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Open Access

Larval superiority of Culex pipiens to Aedes albopictus in a replacement series experiment: prospects for coexistence in Germany

  • Ruth Müller1Email author,
  • Timm Knautz2,
  • Simone Vollroth2,
  • Robert Berger3,
  • Aljoscha Kreß1, 2, 3,
  • Friederike Reuss1, 3,
  • David A. Groneberg1 and
  • Ulrich Kuch1
Parasites & Vectors201811:80

https://doi.org/10.1186/s13071-018-2665-3

Received: 21 August 2017

Accepted: 22 January 2018

Published: 2 February 2018

Abstract

Background

The Asian tiger mosquito Aedes albopictus is an extremely invasive, globally distributed and medically important vector of various human and veterinary pathogens. In Germany, where this species was recently introduced, its establishment may become modulated by interspecific competition from autochthonous mosquito species, especially Culex pipiens (s.l.). While competitive superiority of Ae. albopictus to Cx. pipiens (s.l.) has been described elsewhere, it has not been assessed in the epidemiological conditions of Germany. The present study aimed to determine if such superiority exists under the physicochemical and microclimatic conditions typical for container habitats in Germany.

Methods

In a replacement series experiment, the larval and pupal responses of Ae. albopictus and Cx. pipiens (s.l.) (mortality, development time, growth) to interspecific interaction (five larval ratios) at (sub-)optimal temperatures (15, 20 and 25 °C) and differing food supply (3 and 6 mg animal-based food larva-1) were investigated using a randomized split-plot design. In addition to physicochemical measurements of the test media, natural physicochemical conditions were determined for comparative analyses in mosquito breeding sites across the Rhine-Main metropolitan region of Germany.

Results

Under the physicochemical and microclimatic conditions similar to the breeding sites of the Rhine-Main region, competitive superiority of Cx. pipiens (s.l.) to Ae. albopictus in terms of larval survival was more frequently observed than balanced coexistence. Food regime and multifactorial interactions, but not temperature alone, were controlling factors for interspecific competition. Larval food regime and the larval ratio of Ae. albopictus influenced the physicochemistry and algal growth at 15 °C, with increased Ae. albopictus mortality linked to a decreasing number of Scenedesmus, Oocystis and Anabaena algae.

Conclusions

Under the present environmental conditions, the spread of Ae. albopictus from isolated foci in Germany may generally be slowed by biotic interactions with the ubiquitous Cx. pipiens (s.l.) (and potentially other container-breeding mosquito species) and by limnic microalgae in microhabitats with high resource levels. Detailed knowledge of the context dependency in temperate mosquito ecology, and interrelations of physicochemistry and phycology may help to achieve a better understanding of the upcoming Ae. albopictus colonization processes in central and northern Europe.

Keywords

MicroalgaeInterspecific competitionNiche differentiationResource competitionWater chemistry

Background

The extremely invasive mosquito Aedes (Stegomyia) albopictus Skuse 1894, a known vector of various human pathogens like dengue and chikungunya viruses, is gradually extending its distribution to colder climate regions [1]. At present, the northernmost breeding populations of this species in Europe occur in Freiburg [2], Heidelberg and Jena, Germany (Norbert Becker, personal communication, June 2016). A further spread of Ae. albopictus into the Rhine rift valley and parts of Bavaria and North Rhine-Westphalia, Germany, is very likely due to climatic habitat suitability and man-made features [3, 4]. Apart from the frequently suboptimal low temperatures in these regions, the establishment of Ae. albopictus may be modulated by interactions with resident container-breeding mosquitoes [57].

Aedes albopictus was shown to be a strong competitor for resident mosquito species such as Aedes triseriatus as well as exotic species like Aedes aegypti in the United States of America [7, 8]. However, environmental variation may change competitive balances [9] as recently demonstrated in Japan [10]: At Mount Konpira, Japan, the formerly predominant Ae. albopictus is currently becoming superseded by the formerly rare, but now superior competitor Aedes flavopictus, most probably due to changes in differential responses to environmental variability and altered species interactions [10]. As the competitive superiority of container-breeding mosquitoes strongly depends on context and is reversible, resident mosquitoes may have the potential to act as superior competitors in colder ecoregions due to their better adaptation to low temperatures than the invader Ae. albopictus which has a (sub-) tropical origin [11].

Negative interactions between different mosquito species frequently occur during their aquatic larval developmental stages and can be attributed to direct (chemical/physical interference) or indirect interactions (resource competition), generally known as interspecific competition [7, 9, 12]. Numerous laboratory investigations have corroborated that the pattern of the heterospecific larval response of Ae. albopictus depends not only on the competitor but also on the environmental context [9], food quantity or quality [5, 1317] and food-temperature interactions [18, 19]. Larval resource competition has been considered as a probable reason for the decline of Ae. aegypti in concurrence with the arrival of Ae. albopictus in the United States [15, 20]. Furthermore, species-specific differences in resource utilization are thought to shape microhabitat-specific mosquito assemblages [15, 16, 21]. Water chemistry, as a direct function of available food resources and resource utilization by mosquitoes, has also been considered to control niche differentiation [2224]. For instance, concentrations of nitrite and nitrate and the pH shaped the local distribution of the exotic Aedes notocriptus and the endemic Culex pervigilans in New Zealand [24]. Furthermore, certain microalgal species have been shown to exclude Ae. albopictus from microhabitats in Hawaii [22]; consequently, microalgae have been recommended as agents of biological mosquito control [25].

The common house mosquito Culex pipiens (s.l.) Linnaeus 1758 is a competent vector for West Nile and Usutu viruses and other viral and parasitic pathogens of human and veterinary importance [26]. This species, in particular, could be a strong competitor for Ae. albopictus in Germany because both species often share breeding containers [18]. In the laboratory, a superiority of Ae. albopictus over Cx. pipiens (s.l.) has been demonstrated [18, 27]. A significant niche overlap of Ae. albopictus and Cx. pipiens (s.l.) has been confirmed in Italy (11.3% of 1194 monitored containers) although habitat preferences differed slightly in terms of container volume (Ae. albopictus: < 5 l; Cx. pipiens (s.l.): > 5 l) and microclimate (Ae. albopictus: shaded sites; Cx. pipiens (s.l.): sunny sites) [18]. Both species overlapped greatly in seasonal population growth [18, 27, 28] whereas thermal preferences differ (Ae. albopictus: 29.7 °C [29], Cx. pipiens (s.l.): 20 to 25 °C [30]). The overlapping ecological niche and seasonal population activities in the presence of dissimilar autecological optima render a regionally disparate competitive superiority of Ae. albopictus over Cx. pipiens (s.l.) more than likely [5, 18, 27].

The ecology of container-breeding mosquitoes in temperate regions is still poorly understood. To achieve a better mechanistic understanding of the upcoming Ae. albopictus colonization processes in Germany, we tested whether the described competitive superiority of Ae. albopictus over Cx. pipiens (s.l.) also exists under the physicochemical and microclimatic conditions that are typical for container habitats in the Rhine rift valley of Germany. We hypothesized that (i) competitive superiority depends on the species-specific thermal tolerance spectrum, and therefore Ae. albopictus might be advantaged at higher and Cx. pipiens (s.l.) at lower temperatures, respectively. We further hypothesized that (ii) food regime, as a major determinant for interspecific interactions, interacts with temperature and that (iii) resource competition between Ae. albopictus and Cx. pipiens (s.l.) is directly connected to physicochemical and phycological parameters of their microhabitat.

Methods

Natural microhabitats of mosquito larvae

Physicochemistry

In a radius of 50 km around the city of Frankfurt am Main (Rhine-Main Metropolitan region, northern border of the Rhine rift valley, Germany), intermittent and permanent waterbodies of natural and anthropogenic origin (natural: temporary and permanent puddles, flooded patches/pools in forests and meadows, tree holes, root and rock cavities; anthropogenic: agricultural plastic covers, troughs on paddocks, water storage basins, fountain basins, planters, vases, barrels, car tires, buckets, watering cans) were monitored for the occurrence of mosquito larvae from 27th August to 27th October 2010 (Additional file 1: Figure S1).

Species identification

Species identification was based on sequence analysis of the ‘barcode region’ of the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene. Following proteinase K digestion and phenol-chloroform extraction [31], the DNA of larvae was used for PCR amplification [TrueStartTM Hot Start-Taq DNA polymerase (Fermentas, St. Leon, Germany), 5 pmol LCO1490 primer, 5 pmol HCO 2198 primer, MWG Operon (Eurofins, Ebersberg, Germany)] [32]. After Sanger sequencing using the amplification primers, the ~ 700 bp cox1 mitochondrial DNA barcodes were submitted to the following databases, and compared to entries therein for species identification: Barcode of Life Data System (boldsystems.org), NCBI-BLAST (http://blast.ncbi.nlm.nih.gov/Blast.cgi) and European Nucleotide Archive (ebi.ac.uk/ena).

At a subset of sites positive for Cx. pipiens (s.l.), conductivity (± 2%, LWT-01 sensor, Voltcraft, Hirschau, Germany), temperature (± 0.8 °C, DO-100 sensor, Voltcraft, Hirschau, Germany), oxygen concentration (± 0.4 mg l-1, DO-100 sensor, Voltcraft, Hirschau, Germany) and pH (± 0.01, PHT-02 ATC sensor, Voltcraft, Hirschau, Germany) were measured in situ. Also, 1 L plastic cups were filled with 600 ml water and placed at a sunny and a shaded site in Frankfurt am Main. The water temperature in the transparent containers was recorded at 20-min intervals (64K Pendant® Data Logger UA-002-64, HOBO, OneTemp Pty Ltd., Marleston, Australia) from mid-May to the end of July 2011. During this period, a minimum water volume of 600 ml was maintained.

Replacement series experiment

Experimental design

The interspecific larval interaction of Ae. albopictus and Cx. pipiens molestus under different food regimes and temperatures was investigated in a replacement series experiment using a randomized split-plot design. The replacement series experiment with 4 replications was designed to test the impact of the whole plot factors ‘species’ (2 levels) × ‘larval ratio’ (5 levels) × ‘temperature’ (3 levels) and the subplot factor ‘larval food regime’ (2 levels) on survival, development time and growth of the aquatic life stages of the 2 species.

Mosquito material

The mosquitoes used in the experiments were purchased from Biogents AG (Regensburg, Germany) and reared in-house. We used 2 well-adapted long-term laboratory strains (Ae. albopictus strain with origin Singapore, Cx. pipiens biotype molestus with origin Regensburg, Germany) to minimize unknown impacts of the field histories of the mosquito strains on their responses and better determine the fundamental nature of the context-dependency of interspecific interactions.

The eggs of Ae. albopictus were collected on filter paper, dried at 20 °C and 90% relative humidity, and exposed to a yeast solution stimulating larval hatching [33]. First-instars of Cx. pipiens molestus were released from egg rafts into 1:1 tap: deionized water (hereafter called larval medium). In total, 3600 larvae were investigated.

First-instar larvae of the two species, which had been synchronously released within 24–48 h, were allotted as pure or mixed cohorts of 30 larvae in 1 l test vessels (plastic cups) filled with 600 ml larval medium each.

Larval ratio

In total, 2 pure and 3 mixed larval groups were tested at 3 temperatures and under 2 food conditions. Following the recommendations of Oberg et al. [34], experimental units with only Ae. albopictus larvae (Ae: Cx30:00) and only Cx. pipiens molestus larvae (Ae: Cx00:30) and symmetrically and asymmetrically mixed treatments with a 20:10, 15:15 or 10:20 distribution of Ae. albopictus: Cx. pipiens molestus larvae (Ae: Cx20:10, Ae: Cx15:15, Ae: Cx10:20) were prepared. The larval density of 50 larvae l-1 reproduced natural larval densities: for instance, 1.2–89.5 larvae l-1 had been reported from various containers in Italy [18], 8.3–3080 larvae l-1 in tires in Florida [35], 1.4–80 larvae l-1 in buckets in North Carolina [36] and 0.6–62.4 larvae l-1 in tires and tree holes in Mississippi, USA [37].

Temperature

The experimental units were exposed to 15 °C, 20 °C and 25 °C at a 16:8 h light:dark photocycle in environmental chambers (MKKL 1200, Flohr Instruments GmbH, Utrecht, The Netherlands). These experimental temperatures had been derived from in situ measurements in potential mosquito microhabitats (Fig. 1).
Fig. 1

Physicochemistry in established and potential larval mosquito microhabitats. a Physicochemistry in natural habitats of Cx. pipiens in the periphery of Frankfurt am Main, Germany, during September and October 2010. Temperature (°C), pH, conductivity (μS cm-1) and dissolved oxygen concentration (mg l-1) (mean ± SD) and respective Cx. pipiens abundance [# larvae: number of larvae (n = number of microhabitats)] are shown. Grey bars denote overall 95% confidence intervals. Key: 1, car tires; 2, vases; 3, rain barrels; 4, small reservoirs (bucket, plastic tarpaulin, pot, steel girder); 5, large reservoirs (metal and plastic tub, excavator shovel, water bowl); 6, dung discharge; 7, puddles; 8, ponds. b Spline curve of daily water temperature [mean ± SD] in 1 litre plastic cups logged from mid-May to end of July 2011. The cups representing potential mosquito habitats had been set up in shaded versus sunny localities in Frankfurt am Main

Larval food supply

The larvae were fed with animal-based fish food (Tetra Min®, Tetra, Melle, Germany) using a total amount of either 3 mg larva-1 or 6 mg larva-1. Larval food was provided on days 0, 3, 6, 9, 12, 15, 18 and 21 in the 15 °C treatments, on days 0, 2, 4, 6, 8, 10, 12 and 14 in the 20 °C treatments, and on days 0, 2, 4, 5, 7, 8, 9 and 10 in the 25 °C treatments. Food portions of either 0.25 or 0.5 mg larva-1 were offered during the first to fourth feeding and 0.5 or 1 mg larva-1 at later ones. A total food quantity of 3 mg larva-1 was considered as limited (at least at 20 °C) and 6 mg larva-1 as adequate for Ae. albopictus larval development (at least at 20 °C and 25 °C; see Müller et al. [38]). The feeding protocol also allowed for an optimal development of Cx. pipiens molestus at 20 °C and 25 °C as shown by Kreß et al. [33].

Effect variables

Mortality, pupal development time and pupal growth parameters (size, weight) were examined. During the daily census, pupae were collected and preserved in 70% ethanol. Pupal abdominal length (AL, 3rd to 8th abdominal segment) was measured using a stereo microscope and the software DISKUS (Carl H. Hilgers, Königswinter, Germany) within 14 days after collection. The gender of the pupae was determined by examination of their genital lobes. Species identity was determined using 4 criteria because overlapping morphological traits were occasionally observed: (i) the colour of the prothoracic trumpets (respiratory tubes); (ii) general pigmentation; (iii) paddle marginal spicule hairs; and (iv) compaction of the habitus. After gender and species identification, individual pupae were dried at 60 °C for 14 ± 2 h, and their dry weight (DW) was measured using a microbalance (± 0.01 μg, Sartorius model 708501, Göttingen, Germany).

Physicochemistry in test vessels at 15 °C

Physicochemical effects on larval ratio, temperature and food regime were investigated in the medium of test vessels kept at 15 °C due to long algal growth. After the last pupation had occurred in a given replicate, the following parameters were quantified (in parentheses: test used): standing time of vessel (days until last pupation occurred), concentrations of chlorophyll a (algal cells > 0.45 μm; guideline DIN 38 412 [39]), silicon dioxide (LCW028 test, Hach-Lange, Düsseldorf, Germany), nitrite (Aquaquant/Aquamerck tests, Merck, Darmstadt, Germany), nitrate (LCK 339, Hach-Lange), ammonium (Aquaquant/Aquamerck, Merck), phosphate (Aquamerck, Merck), pH and conductivity (TetraCon 325 and SenTix, WTW, Weilheim, Germany).

In every environmental chamber used in this study, there had previously been some vessels with Scenedesmus obliquus cultures and outdoor mesocosm water which probably led to an aerial proliferation of algal cells. Therefore, 10 ml aliquots of test media kept at 15 °C were diluted (1:10) with tap water, and phytoplankton were collected for 3 h in Utermöhl chambers. The number of algal cells per ml and replicate was estimated based on the microscopic counting of 2 random fields of 116.28 mm2 (Axiovert 40 C microscope, ZEISS; DISKUS software, Carl H. Hilgers, Königswinter). Algae were identified to family or genus level according to Streble et al. [40] and Linne von Berg [41].

Data analysis

Data processing, graphing and statistics were performed using Microsoft Excel, GraphPad Prism version 7 and Statistica version 8 software. Effect variables and physicochemical measures are reported as mean ± standard deviation unless stated otherwise.

Data processing

Means of species- and gender-specific DWs and ALs were tabulated per replicate. Also, species-specific mortalities (%), gender-specific averages of the mean pupation time (PT50) and gender-specific daily biomass accumulation rates (BA) were calculated for every replicate. The PT50 indicated the time when 50% of male or female larvae had metamorphosed to pupae (for detailed calculations see Müller et al. [38]). The BA [mg larva-1 day-1] was calculated as the average for each gender and replicate by the division of the root of the mean DW and the root of the PT50. It should be noted that the BA neglected the initial DW of first-instar larvae.

Treatment-specific relative crowding coefficient values (RCC) were calculated with the AL, BA, DW and PT50 of female and male Ae. albopictus (RCCAe) and Cx. pipiens molestus (RCCCx) from the means of 4 replicates as described by Harper [42] and modified by Novak et al. [13] and Oberg et al. [34]: RCCspecies A = {[0.5 × (species A20:10/ species B20:10) + (species A15:15/ species B15:15) + 2 × (species A10:20/ species B10:20)]/3} / {species A30:00/ species B30:00}. From these parameter-specific RCCs, the difference between species was calculated for each gender and experimental group to improve the comparability of parameter-specific competitiveness between 2 species [43]. Also, integrated RCCs for every gender and species and every experimental group were calculated from 3 parameter-specific RCCs [DW-RCC, AL-RCC, PT50-RCC; mean ± SD].

Statistics

The mosquito and physicochemical datasets were tested for Gaussian distribution using the Kolmogorov-Smirnov test and the D’Agostino Pearson omnibus normality test (P ≤ 0.01). Variance homogeneity of the data was tested using the Cochran’s test at the level of P ≤ 0.01. After confirmation of variance homogeneity for arcsin and log-transformed mortality [%], log-transformed PT50 data (gender-specific) and parameter-specific RCC values, four-factorial ANOVAs (independent factors: species, larval, temperature, food regime) and subsequent Tukey HSD tests were computed for these response variables. The heteroscedastic, but normally distributed dependent variables AL, BA and DW were analyzed with general linearized normal-log models with backward factor removal during model building and subsequent Wald statistics. The total average mortality of the two species (irrespective of experimental treatment) was compared with a t-test.

For reasons of straightforward whole-plot interpretation, polynomic regressions of second order (least squares fit) were computed with mortality data against larval ratio. Also, first order regressions (least squares fit) were computed with AL, DW and BA data against the number of larvae of the other species, respectively. Extra-sum-of-squares-tests were applied to compare independent fits with global fits for mosquito responses to food regime within a temperature block (mortality, corrected by Benjamini-Hochberg (BH) procedure with false discovery rate (FDR) of 0.05) and independent fits with global fits for each species and gender and within each temperature (AL, BA, BH corrected with FDR of 0.05), respectively.

Water quality parameters examined at 15 °C were tested by 2-way ANOVA (P < 0.05) for their dependence on initial larval ratio and food. Water quality parameters were further tested for correlation with algal growth (Spearman). The dependence of algal growth at 15 °C from initial larval ratio or larval mortality and food regime and their interactions were tested with 2-way ANOVAs. The algal growth data at 15 °C were tested for Spearman's correlation with Ae. albopictus mortality at 15 °C and Cx. pipiens mortality at 15 °C, respectively. Also, non-parametric Spearman's correlation analyses with the initial larval quantity of Ae. albopictus or Cx. pipiens molestus and the cell number of specific algal groups were performed for both food-related subplots.

Results

Larval mosquito habitats

During the field survey in the Rhine-Main metropolitan region, 170 intermittent and 8 permanent water bodies were identified as mosquito microhabitats. In total, 1072 mosquito larvae (606 Cx. pipiens (s.l.) (see Fig. 1a), 400 Culex torrentium, 3 Culex hortensis, 1 Culex territans, 37 Culiseta annulata, 1 Aedes geniculata, 3 Anopheles maculipennis, and 12 Anopheles plumbeus) were collected. The physicochemical parameters of a subset of Cx. pipiens (s.l.) positive microhabitats (n = 70) of mainly anthropogenic origin (88.6%) averaged to 15.0 ± 4.56 °C, 6.74 ± 4.02 mg l-1 dissolved oxygen, 390 ± 574 μS cm-1 electrical conductivity and pH 7.60 ± 0.84 (Fig. 1a).

Outdoor versus experimental temperature

From mid-May to the end of July 2011, the water temperature in 1 l plastic cups was on average 19.2 ± 1.8 °C at the shaded site and 20.6 ± 4.8 °C at the sunny site (Fig. 1b). Minimum daily average temperatures were 14.2 °C and 14.1 °C in the shaded and sun-exposed test vessel, and maximum daily average temperatures, 25.3 °C and 27.9 °C, respectively. Thus, the temperatures chosen for the replacement series experiment (15, 20 and 25 °C) mirrored the minimum, mean and maximum daily average water temperatures in potential mosquito microhabitats during spring and summer in Frankfurt am Main, Germany.

Multifactorial impact on mosquito response

General factorial response

The single factors species and temperature influenced every tested effect variable significantly (ANOVA, Table 1). This was analogous to the factor larval food regime except its non-significance for PT50 and male BA. In contrast, the single factor larval ratio had a significant influence on mortality and PT50 only. At the level of two-factorial interactions, food × temperature was always significant and species × food and species × temperature almost always significant for mosquito response variables (16 out of 18 sub-models). In contrast, the larval ratio in combination with species, food or temperature was significant in only 8 out of 27 sub-models. Three-factorial interactions of species × food × larval ratio strongly influenced the data (except AL and male DW), as did species × food × temperature (except BA). The triple interaction species × larval ratio × temperature had a significant influence on mortality, male PT50 and AL, whereas food × larval ratio × temperature had no significant influence on any effect variable. The four-factorial interaction was significant for female BA, female PT50 and male DW only (Table 1).
Table 1

Results for four-factorial significance tests (df, degree of freedom) with the independent factors species (Aedes albopictus, Culex pipiens), food regime (3 and 6 mg larva-1 food supply), larval ratio (0, 10, 15 and 20 larvae of the other species) and temperature (15, 20 and 25 °C). The dependent variables mortality (arcsin log-transformed) and female/male PT50 (log-transformed) were tested with a general linear model (ANOVA, F value) whereas the effect variables female/male abdominal length (AL), dry weight (DW) and biomass accumulation (BA) were tested with a general linearized model (normal-log model with backward removal, Wald value)

  

Mortality

Female PT50

Male PT50

Female AL

Male AL

Female DW

Male DW

Female BA

Male BA

 

df

F

P

F

P

F

P

Wald

P

Wald

P

Wald

P

Wald

P

Wald

P

Wald

P

Intercept

1

1082

<0.001

130948

<0.001

117040

<0.001

83928

<0.001

67157

<0.001

3431

<0.001

21209

<0.001

49678

<0.001

51860

<0.001

Species (S)

1

5.66

0.019

64.00

<0.001

94.01

<0.001

105.6

<0.001

10.58

0.001

562.6

<0.001

1247

<0.001

58.61

<0.001

76.22

<0.001

Food (F)

1

6.26

0.014

3.18

0.076

1.65

0.201

87.39

<0.001

15.03

<0.001

48.55

<0.001

6.20

0.013

4.99

0.026

0.15

0.699

Larval ratio (L)

3

5.20

0.002

3.55

0.016

2.55

0.058

3.23

0.358

3.66

0.301

11.36

0.010

3.75

0.289

7.24

0.065

1.39

0.708

Temperature (T)

2

39.44

<0.001

1839

<0.001

1823

<0.001

71.84

<0.001

152.74

<0.001

127.87

<0.001

79.36

<0.001

1092

<0.001

787.2

<0.001

S × F

1

4.91

0.029

0.88

0.350

15.09

<0.001

53.44

<0.001

8.53

0.003

31.08

<0.001

35.89

<0.001

11.98

0.001

17.38

<0.001

S × L

3

9.89

<0.001

0.82

0.486

1.29

0.280

7.60

0.055

4.41

0.220

14.20

0.003

13.64

0.003

3.43

0.330

3.81

0.283

F × L

3

1.04

0.377

2.34

0.076

2.34

0.076

17.62

0.001

7.57

0.056

2.67

0.445

9.20

0.027

0.44

0.932

2.01

0.570

S × T

2

0.83

0.436

54.59

<0.001

47.60

<0.001

83.33

<0.001

45.81

<0.001

70.79

<0.001

57.32

<0.001

65.30

<0.001

38.20

<0.001

F × T

2

16.80

<0.001

25.62

<0.001

25.95

<0.001

16.87

<0.001

32.41

<0.001

76.47

<0.001

73.05

<0.001

40.15

<0.001

17.68

<0.001

L × T

6

1.78

0.108

0.77

0.596

0.28

0.945

16.47

0.011

15.32

0.018

6.32

0.388

14.85

0.021

10.61

0.101

5.46

0.486

S × F × L

3

3.45

0.019

5.24

0.002

7.98

<0.001

0.75

0.861

7.66

0.053

10.92

0.012

6.41

0.093

11.04

0.011

9.35

0.025

S × F × T

2

7.58

0.001

11.66

<0.001

6.07

0.003

10.04

0.007

37.00

<0.001

46.56

<0.001

38.93

<0.001

0.47

0.791

0.99

0.610

S × L × T

6

3.32

0.005

1.76

0.112

2.33

0.036

27.13

<0.001

14.08

0.029

6.25

0.395

9.86

0.131

5.26

0.511

8.18

0.225

F × L × T

6

0.63

0.709

0.85

0.532

0.77

0.594

4.29

0.637

5.29

0.507

9.00

0.174

12.33

0.055

9.99

0.125

0.93

0.988

S × F × L × T

6

0.67

0.670

2.94

0.010

1.21

0.307

5.05

0.537

4.40

0.622

3.52

0.741

15.07

0.020

15.45

0.017

6.45

0.375

Mortality

The split-plot design used in the 4-factorial replacement series experiment resulted in a larval mortality within the margins of expectation for intra- and interspecific competition with a larval density of 50 larvae l-1 (Fig. 2). The total average mortality (mean ± standard error of the mean; CV - coefficient of variation; n = 96) of Ae. albopictus (34.9 ± 2.67%; 74.8%) was significantly higher than the total average mortality of Cx. pipiens molestus (25.2 ± 1.94%; 75.4%; t(95) = 13.0, P < 0.0001). The highest mortality of Ae. albopictus (66.2 ± 6.04%) and Cx. pipiens molestus (39.6 ± 4.39%) was observed at 15 °C and 6 mg larva-1 food supply. The Cx. pipiens molestus larvae had an equally high mortality (39.6 ± 6.14%) at 25 °C and 3 mg larva-1 food supply. The lowest mortality of Ae. albopictus was 8.54 ± 2.46% and that of Cx. pipiens molestus 10.0 ± 2.17%, both in cohorts exposed to 20 °C and supplied with 6 mg larva-1 food.
Fig. 2

Larval mortality [mean, SD] of Aedes albopictus and Culex pipiens in pure and mixed cohorts (x-axis - the number of Ae. albopictus: Cx. pipiens larvae) in dependence of temperature and larval food regime. Squares symbolize artificially set zero values; solid lines and broken lines - non-linear regression (second-order model); dotted lines - 95% confidence interval

Mortality was mainly influenced by temperature. Further variance was produced by the factors food, species and larval ratio, by the 2-factorial interactions food × temperature, species × larval ratio, species × food, and the 3-factorial interactions species × food × temperature, species × food × larval ratio and species × larval ratio × temperature (Table 1). Between species, regression lines did differ slightly among cohorts fed 6 mg larva-1 and exposed to 20 °C (F(3.34) = 1.87, BH-corrected P = 0.05), and those fed 3 mg larva-1 at 25 °C (F(3.34) = 3.78, BH-corrected P = 0.042). In the other 4 temperature-food treatments, the mortality in pure and mixed cohorts differed strongly between species (15 °C, 3 mg larva-1: F(3.34) = 5.86, BH-corrected P = 0.025; 15 °C, 6 mg larva-1: F(3.34) = 3.78, BH-corrected P = 0.008; 20 °C, 3 mg larva-1: F(3.34) = 3.78, BH-corrected P = 0.033; 25 °C, 6 mg larva-1: F(3.34) = 3.78, BH-corrected P < 0.017; Fig. 2).

The presence of Cx. pipiens molestus larvae significantly increased the mortality of Ae. albopictus in the cohorts fed 6 mg larva-1 at 25 °C (Ae:Cx15:15 vs Ae:Cx30:00, Tukey HSD MS(126) = 0.55, P = 0.046) and 3 mg larva-1 at 20 °C (Ae:Cx15:15 vs Ae:Cx30:00, Tukey HSD MS(126) = 0.55, P = 0.023) and 25 °C (Ae:Cx15:15 vs Ae:Cx30:00, Tukey HSD MS(126) = 0.55, P = 0.009; Ae:Cx20:10 vs Ae:Cx30:00, Tukey HSD MS(126) = 0.55, P = 0.002). The species × food × larval ratio interaction on mortality was demonstrated by several significant differences between food-related Ae. albopictus cohorts: The Ae:Cx15:15 cohort that had been fed 3 mg larva-1 differed from the cohorts Ae:Cx10:20, Ae:Cx20:10 and Ae:Cx30:00 provided with 6 mg food larva-1 at 20 °C (Tukey HSD MS(126) = 0.55, P = 0.004, P = 0.039, P = 0.001, respectively); the Ae:Cx15:15 cohort fed 3 mg larva-1 from the Ae:Cx 30:00 cohort fed 6 mg larva-1 at 25 °C (Tukey HSD MS(126) = 0.55, P = 0.039), and the Ae:Cx30:00 cohort fed 3 mg larva-1 from the Ae:Cx15:15 cohort fed 6 mg larva-1 at 25 °C (Tukey HSD MS(126) = 0.55, P = 0.010). In contrast, the presence of Ae. albopictus larvae did not influence the mortality of Cx. pipiens molestus (Tukey HSD MS(126) = 0.55, P > 0.05).

Mean pupation time (PT50)

Species and temperature were the major factors affecting the PT50 of male and female larvae (Table 1). Also, a larval ratio significantly influenced the PT50 of female larvae while the single factor food had no significant effect (P > 0.08). The PT50 of male and female larvae was additionally influenced by the 2-factorial interactions species × temperature, food × temperature and species × food and the 3-factorial interactions species × food × larval ratio and species × food × temperature. The triple interaction species × larval ratio × temperature had a significant effect on the PT50 of male, and the 4-factorial combination on that of female larvae only.

The PT50 of both genders and species decreased as a function of temperature, with maxima at 15 °C and minima at 25 °C (Fig. 3). At 25 °C, a food supply of 3 mg larva-1 decreased the mean PT50 of females by 2.1 days (Ae. albopictus) and 0.7 days (Cx. pipiens molestus) and the mean PT50 of males by 2.1 days (Ae. albopictus) and 0.5 days (Cx. pipiens molestus) if compared to the cohorts fed 6 mg larva-1 at 25 °C. At lower temperatures, the PT50 had an opposite trend with a higher average PT50 of females of 1.9 days (Ae. albopictus) and 0.8 days (Cx. pipiens molestus) at 15 °C and 1.2 days (Ae. albopictus) at 20 °C if larvae had been supplied with 3 mg food larva-1, although the average PT50 of Cx. pipiens molestus was similar in both food treatments at 20 °C.
Fig. 3

Mean pupation time PT50 (days; box-plot with Tukey whiskers, mean shown as ‘+’) of Aedes albopictus and Culex pipiens in dependence of gender, the larval ratio (number of Ae. albopictus: Cx. pipiens larvae - Ae: Cx), temperature and larval food regime

This general pattern of 2- and 3-factorial interactions of species × food × temperature equally applied to the average PT50 of male larvae that had received 6 mg food larva-1, which was 0.3 days (Ae. albopictus) and 1.6 days (Cx. pipiens molestus) shorter at 15 °C and 0.7 days (Ae. albopictus) and 1.1 days (Cx. pipiens molestus) shorter at 20 °C than the average PT50 of males that had been fed with 3 mg larva-1. With regard to larval ratio, the PT50 of females decreased with an increasing larval ratio of the competing species, but this was only observed in the cohorts of Ae. albopictus provided with 3 mg food larva-1 and those of Cx. pipiens molestus given 6 mg larva-1.

Pupal growth

Pupal growth (AL, DW) was mainly influenced by the single factors species, temperature and food and their 2- and 3-factorial interactions (Table 1). The larval ratio had a significant effect only if it interacted with the factors species (DW), food (AL of females, DW of males) and temperature (AL, DW of males) or in concert with species × food (DW of females) and species × temperature (AL). In the following, the detailed response patterns are described exclusively for the pupal growth of females due to their biological significance for size/weight-fecundity-relationships [21]. However, male response patterns were similar.

The AL of female Ae. albopictus was generally greater (3.12 mm) than that of female Cx. pipiens molestus (2.89 mm), although it was equal at 20 °C and a food supply of 6 mg larva-1 (F(2.28) = 1.25, P = 0.30), and at 15 °C and 3 mg food larva-1 (F(2.27) = 3.14, P = 0.06) (Additional file 2: Figure S2). The greatest AL of female Ae. albopictus was measured in the cohort fed with 3 mg larva-1 at 20 °C, followed by the cohort given 6 mg larva-1 at 20 °C, the significantly different larval food regimes at 25 °C (6 mg larva-1: 3.26 ± 0.15 mm; 3 mg larva-1: 2.99 ± 0.12 mm, F(2.26) = 16.01, P < 0.0001) and finally by both larval food regimes at 15 °C.

The AL of female Cx. pipiens molestus differed significantly among the larval food regimes at every tested temperature (15 °C: F(2.27) = 17.26, P = 0.0001; 20 °C: F(2.28) = 12.25, P = 0.0002; 25 °C: F(2.27) = 23.39, P = 0.0002). The maximum AL of female Cx. pipiens molestus was measured in the cohort with 6 mg food larva-1 at 15 °C, followed by the groups fed 6 mg larva-1 and 3 mg larva-1 at 20 °C, the 3 mg larva-1 cohort at 15 °C, and finally by both larval food regimes at 25 °C.

The mean DW of female Cx. pipiens molestus (0.80 mg) was higher than that of female Ae. albopictus (0.58 mg). In both species, the DW of females was always largest in the cohorts with 6 mg larva-1 food supply (Additional file 3: Figure S3). In the cohorts fed 6 mg larva-1, the DW of Ae. albopictus increased with increasing temperature, whereas the DW of Cx. pipiens molestus decreased with increasing temperature. In the cohorts provided with 3 mg food larva-1, there was no temperature-dependent change of DW. By overall comparison, species-specific differences in DW were minor at 25 °C under both food regimes. In the Cx. pipiens molestus cohorts given 6 mg food larva-1 at 15 °C and 20 °C, a decreased DW became apparent as heterospecific larval numbers increased, but this was not the case at 25 °C nor for Ae. albopictus (Additional file 3: Figure S3).

Daily increase in weight (BA)

The daily increase of weight (BA) was mainly influenced by the factors species (mean BA Ae = 0.053 mg larva-1 day, mean BA Cx = 0.058 mg larva-1 day) and temperature (Table 1). Minimum and maximum values of BA were always found at 15 °C and 25 °C (cohorts fed 3 mg larva-1), demonstrating the strong impact of temperature on BA. The single factor food had a significant influence on females only, but food × species or food × temperature and the 3-factorial interaction species × food × temperature had significant effects on both males and females. The BA of both species was similar to 3 mg larva-1 food supply except Ae. albopictus at 25 °C, but the BA was lower in Ae. albopictus than Cx. pipiens molestus cohorts fed with 6 mg larva-1.

Similar to AL, the BA of Ae. albopictus fed 6 mg larva-1 at 25 °C slightly decreased with an increasing number of Cx. pipiens molestus larvae, whereas the BA of those fed 3 mg larva-1 increased with an increasing number of Cx. pipiens molestus larvae (Fig. 4). In contrast, the BA of Cx. pipiens molestus did not change with different heterospecific larval ratios. This complex pattern led to larval ratio having a significant influence only within the combination species × food × larval ratio or within the 4-factorial interaction (Table 1).
Fig. 4

Daily accumulation of biomass (BA, mean ± SD) of individual female Aedes albopictus and Culex pipiens during their aquatic part of life in dependence of larval ratio, temperature and larval food regime. Solid line - linear regression; dotted lines - 95% confidence interval

Relative crowding coefficient (RCC)

Both species exhibited a weak to intermediate interspecific competitiveness with regard to growth and development as indicated by RCC values between 0.91 and 2.31 (Table 2; Additional file 4: Table S1; [18, 43]). Relative crowding coefficient values < 1.39 were observed in 38 out of 96 experimental groups indicating no competition between species in 39.6% of treatments (Additional file 4: Table S1). Values between 1.4 to 2.31 indicative of low to intermediate competition were most frequently measured (56.3% for RCCAe, 58.3% RCCCx), whereas RCC values ≥ 2.5 signifying high competition were never observed.
Table 2

Relative crowding coefficients (RCCs). Integrated relative crowding coefficient values for female and male Aedes albopictus and Culex pipiens in different experimental treatments indicate a generally low competition between species regarding growth and development due to equilibration of the traits of different species. To evaluate the predomination of one species trait over the other in more detail, the differences between RCCAe and RCCCx that are based on biomass accumulation rate (BA), pupal dry weight (DW), pupal abdominal length (AL) and mean pupation time (PT50) are additionally reported, whereas the respective parameter-specific RCCs can be found in the Additional file 4: Table S1

Larval exposure

Integrated RCCAe (mean ± SD)

Integrated RCCCx (mean ± SD)

AL-RCCAe - AL-RCCCxa

DW-RCCAe - DW-RCCCxa

PT50-RCCAe - PT50-RCCCxa

BA-RCCAe - BA-RCCCxa

6 mg larva-1

15 °C

1.57 ± 0.07

1.34 ± 0.06

0.38

0.16

0.15

-0.29

20 °C

1.52 ± 0.15

1.38 ± 0.12

-0.04

0.46

0.03

-0.50

25 °C

1.67 ± 0.35

1.28 ± 0.24

-0.03

0.13

1.07

-1.35

3 mg larva-1

15 °C

1.40 ± 0.19

1.51 ± 0.19

0.31

-0.38

-0.28

0.75

20 °C

1.45 ± 0.06

1.45 ± 0.07

0.04

0.11

-0.15

0.05

25 °C

1.28 ± 0.13

1.64 ± 0.17

-0.18

-0.20

-0.70

0.88

6 mg larva-1

15 °C

1.59 ± 0.14

1.32 ± 0.12

0.00

0.50

0.32

-0.84

20 °C

1.52 ± 0.19

1.39 ± 0.18

-0.29

0.33

0.36

-0.66

25 °C

1.72 ± 0.46

1.27 ± 0.29

-0.07

0.11

1.32

-1.40

3 mg larva-1

15 °C

1.38 ± 0.11

1.52 ± 0.11

0.12

-0.30

-0.22

0.50

20 °C

1.42 ± 0.36

1.53 ± 0.35

-0.32

0.68

-0.67

0.02

25 °C

1.40 ± 0.02

1.49 ± 0.02

-0.08

-0.06

-0.13

0.19

aA negative value indicates the superiority of a Cx. pipiens trait and positive value the superiority of an Ae. albopictus trait

The pattern of interspecific competitiveness described by parameter-specific RCCs was similar between genders but altered by larval exposure conditions and the parameter. The number of groups with the superiority of Ae. albopictus or Cx. pipiens molestus (groups with negative versus positive parameter-specific RCC differences) was balanced (Table 2). Disregarding temperature, PT50 and BA produced a mirrored RCC difference pattern with the dominance of the PT50-RCC and a parallel lesser BA-RCC in the Ae. albopictus cohorts fed 6 mg larva-1 and the Cx. pipiens molestus cohorts given 3 mg larva-1, respectively. The DW-RCC of Ae. albopictus was larger in the Ae. albopictus cohorts fed 6 mg larva-1 and those fed 3 mg larva-1 at 20 °C. The AL-RCC was higher in Ae. albopictus than in Cx. pipiens molestus cohorts at 15 °C, but the AL-RCC of Cx. pipiens molestus became higher than in Ae. albopictus at a higher temperature.

The integration of RCCs revealed that differences from growth and development specific RCCs were almost balanced out given that the single factors species, gender, food and temperature and almost all interactions had no significant influence. Notably, the integrated RCCAe was higher in the 6 mg larva-1 food treatment and the integrated RCCCx in the 3 mg larva-1 food treatment (species × food interaction, P < 0.001).

Water parameters in test media of 15 °C treatments

The total standing time of test vessels in parallel with the maximum pupation time was altered by larval ratio and larval food regime: The longest standing time of 40 ± 0 days was observed in the Ae:Cx00:30 cohort provided with 6 food mg larva-1and the shortest standing time of 33 ± 2 days in the Ae:Cx00:30 cohort fed with 3 mg larva-1 (Table 3). Physicochemical water parameters depended significantly on the larval food regime (ammonium, nitrate, nitrite, silicon dioxide, oxygen, pH), the larval ratio (nitrate) and their interaction (silicon dioxide). At the end of the half-life-cycle tests where larvae had been fed with 6 mg larva-1, the concentrations of nitrate (2.4 to 6-fold) and potentially toxic nitrite (2-fold) were lower than in the water of 3 mg larva-1 cohorts. In contrast, concentrations of ammonium (1.7 to 18-fold), silicon dioxide (1.8 to 5.6-fold; apart from 0.45-fold in the Ae:Cx30:00 treatment), oxygen (1.1 to 1.3-fold) and pH (1.2 to 1.3-fold) were higher than in the water of cohorts fed with 3 mg larva-1 (Table 3).
Table 3

Physicochemistry of test media at 15 °C. Chemical, physical and biological parameters (mean ± SD) and standing time (d, mean ± SD) of test vessels during the half life-cycle experiment with Aedes albopictus and Culex pipiens based on the larval ratio (number of A. albopictus: C. pipiens larvae; Ae:Cx) and food quantity (3 or 6 mg larva-1 food supply)

Ae:Cx

NO2

NO3

NH4

PO4

pH

Con

O2

Chl a

SiO2

ST

6 mg larva-1 food supply

00:30

0.01 ± 0.00

1.16 ± 0.15

0.88 ± 0.25

2.50 ± 0.58

9.92 ± 0.24

271.5 ± 10.79

15.02 ± 2.99

0.42 ± 0.28

4.53 ± 1.00

40 ± 0

10:20

0.01 ± 0.00

2.72 ± 2.33

0.75 ± 0.29

2.50 ± 0.91

9.75 ± 0.36

297.0 ± 16.02

15.75 ± 1.89

0.24 ± 0.12

1.90 ± 1.54

36 ± 4

15:15

0.01 ± 0.00

0.93 ± 0.52

0.75 ± 0.29

2.50 ± 1.29

9.98 ± 0.13

271.3 ± 16.21

18.18 ± 0.84

0.12 ± 0.03

4.10 ± 2.19

35 ± 5

20:10

0.01 ± 0.00

0.94 ± 0.22

0.75 ± 0.29

3.13 ± 0.85

9.86 ± 0.30

269.7 ± 42.62

17.12 ± 2.30

0.39 ± 0.34

2.70 ± 1.90

38 ± 3

30:00

0.01 ± 0.01

0.84 ± 0.22

2.00 ± 2.0

4.00 ± 1.41

9.78 ± 0.38

313.3 ± 34.73

14.41 ± 2.87

0.41 ± 0.09

0.54 ± 0.16

36 ± 3

3 mg larva-1 food supply

00:30

0.02 ± 0.01

6.15 ± 0.91

0.05 ± 0.00

3.00 ± 0.00

7.97 ± 0.04

238.5 ± 16.05

13.25 ± 0.91

0.88 ± 0.58

0.81 ± 0.03

33 ± 2

10:20

0.02 ± 0.01

6.49 ± 0.97

0.05 ± 0.00

3.00 ± 0.00

7.91 ± 0.07

270.3 ± 55.45

13.05 ± 0.56

0.36 ± 0.15

1.05 ± 0.03

37 ± 1

15:15

0.02 ± 0.01

5.59 ± 0.60

0.23 ± 0.20

3.00 ± 0.00

7.53 ± 0.11

258.5 ± 13.99

13.67 ± 2.28

0.27 ± 0.19

0.92 ± 0.07

39 ± 1

20:10

0.02 ± 0.01

5.34 ± 0.73

0.45 ± 0.06

3.00 ± 0.00

7.97 ± 0.08

255.5 ± 12.47

13.50 ± 0.20

0.36 ± 0.22

0.91 ± 0.08

38 ± 1

30:00

0.02 ± 0.01

3.03 ± 0.71

0.45 ± 0.06

3.00 ± 0.00

8.18 ± 0.04

259.0 ± 23.76

12.99 ± 1.11

0.14 ± 0.05

1.20 ± 0.15

39 ± 1

Abbreviations: NO 2 - nitrite (mg l-1), NO 3 - nitrate (mg l-1), NH 4 + ammonium (mg l-1), PO 4 2- phosphate (mg l-1); pH (acidic or basic character), Con conductivity (μS cm-2), O 2 oxygen content (mg l-1), Chl a, chlorophyll a (mg l-1), SiO 2 silicon dioxide (mg l-1), ST standing time maximum pupation time (d)

In correlation to increased oxygen (Spearman's r(10) = -0.588, P = 0.04), conductivity (Spearman's r(10) = -0.758, P = 0.006), pH (Spearman's r(9) = -0.720, P = 0.014), ammonium (Spearman's r(10) = -0.741, P = 0.007) and decreased nitrate (Spearman's r(10) = 0.842, P = 0.001) and nitrite (Spearman's r(10) = 0.801, P = 0.008), algal growth was significantly reduced in the water with cohorts fed 6 mg larva-1 (F(1.50) = 116.3,, P < 0.001, 46.6% total variation) (Fig. 5). Notably, algal growth was negatively correlated to Ae. albopictus mortality (Spearman's r(8) = -0.86, P = 0.011). Algal growth especially declined with an increasing number of initial Ae. albopictus larvae (larval ratio: F(4.50) = 13.17, P < 0.001, 21.1% total variation; food × larval ratio: F(4.50) = 4.58, P = 0.003, 7.3% total variation).
Fig. 5

Algal growth in test vessels at 15 °C. Algal cells (mean and SD) grown in test media with pure and mixed cohorts of Aedes albopictus and Culex pipiens larvae in dependence of larval ratio (Ae:Cx - number of Ae. albopictus: Cx. pipiens larvae) and larval food regime with (a) 3 mg larva-1 larval food supply versus (b) 6 mg larva-1 larval food supply at 15 °C. Algal growth was not observed in test vessels at 20 °C and 25 °C

Discussion

In the present study, we show that the physicochemical and microclimatic conditions of potential and actual mosquito breeding sites in Germany offer suitable conditions for a balanced coexistence and frequent competitive superiority of Cx. pipiens molestus over Ae. albopictus with regard to larval and pupal survival. We demonstrate that the interspecific competition pattern of these two mosquito species does not directly depend on the thermal regime (rejection of hypothesis i), but is rather controlled by food regime and species-food and species-temperature interactions (support of hypothesis ii). Furthermore, we discuss how the resource competition between Ae. albopictus and Cx. pipiens molestus is directly connected to species-specific foraging behaviour and physicochemical as well as phycological microhabitat parameters (support of hypothesis iii).

Larval coexistence under present climatic conditions in Germany

In general agreement with our data, Ae. albopictus and Cx. pipiens molestus had an almost identical population growth at 20 °C but a divergent one at a higher temperature [33]. A simple extrapolation of the laboratory optimum to mosquito co-occurrence in the field suggests that microhabitats in Germany with a typical median temperature of 20 °C in spring and summer offer favourable conditions for the coexistence of these two species, even if the food regime may ultimately restrict the success of Ae. albopictus in mixed larval habitats. The longer development time of Ae. albopictus at 15 °C and Cx. pipiens molestus at 20 °C and 25 °C will directly translate to the population level [44] suggesting a shift from Cx. pipiens (s.l.) with advantaged population activity at 15 °C to Ae. albopictus at a higher temperature. During warmer seasons, Ae. albopictus may make up its population delay by faster development time, faster growth and superior biomass accumulation. This experimentally based expectation has support from field observations elsewhere [18, 27, 28]: In Italy, for example, the population growth of Cx. pipiens (s.l.) starts in mid-April and peaks in July, and that of Ae. albopictus in mid-May and September, respectively [18]. Marini et al. [28] calculated for Ae. albopictus an average delayed population growth by 29 days when compared with Cx. pipiens (s.l.) from the same location and year.

Our results indicate a general adaptation of Cx. pipiens to low temperatures although the thermal optimum of Cx. pipiens (s.l.) is between 20 °C and 25/27 °C (present study, [45, 46]). In contrast, the higher total average mortality of Ae. albopictus and the especially high mortality at 15 °C suggest that Ae. albopictus is less adapted to colder ecoregions than its autochthonous competitor. However, despite these differences in thermal biology, the outcomes from available competition studies for Ae. albopictus and Cx. pipiens (s.l.) might be transferrable to colder ecoregions [18, 27], because the temperature did not influence the interspecific pattern at other than optimal test conditions. We, therefore, reject hypothesis i, that a competitive superiority depends on the species-specific thermal tolerance spectrum and therefore Ae. albopictus might be advantaged at higher and Cx. pipiens (s.l.) at lower temperatures. Similarly, temperatures ranging from 24 to 30 °C did not change the larval competition between Ae. albopictus and A. aegypti [6], and a range of 15 °C to 31 °C did not alter the larval competition between Ae. albopictus and O. triseriatus [5].

The competitive superiority of Culex pipiens over Aedes albopictus - or vice versa?

All available evidence from this study suggests a competitive superiority of Cx. pipiens molestus over Ae. albopictus with regard to survival at every other temperature and food treatment than 20 °C and 6 mg larva-1. Agonistic growth and development strategies levelled out the parameter-specific sublethal crowding effects. Mortalities comparable to the ones registered in the present study have been observed in Cx. pipiens (s.l.) cohorts regardless of the presence or absence of larvae of a second mosquito species, whereas the presence of Cx. pipiens larvae especially affected Ae. albopictus survival and biomass accumulation at 25 °C and 3 mg larva-1 food supply. Carrieri et al. [18] observed balanced survival and development in their Ae. albopictus and Cx. pipiens (s.l.) cohorts exposed to 25 °C with 2.85 mg larva-1 food supply, but growth differed significantly between species indicating a slightly suboptimal food supply especially for Cx. pipiens (s.l.). As an additional contrast to our study, the powerful competitive superiority of Ae. albopictus over Cx. pipiens (s.l.) became apparent at poorer larval food conditions at 20 °C and 25 °C [18]. Likewise, Costanzo et al. [27] reported competitive superiority of Ae. albopictus over Cx. pipiens (s.l.).

The contrary outcomes from the replacement series experiments in this study and those by Carrieri et al. [18] and Costanzo et al. [27] are not expected to result from the use of different Ae. albopictus mosquito strains [47, 48] but more likely from the investigation of different members of the Cx. pipiens complex with differential autecological characteristics [26]. The autogenic and stenogamic biotype Cx. pipiens molestus shows a homodynamic development, strong synanthropic behaviour, mammal feeding type and urban to suburban distribution [30, 49, 50]. This ecotype often occupies biotopes different from those of the ornithophilic Cx. pipiens pipiens in regions with cold temperate climate (hypogeous, [30]), although Cx. pipiens molestus is not excluded from epigeous breeding sites in the Rhine rift valley and warm temperate European regions [49, 51, 52].

However, resource competition may give a more reliable explanation for the opposite competition pattern observed between the present study and those by Carrieri et al. [18] and Costanzo et al. [27], because different types of food were supplied (animal-based food versus plant-based detritus), species-specific food quantity responses can be assumed [17, 18] and differential food quantity-temperature interactions may have taken place [5, 33, 38].

Larval food supply is decisive for competitive superiority

Hypothesis ii stating food regime is a major determinant for interspecific interactions with temperature becomes supported by the comparison of our data with literature. In an overall comparison of three competition studies (present study, [18, 27]), the food supply in our experiment was perhaps always ad libitum as implied by evenly balanced biomass accumulation rates. On the other hand, pupal sizes and weights were always larger in the cohorts fed with 6 mg larva-1. This surplus of food may especially have favoured Cx. pipiens molestus in our study, whereas the progressively limited food resources in the studies by Carrieri et al. [18] and Costanzo et al. [27] may have resulted in a competitive superiority of Ae. albopictus. Similarly, O. triseriatus had a better survival than Ae. albopictus in water sampled from tree-holes with significant detritus whereas probably limited resource levels in tire water favoured Ae. albopictus [21]. In another replacement series experiment, the larval superiority of Ae. albopictus over O. triseriatus was observed under 0.5 mg but diminished under two mg larval food supply [5].

Food quality could have alternatively triggered the different competitive interactions between Ae. albopictus and Cx. pipiens molestus given that spatiotemporal natural variation in detritus quality (in terms of the animal to plant-based ratio and associated microbial growth) can shape mosquito assemblages [53]. It has been shown that the detritus type, its decay rate and associated microbial growth influence the interspecific competition between Ae. albopictus and O. triseriatus and Ae. aegypti, respectively [17, 53]. It has also been shown that Ae. aegypti is weaker in competition than Ae. albopictus if larvae feed on oak, pine, or insect detritus (related to low to intermediate microbial growth) but face competition if feeding on grass which is related to high microbial growth [17]. Aedes albopictus has been described as an opportunistic feeder [54] with great resistance to starvation [14], but animal-based detritus and associated microorganisms always yield higher performance of Ae. albopictus [20, 38, 53, 55, 56].

Under resource-limiting conditions (limiting at least for Cx. pipiens molestus), Ae. albopictus with its continuously foraging behaviour may usurp detritus, and specifically essential nutrients, more quickly than the selectively feeding larvae of Cx. pipiens (s.l.) which rest between meals [22, 56, 57]. Culex pipiens molestus larvae are known to react to phagostimulants released from high-value food which may allow the species to control foraging and filtering activity to usurp mostly food particles with high nutritional value [57, 58]. However, the opportunistic features of Ae. albopictus might not be successful in microhabitats with eutrophic conditions, as observed at 25 °C and 6 mg larva-1 food supply where the BA of Ae. albopictus even decreased, and the PT50 increased if compared to 3 mg larva-1 cohorts.

Phycology is a crucial factor for Ae. albopictus survival

Our study demonstrates that resource competition between Ae. albopictus and Cx. pipiens molestus is directly connected to physicochemical and phycological parameters of their microhabitat (hypothesis iii). The competitive disadvantage of Ae. albopictus with lower survival under eutrophic conditions became very substantial in the cohorts fed 6 mg larva-1 at 15 °C. Here, the algal growth-mortality relationship directly pointed towards the importance of phycology in temperate mosquito ecology. Notably, Ae. albopictus ingests every available particle very quickly, especially green algal cells with an average of 22.4 ± 0.33 cells per second [59]. The green and blue-green algal growth in our study (Oocystis, Scenedesmus, Anabaena) were in fact negatively correlated to the initial Ae. albopictus larval ratio when 6 mg larva-1 food was supplied. Marten [22, 25] demonstrated a starvation effect of a variety of algal species, and in particular the lethal effect on first- and second-instars of Ae. albopictus if feeding on indigestible algal cells. For instance, larval feeding on Scenedesmus obliquus (the likely Scenedesmus species in our study) almost always killed Ae. albopictus first- and second-instar larvae, whereas Ae. albopictus first- and second-instar larvae survived feeding on some (but not every) species of Scenedesmus, Oocystis and Anabaena [25].

Water physicochemistry suits both mosquito species

Water physicochemistry in the test vessels well resembled that of Cx. pipiens (s.l.) microhabitats at the northern border of the Rhine rift valley in Germany. The mosquito larvae had a direct impact on nutrient cycling (e.g. nitrate, silicon dioxide concentration) and algal growth, at least in our test vessels. According to physicochemistry, the ecology of Cx. pipiens (s.l.) is described as euryoecious [26] (β-mesosaprob microhabitats, [60]). For Ae. albopictus, however, the physicochemical parameters in the replacement series experiment were sometimes higher than reported in the few published studies in their natural microhabitats (pH = 7.97, dissolved oxygen ≤ 9.7 mg l-1, [23, 61]). A pH between 6.8 and 7.6 seems optimal, although Ae. albopictus larvae were observed within a pH range of 5.2 to 8.4 and at dissolved oxygen levels of 1.3 mg l-1 [60]. Wu & Chang [62] reported the greatest rate of food displacement of Ae. albopictus in water with a pH of 5.5. However, given that physicochemical conditions similar to our experiment were interference-free for Ae. albopictus larval/pupal performance at 20 °C to 30 °C [38], the physicochemistry of natural Culex spp. microhabitats in warmer regions of Germany, specifically in the Rhine-Main Metropolitan region, would not prevent an establishment of Ae. albopictus.

Conclusions

Given recent distribution models [3, 4] and the fact that 67% of collected Ae. albopictus larvae shared their microhabitats with Cx. pipiens (s.l.) larvae in an Italian monitoring study [18], regular encounters between Ae. albopictus and Cx. pipiens (s.l.) larvae can be anticipated in suitable (container-like) temporary water bodies in Germany. Our results indicate that larval coexistence of Cx. pipiens molestus and Ae. albopictus is possible at present and forecasted future spring-to-autumn climate conditions of the Rhine rift valley, but competitive exclusion due to reduced larval survival of Ae. albopictus may be the more frequent condition in microhabitats where Cx. pipiens (s.l.) populations are established.

Next to the competitive superiority of Cx. pipiens (s.l.), the advantage of the faster development of Cx. pipiens (s.l.) at 15 °C may be of highest importance when available containers are initially colonized in spring and early summer, and to maintain population growth in autumn. However, the temperature was not a controlling factor for interspecific competition in our study, but food quantity, multifactorial interactions and food-related water parameter changes. Species-specific foraging behaviour influenced the physicochemistry and phycology in experimental mosquito microhabitats, and algal growth controlled especially the performance of Ae. albopictus. Thus, detailed examination of context dependence in interspecific interactions of these two important vector species and the related changes of water chemistry may lead to a better understanding of Ae. albopictus colonization in Germany [63] and elsewhere.

Abbreviations

Ae: Cx00:30

Experimental units with only Cx. pipiens molestus larvae

Ae: Cx10:20

Asymmetrically mixed treatment with a 10:20 distribution of Ae. albopictus: Cx. pipiens molestus larvae

Ae: Cx15:15

Symmetrically mixed treatment with a 15:15 distribution of Ae. albopictus: Cx. pipiens molestus larvae

Ae: Cx20:10

Asymmetrically mixed treatment with a 20:10 distribution of Ae. albopictus: Cx. pipiens molestus larvae

Ae: Cx30:00

Experimental units with only Ae. albopictus larvae

AL: 

Abdominal length (mm)

AL-RCC: 

AL-specific relative crowding coefficient

ANOVA: 

Analysis of variance

BA: 

Gender-specific daily biomass accumulation rate (mg larva-1 day-1)

BA-RCC: 

BA-specific relative crowding coefficient

BH: 

Benjamini-Hochberg

cox1: 

Mitochondrial cytochrome c oxidase subunit 1

CV: 

Coefficient of variation

DW: 

Dry weight (mg)

DW-RCC: 

DW-specific relative crowding coefficient

FDR: 

False discovery rate

HSD: 

Honest significant difference

PT50

Gender-specific averages of the mean pupation time (days)

PT50-RCC: 

PT50-specific relative crowding coefficient

RCC: 

Relative crowding coefficient

RCCAe

Integrated relative crowding coefficient for Ae. albopictus

RCCCx

Integrated relative crowding coefficient for Cx. pipiens molestus

Declarations

Acknowledgements

The authors wish to thank Jörg Oehlmann for his comprehensive support of the study. We are also thankful to two anonymous reviewers for their helpful comments.

Funding

The present study was partly funded by the research funding programme ´LOEWE - Landes-Offensive zur Entwicklung Wissenschaftlich-ökonomischer Exzellenz’ of the Ministry of Higher Education, Research, and the Arts of the State of Hesse, Germany. Financial support was furthermore provided by the Hessian Centre on Climate Change (FZK) of the Hessian Agency for Nature Conservation, Environment and Geology (HLNUG) with co-funding by the Rhineland-Palatinate Centre of Excellence for Climate Change Impacts.

Availability of data and materials

All datasets on which the conclusions of the manuscript rely are presented in the article and its additional files.

Authors’ contributions

All authors conceived and designed the study. SV, TK and RB conducted the experiments and RM, TK, SV, FR and RB analyzed the data. RM, DAG and UK provided laboratory, rearing and other infrastructure. All authors read and approved the final manuscript.

Ethics approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

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Authors’ Affiliations

(1)
Goethe University Frankfurt am Main, Institute for Occupational Medicine, Social Medicine and Environmental Medicine, Frankfurt am Main, Germany
(2)
Goethe University Frankfurt am Main, Institute of Ecology, Evolution and Diversity, Frankfurt am Main, Germany
(3)
Senckenberg Biodiversity and Climate Research Centre, Frankfurt am Main, Germany

References

  1. European Centre for Disease Prevention and Control. Aedes albopictus. available from: http://ecdc.europa.eu/en/healthtopics/vectors/mosquitoes/Pages/aedes-albopictus.aspx. Accessed 15 Apr 2016.
  2. Werner D, Kampen H. Aedes albopictus breeding in southern Germany, 2014. Parasitol Res. 2015;114(3):831–4.View ArticlePubMedGoogle Scholar
  3. Erguler K, Smith-Unna SE, Waldock J, Proestos Y, Christophides GK, Lelieveld J, Parham PE. Large-scale modelling of the environmentally-driven population dynamics of temperate Aedes albopictus (Skuse). PloS One. 2016;11(2):e0149282.View ArticlePubMedPubMed CentralGoogle Scholar
  4. Koch LK, Cunze S, Werblow A, Kochmann J, Dörge DD, Mehlhorn H, Klimpel S. Modeling the habitat suitability for the arbovirus vector Aedes albopictus (Diptera: Culicidae) in Germany. Parasitol Res. 2015;115(3):957-64.Google Scholar
  5. Teng HJ, Apperson CS. Development and survival of immature Aedes albopictus and Aedes triseriatus (Diptera: Culicidae) in the laboratory: effects of density, food, and competition on response to temperature. J Med Entomol. 2000;37(1):40–52.View ArticlePubMedGoogle Scholar
  6. Lounibos LP, Suárez S, Menéndez Z, Nishimura N, Escher RL, SM OC, Rey JR. Does temperature affect the outcome of larval competition between Aedes aegypti and Aedes albopictus? J Vector Ecol. 2002;27:86–95.PubMedGoogle Scholar
  7. Juliano SA, Lounibos PL. Ecology of invasive mosquitoes: effects on resident species and on human health. Ecol Lett. 2005;8(5):558–74.View ArticlePubMedPubMed CentralGoogle Scholar
  8. Rochlin I, Gaugler R, Williges E, Farajollahi A. The rise of the invasives and decline of the natives: insights revealed from adult populations of container-inhabiting Aedes mosquitoes (Diptera: Culicidae) in temperate North America. Biol Invasions. 2013;15(5):991–1003.View ArticleGoogle Scholar
  9. Juliano SA. Species interactions among larval mosquitoes: context dependence across habitat gradients. Annu Rev Entomol. 2009;54:37.View ArticlePubMedPubMed CentralGoogle Scholar
  10. Chaves LF. Globally invasive, withdrawing at home: Aedes albopictus and Aedes japonicus facing the rise of Aedes flavopictus. Int J Biometeorol. 2016;60(11):1727-38.Google Scholar
  11. Manni M, Guglielmino CR, Scolari F, Vega-Rúa A, Failloux AB, Somboon P, et al. Genetic evidence for a worldwide chaotic dispersion pattern of the arbovirus vector, Aedes albopictus. PLoS NTD. 2017;11(1):e0005332.Google Scholar
  12. Begon M, Townsend CR, Harper JL. Ecology: from individuals to ecosystems. 4th ed. New York: Wiley; 2011.Google Scholar
  13. Novak MG, Higley LG, Christianssen CA, Rowley WA. Evaluating larval competition between Aedes albopictus and Ae. triseriatus (Diptera: Culicidae) through replacement series experiments. Environ Entomol. 1993;22:311–8.View ArticleGoogle Scholar
  14. Barrera R. Competition and resistance to starvation in larvae of container-inhabiting Aedes mosquitoes. Ecol Entomol. 1996;21(2):117–27.View ArticleGoogle Scholar
  15. Juliano SA. Species introduction and replacement among mosquitoes: interspecific resource competition or apparent competition? Ecology. 1998;79(1):255–68.View ArticleGoogle Scholar
  16. Griswold MW, Lounibos LP. Does differential predation permit invasive and native mosquito larvae to coexist in Florida? Ecol Entomol. 2005;30(1):122–7.View ArticlePubMedPubMed CentralGoogle Scholar
  17. Murrell EG, Juliano SA. Detritus type alters the outcome of interspecific competition between Aedes aegypti and Aedes albopictus (Diptera: Culicidae). J Med Entomol. 2008;45(3):375–83.View ArticlePubMedPubMed CentralGoogle Scholar
  18. Carrieri M, Bacchi M, Bellini R, Maini S. On the competition occurring between Aedes albopictus and Culex pipiens (Diptera: Culicidae) in Italy. Environ Entomol. 2003;32(6):1313–21.View ArticleGoogle Scholar
  19. Buckner EA, Alto BW, Lounibos LP. Larval temperature-food effects on adult mosquito infection and vertical transmission of dengue-1 virus. J Med Entomol. 2015;53(1):91–8.View ArticlePubMedPubMed CentralGoogle Scholar
  20. Daugherty MP, Alto BW, Juliano SA. Invertebrate carcasses as a resource for competing Aedes albopictus and Aedes aegypti (Diptera: Culicidae). J Med Entomol. 2000;37(3):364–72.View ArticlePubMedPubMed CentralGoogle Scholar
  21. Livdahl TP, Willey MS. Prospects for an invasion: competition between Aedes albopictus and native Aedes triseriatus. Science. 1991;253(5016):189–91.View ArticlePubMedGoogle Scholar
  22. Marten GG. Impact of the copepod Mesocyclops leuckarti pilosa and the green alga Kirchneriella irregularis upon larval Aedes albopictus (Diptera: Culicidae). Bull Soc Vector Ecol. 1984;9(1):1–5.Google Scholar
  23. Akram W, Lee JJ. Effect of habitat characteristics on the distribution and behavior of Aedes albopictus. J Vector Ecol. 2004;29(2):379–82.PubMedGoogle Scholar
  24. Leisnham PT, Slaney DP, Lester PJ, Weinstein P, Heath ACG. Mosquito density, macroinvertebrate diversity, and water chemistry in water-filled containers: relationships to land use. New Zeal J Z. 2007;34(3):203–18.View ArticleGoogle Scholar
  25. Marten GG. Mosquito control by plankton management: the potential of indigestible green algae. J Trop Med Hyg. 1986;89:213–22.PubMedGoogle Scholar
  26. Farajollahi A, Fonseca DM, Kramer LD, Marm Kilpatrick A. “Bird biting” mosquitoes and human disease: a review of the role of Culex pipiens complex mosquitoes in epidemiology. Infect Genet Evol. 2011;11:1577–85.View ArticlePubMedPubMed CentralGoogle Scholar
  27. Costanzo KS, Mormann K, Juliano SA. Asymmetrical competition and patterns of abundance of Aedes albopictus. J Med Entomol. 2005;42(4):559–70.View ArticlePubMedPubMed CentralGoogle Scholar
  28. Marini G, Guzzetta G, Baldacchino F, Arnoldi D, Montarsi F, Capelli G, et al. The effect of interspecific competition on the temporal dynamics of Aedes albopictus and Culex pipiens. Parasit Vectors. 2017;10(1):102.View ArticlePubMedPubMed CentralGoogle Scholar
  29. Delatte H, Gimonneau G, Triboire A, Fontenille D. Influence of temperature on immature development, survival, longevity, fecundity, and gonotrophic cycles of Aedes albopictus, vector of chikungunya and dengue in the Indian Ocean. J Med Entomol. 2009;46(1):33–41.View ArticlePubMedGoogle Scholar
  30. Vinogradova EB, Ivshina EV, Shaikevich EV. A study of the mosquito Culex pipiens (Diptera, Culicidae) population structure in the Transcaucasia using molecular identification methods. Entomol Rev. 2013;93(1):14–8.View ArticleGoogle Scholar
  31. Sambrook J, Fritsch EF, Maniatis T. Molecular cloning: a laboratory manual. New York: CSHL Press; 1989.Google Scholar
  32. Vrijenhoek R. DNA primers for amplification of mitochondrial cytochrome c oxidase subunit I from diverse metazoan invertebrates. Mol Mar Biol Biotechnol. 1994;3(5):294–9.Google Scholar
  33. Kreß A, Kuch U, Oehlmann J, Müller R. Impact of temperature and nutrition on the toxicity of the insecticide λ-cyhalothrin in full-lifecycle tests with the target mosquito species Aedes albopictus and Culex pipiens. J Pest Sci. 2014;87(4):739–50.View ArticleGoogle Scholar
  34. Oberg AL, Young LJ, Higley LG. A comparison of two measures of competition. J Agric Biol Environ Stat. 1996;1:393–403.View ArticleGoogle Scholar
  35. Alto BW, Lounibos LP, Higgs S, Juliano SA. Larval competition differentially affects arbovirus infection in Aedes mosquitoes. Ecology. 2005;86(12):3279–88.View ArticlePubMedPubMed CentralGoogle Scholar
  36. Walsh RK, Bradley C, Apperson CS, Gould F. An experimental field study of delayed density dependence in natural populations of Aedes albopictus. PLoS One. 2012;7(4):e35959.View ArticlePubMedPubMed CentralGoogle Scholar
  37. Yee DA, Allgood D, Kneitel JM, Kuehn KA. Constitutive differences between natural and artificial container mosquito habitats: vector communities, resources, microorganisms, and habitat parameters. J Med Entomol. 2012;49(3):482–91.View ArticlePubMedGoogle Scholar
  38. Müller R, Knautz T, Völker J, Kreß A, Kuch U, Oehlmann J. Appropriate larval food quality and quantity for Aedes albopictus (Diptera: Culicidae). J Med Entomol. 2013;50(3):668–73.View ArticlePubMedGoogle Scholar
  39. DIN 38412 Teil 16. Bestimmung des Chlorophyll-a-Gehaltes von Oberflächenwasser (L42). Deutsche Einheitsverfahren zur Wasser-, Abwasser- und Schlammuntersuchung; Testverfahren mit Wasserorganismen (Gruppe L). 1985;16. Lieferung 1986.Google Scholar
  40. Linne von Berg KH, Hoef-Emden K, Melkonian M. Der Kosmos-Algenführer. Die wichtigsten Süßwasseralgen im Mikroskop. Stuttgart: Kosmos Gesellschaft der Naturfreunde, Franckh'sche Verlagshandlung; 2004.Google Scholar
  41. Streble H, Krauter D. Das Leben im Wassertropfen. Mikroflora und Fauna des Süßwassers. Ein Bestimmungsbuch. Stuttgart: Kosmos Gesellschaft der Naturfreunde, Franckh'sche Verlagshandlung; 1973.Google Scholar
  42. Harper JL. Population biology of plants. New York: Academic Press; 1977.Google Scholar
  43. Zarochentseva O. Adaptation of methodology calculation relative crowding coefficient for evaluation competition of tree species in polyculture. In: Economics for ecology ISCS'2012: 18th International Scientific Conference, Sumy, Аpril 27-30, 2012. Sumi: SumDU; 2012, p. 196-197.Google Scholar
  44. Noden BH, O'Neal PA, Fader JE, Juliano SA. Impact of inter- and intra-specific competition among larvae on larval, adult, and life-table traits of Aedes aegypti and Aedes albopictus females. Ecol Entomol. 2015;41:192–200.View ArticlePubMedPubMed CentralGoogle Scholar
  45. Eldridge BF. Environmental control of ovarian development in mosquitoes of the Culex pipiens complex. Science. 1966;151(3712):826–8.View ArticlePubMedGoogle Scholar
  46. Ruybal JE, Kramer LD, Kilpatrick AM. Geographic variation in the response of Culex pipiens life history traits to temperature. Parasit Vectors. 2016;9(1):1.View ArticleGoogle Scholar
  47. Black WC, Rai KS, Turco BJ, Arroyo DC. Laboratory study of competition between United States strains of Aedes albopictus and Aedes aegypti (Diptera: Culicidae). J Med Entomol. 1989;26(4):260–71.View ArticlePubMedGoogle Scholar
  48. Leisnham PT, Lounibos LP, O'Meara GF, Juliano SA. Interpopulation divergence in competitive interactions of the mosquito Aedes albopictus. Ecology. 2009;90(9):2405–12.View ArticlePubMedPubMed CentralGoogle Scholar
  49. Gomes B, Sousa CA, Vicente JL, Pinho L, Calderón I, Arez E, et al. Feeding patterns of molestus and pipiens forms of Culex pipiens (Diptera: Culicidae) in a region of high hybridization. Parasit Vectors. 2013;6:93.View ArticlePubMedPubMed CentralGoogle Scholar
  50. Asgharian H, Chang PL, Lysenkov S, Scobeyeva VA, Reisen WK, Nuzhdin SV. Evolutionary genomics of Culex pipiens: global and local adaptations associated with climate, life-history traits and anthropogenic factors. Proc R Soc Lond B Biol Sci. 2015;282(1810):20150728.View ArticleGoogle Scholar
  51. Weitzel T, Collado A, Jöst A, Pietsch K, Storch V, Becker N. Genetic differentiation of populations within the Culex pipiens complex and phylogeny of related species. J Am Mosq Control Assoc. 2009;25(1):6–17.View ArticlePubMedGoogle Scholar
  52. Becker N, Jöst A, Weitzel T. The Culex pipiens complex in Europe. J Am Mosq Control Assoc. 2012;28(4s):53–67.View ArticlePubMedGoogle Scholar
  53. Yee DA, Kesavaraju B, Juliano SA. Direct and indirect effects of animal detritus on growth, survival, and mass of invasive container mosquito Aedes albopictus (Diptera: Culicidae). J Med Entomol. 2007;44(4):580–8.View ArticlePubMedPubMed CentralGoogle Scholar
  54. Hawley WA. The biology of Aedes albopictus. J Am Mosq Control Assoc. 1988;Supplement 1:1–39.Google Scholar
  55. Yee DA, Juliano SA. Consequences of detritus type in an aquatic microsystem: effects on water quality, micro-organisms and performance of the dominant consumer. Freshw Biol. 2006;51(3):448–59.View ArticlePubMedPubMed CentralGoogle Scholar
  56. Winters AE, Yee DA. Variation in performance of two co-occurring mosquito species across diverse resource environments: insights from nutrient and stable isotope analyses. Ecol Entomol. 2012;37(1):56–64.View ArticleGoogle Scholar
  57. Dadd RH. Comparison of rates of ingestion of particulate solids by Culex pipiens larvae: Phagostimulation effect on water-soluble yeast extract. Entomol Exp Appl. 1970;13(4):407–19.View ArticleGoogle Scholar
  58. Rashed SS, Mulla MS. Factors influencing ingestion of particulate materials by mosquito larvae (Diptera: Culicidae). J Med Entomol. 1989;26(3):210–6.View ArticlePubMedGoogle Scholar
  59. Ho BC, Khoo HG, Chew LM, Wong KP. Ewert A. Food ingestion and digestive enzymes in larval Aedes aegypti and Ae. albopictus (Diptera: Culicidae). J Med Entomol. 1992;29(6):960–4.Google Scholar
  60. Estrada-Franco JG, Craig GB. Biology, disease relationship, and control of Aedes albopictus. Washington DC. Pan American Health Organisation; 1995. Technical Paper 42.Google Scholar
  61. Dom NC, Madzlan MF, Hasnan SNA, Misran N. Water quality characteristics of dengue vectors breeding containers. Int J Mosq Res. 2016;3(1):25–9.Google Scholar
  62. Wu HH, Chang NT. Influence of temperature, water quality and pH value on ingestion and development of Aedes aegypti and Aedes albopictus (Diptera: Culicidae) larvae. Chin J Entomol. 1993;13:33–44.Google Scholar
  63. Becker N, Schön S, Klein A-M, Ferstl I, Kizgin A, Tannich E, Kuhn C, Pluskota B, Jöst A. First mass development of Aedes albopictus (Diptera: Culicidae) - its surveillance and control in Germany. Parasitol Res. 2017; https://doi.org/10.1007/s00436-016-5356-z.

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