatures were collected from approximately four fragments of the Atlantic Forest. RESULTS: This study presents new occurrences of 18 Culicidae species in Pernambuco state: Anopheles kompi, Georgecraigius fluviatilis, Culex bidens, Culex chidesteri, Culex bastagarius, Culex imitator, Mansonia humeralis, Wyeomyia incaudata, Uranotaenia apicalis, Culex mollis, Culex usquatus, Culex dunni, Culex serratimarge, Culex ybarmis, Culex microphyllus, Sabethes purpureus, Wyeomyia pilicauda, and Wyeomyia airosai. The last nine species were also new records for the northeast region. CONCLUSIONS: With the inclusion of these newly recorded species, the total number of mosquitoes documented in Pernambuco state now rises to 94.

New occurrences of mosquitoes (Diptera: Culicidae) in the Atlantic Forest biome of the Brazilian Northeast

ABSTRACT Background: Information regarding the distribution of Culicidae species in the northeastern region of Brazil is scarce. Methods: Immatures were collected from approximately four fragments of the Atlantic Forest. Results: This study presents new occurrences of 18 Culicidae species in Pernambuco state: Anopheles kompi, Georgecraigius fluviatilis, Culex bidens, Culex chidesteri, Culex bastagarius, Culex imitator, Mansonia humeralis, Wyeomyia incaudata, Uranotaenia apicalis, Culex mollis, Culex usquatus, Culex dunni, Culex serratimarge, Culex ybarmis, Culex microphyllus, Sabethes purpureus, Wyeomyia pilicauda, and Wyeomyia airosai. The last nine species were also new records for the northeast region. Conclusions: With the inclusion of these newly recorded species, the total number of mosquitoes documented in Pernambuco state now rises to 94.

The aquatic macrophyte Salvinia molesta mitigates herbicides (glyphosate and aminomethylphosphonic acid) effects to aquatic invertebrates.

We evaluated the individual and combined effects of different environmentally representative concentrations of glyphosate (0, 25, 50, 75, and 100 µg l-1) and aminomethylphosphonic acid (AMPA; 0, 12.5, 25, 37.5, and 50 µg l-1) on the physiology of Aedes aegypti larvae, as well as the capacity of the aquatic macrophyte Salvinia molesta to attenuate those compounds' toxicological effects. Larvae of Ae. aegypti (between the third and fourth larval stages) were exposed for 48 h to glyphosate and/or AMPA in the presence or absence of S. molesta. Glyphosate and AMPA induced sublethal responses in Ae. aegypti larvae during acute exposures. Plants removed up to 49% of the glyphosate and 25% of AMPA from the water, resulting in the exposure of larvae to lower concentration of those compounds in relation to media without plants. As a result, lesser effects of glyphosate and/or AMPA were observed on larval acetylcholinesterase, P450 reductase, superoxide dismutase, mitochondrial electron transport chain enzymes, respiration rates, and lipid peroxidation. In addition to evidence of deleterious effects by media contamination with glyphosate and AMPA on aquatic invertebrates, our results attest to the ability of S. molesta plants to mitigate the toxicological impacts of those contaminants.

Surveillance of Aedes aegypti and Aedes albopictus (Diptera: Culicidae) as a Method for Prevention of Arbovirus Transmission in Urban and Seaport Areas of the Southern Coast of Brazil.

Entomological surveillance is a traditional method to measure presence, distribution, and seasonal variation of vectors in urban areas, and is essential to targeted control activities to prevent arbovirus transmission. Ovitraps as one of the main components of surveillance programs, enable determination of female oviposition behavior, as well as identification of seasonal variations of the vector. The goals of this study were 1) to detect the mosquitos (Aedes aegypti Linnaeus) and (Aedes albopictus Skuse) (Diptera:Culicidae), in Paranaguá city, 2) to assess ovitrap positive index (OPI), egg density index (EDI), and their relationship with meteorological variables, and 3) to evaluate the vertical transmission of dengue, Zika, and chikungunya in Ae. aegypti and Ae. albopictus. The study was carried out in urban areas of Paranaguá city, an important port region of Brazil, from June 2017 to November 2018. The city was divided into 16 area-clusters. Three-hundred and thirty-one ovitraps were installed monthly, remaining for four days in selected places. Kernel density maps were done to compare the spatiotemporal distribution of collected eggs. Areas which maintained constant oviposition associated with vector activity were identified and were found to overlap the area-clusters with the highest EDI. As viral RNA was not detected, vertical transmission was likely not a maintenance mechanism of arbovirus circulation in Ae. aegypti and Ae. albopictus vectors. This study reiterates the importance, efficiency, and feasibility of ovitraps to monitor the presence and dynamics of Aedes spp. populations.

Genetic diversity and Kdr mutations of natural Aedes (Stegomyia) aegypti (Diptera: Culicidae) populations of Brazil

ABSTRACT Aedes (Stegomyia) aegypti is an important vector of dengue, yellow fever, chikungunya and Zika virus. It is well known that resistance monitoring and genetic diversity data help designing the vector control programs. This study aimed to evaluate resistance to pyrethroids (PYs) through the frequency of kdr mutations Val1016IIe and F1534C, and the genetic variation of the mitochondrial gene ND4 in six natural populations of A. aegypti from Paraná - Brazil. Adults were obtained from eggs collected from Alvorada do Sul, Marilena, Maringá, Nova Londrina, Paranavaí and São Carlos do Ivaí. From these adults, 345 were used to identify the 1016 and 1534 sites, and 120 were used to perform the ND4 gene analysis. The studied populations from Paraná showed PYs resistance, low gene flow and genetic diversity. Additionally, a relationship was observed among the haplotypes of populations from the Amazon and Southeastern Brazil, Peru, Mexico, and North America.

Do anti-HIV drugs pose a threat to photosynthetic microorganisms?

We investigated the occurrence and risk assessment of three anti-HIV drugs [(tenofovir (TNF), lamivudine (LMV) and efavirenz (EFV)] in urban rivers from Curitiba (Brazil), as well as the individual and combined effects of their environmental representative concentrations on the freshwater periphytic species Synechococcus elongatus (Cyanobacteria) and Chlorococcum infusionum (Chlorophyta). The three studied drugs, except TNF, were found in 100% of the samples, and concentrations in samples ranged from 165 to 412 ng TNF L-1, 173-874 ng LMV L-1 and 13-1250 ng EFV L-1. Bioassays using artificial contaminated water showed that at environmental concentrations, TNF and LMV did not represent environmental risks to the studied photosynthetic organisms. However, EFV was shown to be toxic, affecting photosynthesis, respiration, and oxidative metabolism. The studied drugs demonstrated interactive effects. Indeed, when submitted to the combination of TNF and LMV, decreased photosynthesis was observed in C. infusionum cells. Moreover, the toxic effects of EFV were amplified in both species when TNF and/or LMV were added to the media. The simultaneous presence of TNF, LMV and EFV in environmental matrices associated with their interactive effects, lead to increased toxicological effects of water contaminated by anti-HIV drugs and thus to an ecological threat to photosynthetic microorganisms.

Accumulation of nylon microplastics and polybrominated diphenyl ethers and effects on gut microbial community of Chironomus sancticaroli.

Microplastics (MP) are emerging contaminants with the capacity to bind and transport hydrophobic organic compounds of environmental concern, such as polybrominated diphenyl ethers (PBDEs). The aim of this study was to investigate the ingestion of nylon (polyamide) MP alone and when associated with PBDEs and their effects on Chironomus sancticaroli larvae survival and microbiome structure. Survival, PBDE uptake and microbial community composition were measured in fourth instar larvae exposed for 96 h to BDEs- 47, 99, 100 and 153 in the presence and absence of 1% w/w MP in sediment. Microbiome community structures were determined through high throughput sequencing of 16S small subunit ribosomal RNA gene (16S rRNA). Initial experiments showed that larvae ingested MP faster at 0.5% w/w MP, while depuration was more efficient at 1% w/w MP, although retention of MP was seen even after 168 h depuration. No mortality was observed as a result of PBDEs and MP exposure. MP had a negative effect on PBDE concentration within larvae (η2 = 0.94) and a negative effect on sediment concentrations (η2 = 0.48). In all samples, microbial communities were dominated by Alphaproteobacteria, Betaproteobacteria, Actinobacteria and Gammaproteobacteria. Bacterial alpha diversity was not significantly affected by PBDEs or MP exposure. However, the abundance of discrete bacterial taxa was more sensitive to MP (X2 = 45.81, p = 0.02), and PBDE exposure. Our results highlight that C. sancticaroli showed no acute response to MPs and PBDEs, but that MPs influenced bacterial microbiome structure even after only short-term acute exposure.

Gene Flow Patterns of the Aedes aegypti (Diptera: Culicidae) Mosquito in Colombia: a Continental Comparison Suggests Multiple Invasion Routes and Gene Exchange

ABSTRACT In Colombia Aedes aegypti is present in 80% of the country up to 2,300 m; however, little is known of its genetic relations within a country context and, hence, within a broader context, for example, America. The aforementioned, herein, analyzed the gene flow within a context of the Americas, its directionality and genetic diversity of the mitochondrial lineages in the A. aegypti populations for Colombia. This called for the use of the sequences for A. aegypti available of the mitochondrial ND4 gene in the GenBank for Colombia and the American continent. No presence was found of nuclear mitochondrial pseudogenes (NUMTs) for Colombia. It is estimated that in Colombia the gene flow of the A. aegypti populations is occurring from the southeast and northeast toward the center of the country. In comparison with the mitochondrial sequences for America, the vector's haplotypes in Colombia suggest connections between the populations of mosquitoes from the south with those from the north of the continent. The gene flow model at continental scale suggests bidirectional connections between the populations from the north of the continent with those from the south, while at South American scale it proposes the gene flow in all the directions with respect to the Colombian. The Colombian A. aegypti vector monitoring and control strategies must pay special attention to the vector's points of entry into Colombia related with Peru, Venezuela, Brazil, Mexico, and North America to avoid the entry of populations with characteristics like resistance to insecticides or vector competition.

Isolated and combined effects of glyphosate and its by-product aminomethylphosphonic acid on the physiology and water remediation capacity of Salvinia molesta.

We evaluate the isolated and combined effects of glyphosate and its by-product aminomethylphosphonic acid (AMPA) and the potential of the aquatic macrophyte Salvinia molesta to remove these chemicals from contaminated water. Plants were exposed to environmentally relevant concentrations of glyphosate (0, 20, 40, 60, 80 and 100 µg l-1) or AMPA (0, 10, 20, 30, 40 and 50 µg l-1) for seven days. Then, based on the effective concentrations of glyphosate found to reduce photosynthetic rates by 10% (EC10) and 50% (EC50), the plants were exposed to combinations of 0, 16 and 63.5 µg glyphosate l-1 and 0, 5, 15, 25 µg AMPA l-1. The EC(10) and EC(50) were lower for AMPA (6.1 µg l-1 and 28.4 µg l-1 respectively) than for glyphosate (16 and 63.5 µg glyphosate l-1 respectively). When occurring together, the deleterious effects of those chemicals to plants increased. S. molesta plants removed up to 74.15% of glyphosate and 71.34% of AMPA from culture water. Due to its high removal efficiency, S. molesta can be used in phytoremediation programs. It will be important to evaluate the combined effects of glyphosate and AMPA in any toxicological studies of the herbicide.

New traps for the capture of Aedes aegypti (Linnaeus) and Aedes albopictus (Skuse) (Diptera: Culicidae) eggs and adults.

The control of arboviruses carried by Aedes aegypti (Linnaeus) and Aedes albopictus (Skuse) can be performed with tools that monitor and reduce the circulation of these vectors. Therefore, the efficiency of four types of traps in capturing A. aegypti and A. albopictus eggs and adults, with the biological product Vectobac WG, was evaluated in the field. For this, 20 traps were installed in two locations, which were in the South (Londrina, Paraná) and North (Manaus, Amazonas) Regions of Brazil, from March to April 2017 and January to February 2018, respectively. The UELtrap-E (standard trap) and UELtrap-EA traps captured A. aegypti and A. albopictus eggs: 1703/1866 eggs in Londrina, and 10268/2149 eggs in Manaus, respectively, and presented high ovitraps positivity index (OPI) values (averages: 100%/100% in Londrina, and 100%/96% in Manaus, respectively); and high egg density index (EDI) values (averages: 68/75 in Londrina, and 411/89 in Manaus, respectively), so they had statistically superior efficiency to that of the CRtrap-E and CRtrap-EA traps in both regions, that captured less eggs and adults: 96/69 eggs in Londrina, and 1091/510 eggs in Manaus, respectively. Also presented lower OPI values (averages: 28%/4% in Londrina, and 88%/60% in Manaus, respectively); and lower EDI values (averages: 10.5/9 in Londrina, and 47/30 in Manaus, respectively). The capture ratios of Aedes adults in the UELtrap-EA and CRtrap-EA traps in Londrina and Manaus were 53.3%/29.5% and 0%/9.8%, respectively. UELtrap-EA can be adopted as efficient tool for Aedes monitoring due to their high sensitivity, low cost and ease of use.

Molecular and biochemical evaluation of effects of malathion, phenanthrene and cadmium on Chironomus sancticaroli (Diptera: Chironomidae) larvae.

In-vitro effects of sub-lethal concentrations of malathion, phenanthrene (Phe) and cadmium (Cd) were tested on Chironomus sancticaroli larvae in acute bioassays by measuring biochemical and molecular parameters. Malathion was evaluated at 0.001, 0.0564 and 0.1006 mg L-1; Phe at 0.0025, 1.25 and 2.44 mg L-1; and Cd at 0.001, 3.2 and 7.4 mg L-1. The recovery test carried out at the highest concentration of each compound showed that survival of larvae exposed to Phe ranged from 4% to 5%, while the effects of malathion and Cd were irreversible, not allowing the emergence of adults. Results showed that malathion and Cd inhibited AChE, EST-α and ES-ß activities at the two highest concentrations. Phe at 0.0025, 1.25 and 2.44 mg L-1; and Cd at 3.2 and 7.4 mg L-1 inhibited glutathione S-transferase activity. Oxidative stress was exclusively induced by the lowest concentration of malathion considering SOD activity once CAT was unaffected by the stressors. Lipid peroxidation was registered exclusively by malathion at the two highest concentrations, and total hemoglobin content was only reduced by Cd at the two highest concentrations. The relationship among biochemical results, examined using the PCA, evidenced that malathion and Cd concentrations were clustered into two groups, while Phe only formed one group. Four hemoglobin genes of C. sancticaroli were tested for the first time in this species, with Hemoglobin-C being upregulated by malathion. The toxicity ranking was malathion > Phe > Cd, while biochemical and molecular results showed the order malathion > Cd > Phe. Our results highlight the importance of combining different markers to understand the effects of the diverse compounds in aquatic organisms.

Individual and combined effects of amoxicillin, enrofloxacin, and oxytetracycline on Lemna minor physiology.

We investigated individual and combined effects of environmentally representative concentrations of amoxicillin (AMX; 2 µg l-1), enrofloxacin (ENR; 2 µg l-1), and oxytetracycline (OXY; 1 µg l-1) on the aquatic macrophyte Lemna minor. While the concentrations of AMX and ENR tested were not toxic, OXY decreased plant growth and cell division. OXY induced hydrogen peroxide (H2O2) accumulation and related oxidative stress through its interference with the activities of mitochondria electron transport chain enzymes, although those deleterious effects could be ameliorated by the presence of AMX and/or ENR, which prevented the overaccumulation of ROS by increasing catalase enzyme activity. L. minor plants accumulated significant quantities of AMX, ENR and OXY from the media, although competitive uptakes were observed when plants were submitted to binary or tertiary mixtures of those antibiotics. Our results therefore indicate L. minor as a candidate for phytoremediation of service waters contaminated by AMX, ENR, and/or OXY.

Cell damage induced by polybrominated diphenyl ethers on Chironomus sancticaroli (Diptera: Chironomidae)

Abstract Histological effects of polybrominated diphenyl ethers (PBDEs) were observed in Chironomus sancticaroli larvae which underwent acute exposure. 2,2′,4-triBDE (BDE-17), 2,2′,4,4′-tetraBDE (BDE-47) and 2,2′,4,4′,5-pentaBDE (BDE-99) were evaluated at 0.5, 2.0 and 20 μg L-1. Cytoplasm vacuolisation of oenocytes was observed in the larvae exposed to BDE-17 and BDE-47. Cuénot cells were disrupted at the brush border as an effect of the three evaluated congeners highlighting BDE-47 at 2.0 μg L-1; 60% of larvae displayed this disruption. The midgut showed changes in the morphology of apex cells located next to the lumen of region I exposed to BDE-17 and BDE-47, while BDE-99 induced a narrowing of the lumen diameter. Significant cytoplasm vacuolisation of the larvae exposed to BDE-47 and BDE-99 was observed in region II of the midgut. Salivary glands showed acidophilic granules in the cytoplasm exposed to BDE-17 and BDE-47. The results showed that the tissues of C. sancticaroli were sensitive to flame retardants; these histopathologies can compromise the health and physiology of this organism, highlighting the concern with the presence of PBDEs in freshwater sediments.

Data of insecticide effects of natural compounds against third instar larvae of Cochliomyia macellaria.

Morphological biomarkers can be used to establish a diagnosis of fly larvae structural damage and toxicity to target cells by biopesticide candidates. Insecticide activity of natural compounds such as Curcuma longa essential oil (CLLEO) extracted from leaves, and its major constituent α-phellandrene have proven to be a novel biopesticide candidate against third instar larvae (L3) of Cochliomyia macellaria. In this way, groups of 20 L3 were placed on filter paper impregnated with different concentrations of CLLEO, from 0.31 to 2.86 µL/cm2 and α-phellandrene, from 0.29 to 1.47 µL/cm2. The extracts were solubilized in ethanol. Data shown in this article is related to the research article "Can an overlooked by-product from turmeric industry be effective for myiasis control?" Chaaban et al., 2019. Data on L3 toxicity was observed after 6 and 24h of contact with both extracts, as well as a marked reduction of L3 movement, color changes in the cuticle and progressive darkening in their body. Major cuticle damage and L3 mortality were reported.

Novel molecular evidence of population structure in Anopheles (Kerteszia) bellator from Brazilian Atlantic Forest.

Anopheles bellator is a primary malaria vector in the Atlantic Forest. Partial sequences of timeless and Clock genes were used to assess the genetic differentiation of five Brazilian populations, which showed strong population structure (e.g. high F ST values and fixed differences) in all pairwise comparisons between Bahia sample and the others from Paraná, São Paulo and Rio de Janeiro states. Also, the resulting phylogenetic trees clearly grouped the sequences from Bahia in a different cluster with high bootstrap values. Among southern and southeastern populations low levels of genetic differentiation were found suggesting a general stability of the genetic structure.

Molecular identification of blood meals in mosquitoes (Diptera, Culicidae) in urban and forested habitats in southern Brazil.

The study of host associations of mosquitoes (Diptera, Culicidae) provides valuable information to assist in our understanding of a variety of related issues, from their life-history to the entomological surveillance of pathogens. In this study, we identified and characterized mosquito blood meals from both urban and forested areas in the city of Paranaguá, state of Paraná, Brazil, by analyzing the amplification of host DNA ingested by mosquitoes under different storage conditions and digestion levels. Host DNA preservation was evaluated in fresh blood meals according to storage duration (30 to 180 days) and temperature (-20°C / -80°C) and, in digested blood, according the degree of digestion classified on the Sella scale. Molecular analysis of blood meals was based on DNA extraction and amplification of a fragment of the mitochondrial COI gene. We determined that, up to180 days of storage, the evaluated temperatures did not influence the preservation of fresh blood meals DNA, whereas the amplification success was increasingly reduced over the course of the digestion process. The species Anopheles cruzii, Aedes fluviatilis, Aedes scapularis, Psorophora ferox, Culex quinquefasciatus, Culex mollis, and Culex intrincatus, together with specimens representing four subgenera and one genus of Culicidae [Ae. (Ochlerotatus), Cx. (Culex), Cx. (Melanoconion), Cx. (Microculex), and Limatus, respectively] had their blood meals identified. Their diverse host use was evidenced by the identification of 19 species of vertebrate host, namely two amphibians, three mammals and 14 birds. Birds were the most commonly identified host in blood meals. These results not only show the diversity of mosquito hosts, but also underscore the challenges involved in monitoring arboviruses of public health importance, given potential combinations of host use for each mosquito species.

Effects of Ciprofloxacin and Roundup on seed germination and root development of maize.

Their continuous release into the environment, associated with their inherent biological activity, has motivated investigations into the detrimental effects of antibiotics and herbicides in natural and agricultural ecosystems. In this study, the interactive effects of the antibiotic ciprofloxacin (Cipro) and the herbicide Roundup on seed germination and root development were investigated. Although both compounds act as inhibitors of the mitochondrial electron transport chain in seeds, neither Cipro nor Roundup disrupted germinability of maize seeds. However, Cipro accelerated germination by promoting ROS accumulation in seeds, while the stimulatory effect of Roundup on ROS-scavenging enzymes (catalase and ascorbate peroxidase) seems to prevent ROS-signaling, delaying the germination process. Roundup reduced root elongation, possibly due to its interference with auxin production, thereby preventing cell division, while Cipro stimulated root elongation by increasing root oxidative status. Cipro and Roundup showed antagonistic effects on maize seeds and root physiology. The presence of the antibiotic is likely not to disturb plant development; however, its stimulatory effects were not sufficient to overcome the deleterious effects of Roundup. According to our results, glyphosate-based herbicides must be carefully used during maize cropping and although antibiotics such as Cipro may not negatively impact agricultural production, their accumulation by crops must be investigated since this can be a pathway of antibiotic-insertion into the food chain.

Cuticular damage of Lucilia cuprina larvae exposed to Curcuma longa leaves essential oil and its major compound α-phellandrene.

Morphological biomarkers as the histopathological assessment and scanning electron microscopy can be used to establish a diagnosis of structure damage and intoxication of target cells by new biopesticide candidate. In this sense, cuticle damage caused by active substances in larvae exposed to biopesticides can help to elucidate the mode action. Thus, insecticide activity analysis of essential oil of Curcuma longa leaves and its major compound α-phellandrene have proven to be a new biopesticide candidate against third instar larvae (L3) of the Australian blowfly Lucilia cuprina. In this way, groups of 20 L3 were placed on filter paper, impregnated with ranging concentrations (from 0.15 to 2.86 µL/cm2) of C. longa leaves EO and (0.29-1.47 µL/cm2) to α-phellandrene. The extracts were solubilized in ethanol. Progressive darkening in the body of L3, marked reduction of movement, color changes in larval cuticle and dead were observed 6 and 24 h after contact with both extracts.