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Arboviruses including dengue, Zika, and chikungunya are amongst the most significant public health concerns worldwide. Arbovirus control relies on the use of insecticides to control the vector mosquito Aedes aegypti (Linnaeus), the success of which is threatened by widespread insecticide resistance. The work presented here profiled the gene expression of Ae. aegypti larvae from field populations of Ae. aegypti with differential susceptibility to temephos originating from two Colombian urban locations, Bello and Cúcuta, previously reported to have distinctive disease incidence, socioeconomics, and climate. We demonstrated that an exclusive field-to-lab (Ae. aegypti strain New Orleans) comparison generates an over estimation of differential gene expression (DGE) and that the inclusion of a geographically relevant field control yields a more discrete, and likely, more specific set of genes. The composition of the obtained DGE profiles is varied, with commonly reported resistance associated genes including detoxifying enzymes having only a small representation. We identify cuticle biosynthesis, ion exchange homeostasis, an extensive number of long noncoding RNAs, and chromatin modelling among the differentially expressed genes in field resistant Ae. aegypti larvae. It was also shown that temephos resistant larvae undertake further gene expression responses when temporarily exposed to temephos. The results from the sampling triangulation approach here contribute a discrete DGE profiling with reduced noise that permitted the observation of a greater gene diversity, increasing the number of potential targets for the control of insecticide resistant mosquitoes and widening our knowledge base on the complex phenotypic network of the Ae. aegypti response to insecticides.

Insecticide resistance is a significant challenge facing the successful control of mosquito vectors globally. Bioassays are currently the only method for phenotyping resistance. They require large numbers of mosquitoes for testing, the availability of a susceptible comparator strain and often insectary facilities. This study aimed to trial the novel use of rapid evaporative ionisation mass spectrometry (REIMS) for the identification of insecticide resistance in mosquitoes. No sample preparation is required for REIMS and analysis can be rapidly conducted within hours. Temephos resistant Aedes aegypti (Linnaeus) larvae from Cúcuta, Colombia and temephos susceptible larvae from two origins (Bello, Colombia, and the lab reference strain New Orleans) were analysed using REIMS. We tested the ability of REIMS to differentiate three relevant variants: population source, lab versus field origin and response to insecticide. The classification of these data was undertaken using linear discriminant analysis (LDA) and random forest. Classification models built using REIMS data were able to differentiate between Ae. aegypti larvae from different populations with 82% (± 0.01) accuracy, between mosquitoes of field and lab origin with 89% (± 0.01) accuracy and between susceptible and resistant larvae with 85% (± 0.01) accuracy. LDA classifiers had higher efficiency than random forest with this data set. The high accuracy observed here identifies REIMS as a potential new tool for rapid identification of resistance in mosquitoes. We argue that REIMS and similar modern phenotyping alternatives should complement existing insecticide resistance management tools.

Arboviruses including dengue, Zika and chikungunya are amongst the most significant public health concerns worldwide and their control relies heavily on the use of insecticides to control the vector mosquito Aedes aegypti. The success of controlling these vector-pathogen systems is threatened by widespread insecticide resistance. The work presented here profiled the gene expression of the larvae from two field populations of Ae. aegypti with differential susceptibility to temephos. The contrasting phenotypes originated from two Colombian urban locations, Bello and Cúcuta, that we have previously reported to have distinctive disease incidence, socioeconomics, and climate. The closeness of the geographical origin of the study populations was suspected to be highly influential in the profiling of the gene expression of resistance since the mosquito’s resistance levels themselves are highly dependent upon environmental variables. We demonstrated that an exclusive field-to-lab (Ae. aegypti reference strain New Orleans) comparison generates an over estimation of differential gene expression (DGE) and that the inclusion of a geographically relevant field control, as used here, yields a more discrete, and likely, more specific set of genes. The composition of the obtained DGE profiles is varied, with commonly reported resistance associated genes such as detoxifying enzymes having only a small representation. We identify cuticle biosynthesis, ion exchange homeostasis, an extensive number of long non-coding RNAs, and chromatin modelling among the specifically and differentially expressed genes in field resistant Ae. aegypti larvae. It was also shown that temephos resistant larvae undertake further gene expression responses when temporarily exposed to this insecticide. The results from the sampling triangulation approach undertaken here contributes a discrete DGE profiling with reduced noise that permitted the observation of a greater gene diversity. This deeper gene granularity significantly increases the number of potential targets for the control of insecticide resistant mosquitoes and widens our knowledge base on the complex phenotypic network of the Ae. aegypti mosquito responses to insecticides.Author SummaryAedes aegypti mosquitoes are vectors for several significant human viruses including dengue, Zika and chikungunya. The lack of widely available vaccines and specific antiviral treatments for these viruses means that the principal method for reducing disease burden is through controlling the vector mosquitoes. Mosquito control relies heavily on the use of insecticides and successful vector control is threatened by widespread insecticide resistance in Ae. aegypti. Here, we examined changes in gene expression that occur in temephos resistant populations of Ae. aegypti from two field populations in Colombia. We compare gene expression in resistant larvae from Cúcuta with susceptible larvae from Bello and a susceptible laboratory strain of Ae. aegypti (New Orleans). We also compare mosquitoes from Cúcuta with and without temephos exposure. We report several differentially expressed genes beyond those usually reported in resistant mosquitoes. We also demonstrate the over estimation in differential gene expression that can occur when field resistant populations are compared against lab susceptible populations only. The identification of new mechanisms involved in the development of insecticide resistance is crucial to fully understanding how resistance occurs and how best it can be reduced.

Dengue, Zika and chikungunya are diseases of global health significance caused by arboviruses and transmitted by the mosquito Aedes aegypti, which is of worldwide circulation. The arrival of the Zika and chikungunya viruses to South America increased the complexity of transmission and morbidity caused by these viruses co-circulating in the same vector mosquito species. Here we present an integrated analysis of the reported arbovirus cases between 2007 and 2017 and local climate and socio-economic profiles of three distinct Colombian municipalities (Bello, Cúcuta and Moniquirá). These locations were confirmed as three different ecosystems given their contrasted geographic, climatic and socio-economic profiles. Correlational analyses were conducted with both generalised linear models and generalised additive models for the geographical data. Average temperature, minimum temperature and wind speed were strongly correlated with disease incidence. The transmission of Zika during the 2016 epidemic appeared to decrease circulation of dengue in Cúcuta, an area of sustained high incidence of dengue. Socio-economic factors such as barriers to health and childhood services, inadequate sanitation and poor water supply suggested an unfavourable impact on the transmission of dengue, Zika and chikungunya in all three ecosystems. Socio-demographic influencers were also discussed including the influx of people to Cúcuta, fleeing political and economic instability from neighbouring Venezuela. Aedes aegypti is expanding its range and increasing the global threat of these diseases. It is therefore vital that we learn from the epidemiology of these arboviruses and translate it into an actionable local knowledge base. This is even more acute given the recent historical high of dengue cases in the Americas in 2019, preceding the COVID-19 pandemic, which is itself hampering mosquito control efforts., Author summary Viruses transmitted by Ae. aegypti mosquitoes (dengue, Zika, chikungunya) are amongst the most significant public health concerns of recent years due to the increase in global cases and the rapid spread of the vectors. The primary method of controlling the spread of these arboviruses is through mosquito control. Understanding factors associated with risk of these viruses is key for informing control programmes and predicting when outbreaks may occur. Climate is an important driver in mosquito development and virus reproduction and hence the association of climate with disease risk. Socio-economic factors contribute to perpetuate disease risk. Areas of high poverty have abundance of suitable habitat for Ae. aegypti (e.g. due to poor housing and sanitation). This study investigated the factors affecting arbovirus incidence in three distinct regions of Colombia: Bello, Cúcuta and Moniquirá. The results show significant relationships between disease incidence and temperature, precipitation and wind speed. A decline in dengue following outbreaks of Zika (2016) is also reported. Measures of poverty, including critical overcrowding and no access to improved water source were also found to be higher in areas of higher disease incidence. The results of this study highlight the importance of using a multifactorial approach when designing vector control programs in order to effectively distribute local health care resources.

Bacteria can survive high doses of antibiotics through stochastic phenotypic diversification. We present initial evidence that folate metabolism could be involved with the formation of persisters. The aberrant expression of the folate enzyme gene fau seems to reduce the incidence of persisters to antibiotics. Folate-impaired bacteria had a lower generation rate for persisters to the antibiotics ampicillin and ofloxacin. Persister bacteria were detectable from the outset of the exponential growth phase in the complex media. Gene expression analyses tentatively showed distinctive profiles in exponential growth at times when bacteria persisters were observed. Levels of persisters were assessed in bacteria with altered, genetically and pharmacologically, folate metabolism. This work shows that by disrupting folate biosynthesis and usage, bacterial tolerance to antibiotics seems to be diminished. Based on these findings there is a possibility that bacteriostatic antibiotics such as anti-folates could have a role to play in clinical settings where the incidence of antibiotic persisters seems to drive recalcitrant infections.

Bacteria can survive high doses of antibiotics through stochastic phenotypic diversification.We present initial evidence that folate metabolism could be involved with the formation of persisters. The aberrant expression of the folate enzyme genefauseems to reduce the incidence of persisters to antibiotics. Folate impaired bacteria had a lower generation rate for persisters to both antibiotics ampicillin and ofloxacin. Persister bacteria were detectable from the outset of the exponential growth phase in the complex media. Gene expression analyses showed tentatively distinctive profiles in exponential growth at times when bacteria persisters were observed. Levels of persisters were assessed in bacteria with altered, genetically and pharmacologically, folate metabolism. This work shows that by disrupting folate biosynthesis and usage, bacterial tolerance to antibiotics seems to be diminished. Based on these findings there is a possibility that bacteriostatic antibiotics such as antifolates could have a role to play in clinical settings where the incidence of antibiotic persisters seem to drive recalcitrant infections.

Post-mitotic, differentiated cells exhibit a variety of characteristics that contrast with those of actively growing neoplastic cells, such as the expression of cell-cycle inhibitors and differentiation factors. We hypothesized that the gene expression profiles of these differentiated cells could reveal the identities of genes that may function as tumour suppressors. Here we show, using in vitro and in vivo studies in mice and humans, that the mitochondrial protein LACTB potently inhibits the proliferation of breast cancer cells. Its mechanism of action involves alteration of mitochondrial lipid metabolism and differentiation of breast cancer cells. This is achieved, at least in part, through reduction of the levels of mitochondrial phosphatidylserine decarboxylase, which is involved in the synthesis of mitochondrial phosphatidylethanolamine. These observations uncover a novel mitochondrial tumour suppressor and demonstrate a connection between mitochondrial lipid metabolism and the differentiation program of breast cancer cells, thereby revealing a previously undescribed mechanism of tumour suppression.