Generalized spectral light scatter models of diverse bacterial colony morphologies

An optical forward‐scatter model was generalized to encompass the diverse nature of bacterial colony morphologies and the spectral information. According to the model, the colony shape and the wavelength of incident light significantly affect the characteristics of a forward elastic‐light‐scattering pattern. To study the relationship between the colony morphology and the scattering pattern, three‐dimensional colony models were generated in various morphologies. The propagation of light passing through the colony model was then simulated. In validation of the theoretical modeling, the scattering patterns of three bacterial genera, Staphylococcus, Exiguobacterium and Bacillus, which grow into colonies having convex, crateriform and flat elevations, respectively, were qualitatively compared to the simulated scattering patterns. The strong correlations observed between simulated and experimental patterns validated the scatter model. In addition, spectral effect on the scattering pattern was studied using the scatter model, and experimentally investigated using Staphylococcus, whose colony has circular form and convex elevation. Both simulation and experiment showed that changes in wavelength affected the overall pattern size and the number of rings.     

Streptomycin Induced Stress Response in Salmonella enterica Serovar Typhimurium Shows Distinct Colony Scatter Signature

We investigated the streptomycin-induced stress response in Salmonella enterica serovars with a laser optical sensor, BARDOT (bacterial rapid detection using optical scattering technology). Initially, the top 20 S. enterica serovars were screened for their response to streptomycin at 100 μg/mL. All, but four S. enterica serovars were resistant to streptomycin. The MIC of streptomycin-sensitive serovars (Enteritidis, Muenchen, Mississippi, and Schwarzengrund) varied from 12.5 to 50 μg/mL, while streptomycin-resistant serovar (Typhimurium) from 125–250 μg/mL. Two streptomycin-sensitive serovars (Enteritidis and Mississippi) were grown on brain heart infusion (BHI) agar plates containing sub-inhibitory concentration of streptomycin (1.25–5 μg/mL) and a streptomycin-resistant serovar (Typhimurium) was grown on BHI containing 25–50 μg/mL of streptomycin and the colonies (1.2 ± 0.1 mm diameter) were scanned using BARDOT. Data show substantial qualitative and quantitative differences in the colony scatter patterns of Salmonella grown in the presence of streptomycin than the colonies grown in absence of antibiotic. Mass-spectrometry identified overexpression of chaperonin GroEL, which possibly contributed to the observed differences in the colony scatter patterns. Quantitative RT-PCR and immunoassay confirmed streptomycin-induced GroEL expression while, aminoglycoside adenylyltransferase (aadA), aminoglycoside efflux pump (aep), multidrug resistance subunit acrA, and ribosomal protein S12 (rpsL), involved in streptomycin resistance, were unaltered. The study highlights suitability of the BARDOT as a non-invasive, label-free tool for investigating stress response in Salmonella in conjunction with the molecular and immunoassay methods.     

A review of Gammaridae (Crustacea: Amphipoda): the family extent,its evolutionary history,and taxonomic redefinition of genera

By molecular analysis of a high number of gammarids, including 29 out‐group genera, we could assure the monophyly of Gammaridae. To avoid the paraphyly of the family, we propose the omission of Pontogammaridae, Typhlogammaridae, and all Baikalian families. Similarly, the genera Fontogammarus, Sinogammarus, Lagunogammarus, Pephredo, Neogammarus, and Laurogammarus may be cancelled. But, tens of Baikal genera, nested within Gammarus, are so diverse that they must be retained, although rendering Gammarus paraphyletic. Besides we propose the polyphyletic Echinogammarus–Chaetogammarus group to be divided into monophyletic genera Echinogammarus s. str., Homoeogammarus, Parhomoeogammarus, Marinogammarus, R elictogammarus gen. nov. , Chaetogammarus, and T richogammarus gen. nov. These solutions made it possible to complete the first analysis of the family evolution in light of its phylogeny. Perimarine clades are mainly basally split clades, whereas in some ancient lakes extremely rich endemic faunas had developed polyphyletically. The troglobiotic Typhlogammarus group from Dinarides and Caucasus formed a monophylum, whereas the troglobiotic assemblage of Gammarus species is highly polyphyletic. Reduction of the uropod III endopodite, which classically distinguishes between the genera Gammarus and Echinogammarus, appeared to be highly polyphyletic. Protective dorsal pleonal projections occur scattered across the family and beyond, whereas lateral projections were limited to species of ancient lakes, so both structures were polyphyletic. The evolutionary history of Gammaridae was investigated with ten different calibration schemes, which produced incompatible results; however, the most probable scenario is a late rise of the family, which can only explain the absence of Gammaridae species around the Indo‐Pacific. © 2015 The Linnean Society of London     

Isolation and Identification of Bacteria Associated with Adult Laboratory Mexican Fruit Flies, Anastrepha ludens (Diptera: Tephritidae)

From the guts of new and old colonies (female and male) of Mexican fruit flies, Anastrepha ludens (Diptera: Tephritidae), we identified a total of 18 different bacterial species belonging to the family Enterobacteriaceae, Pseudomonadaceae, Vibrionaceae, Micrococcaceae, Deinococcacea, Bacillaceae, and the genus Listeria. Enterobacter, Providencia, Serratia, and Staphylococcus spp. were the most frequently isolated genera, with Citrobacter, Streptococcus, Aerococcus, and Listeria found less frequently. We found Bacillus cereus, Enterobacter sakazakii, Providencia stuartii, and Pseudomonas aeruginosa only in the new colony, Aeromonas hydrophila and Klebsiella pneumoniae spp. pneumoniae only in the old colony. We also studied resistance/sensitivity to 12 antibiotics for six bacterial isolates such as Enterobacter cloacae, E. sakazakii, K. pneumoniae spp., Providencia rettgeri, P. aeruginosa, and Bacillus cereus. Isolates on the whole were resistant to penicillin and ampicillin (five of six isolates) and sensitive to rifampin and streptomycin (six of six isolates). Antibiotic resistance profiles might be useful characteristics for distinguishing among species and strains of these bacteria, probably having ecological significance with respect to intra- and inter-specific competition within host cadavers, and could have implications for the utility of these organisms for biological control, including the alternative control strategy, paratransgenesis. Received: 28 August 2000 / Accepted: 2 October 2000     

Detection by using monoclonal antibodies of Yersinia enterocolitica in artificially-contaminated pork

Monoclonal antibodies against Yersinia enterocolitica were produced by fusion of NS‐1 mouse myeloma cells with spleen cells of ICR mice immunized with heat‐killed and heat‐killed plus SDS‐mercaptoethanol treated forms of Y. enterocolitica ATCC 27729 alone or mixed with Y. enterocolitica MU. The twenty‐five MAbs obtained from five fusions were divided into nine groups according to their specificities to different bacterial strains and species, as determined by dot blotting. The first five groups of MAbs were specific only to Y. enterocolitica, but did not recognize all of the isolates tested. MAbs in groups 6 and 7 reacted with all isolates of Y. enterocolitica tested but showed cross‐reaction with some Yersinia spp. and Edwardsiella tarda, especially in the case of group 7. MAbs in groups 8 and 9 reacted with all isolates of Y. enterocolitica and Yersinia spp., as well as other Gram‐negative bacteria that belong to the family Enterobacteriaceae. These MAbs recognized Y. enterocolitica antigens with apparent molecular weights ranging from 10 – 43 kDa by Western blotting, and could detect Y. enterocolitica from ～10³– 10⁵ colony forming units (CFUs) by dot blotting. The hybridoma clone YE38 was selected for detection of Y. enterocolitica in pork samples which had been artificially‐contaminated by inoculation with Y. enterocolitica ATCC 27729 at concentrations of ～10⁴– 10⁶ CFUs/g and incubation in peptone sorbitol bile broth at 4°C. Samples were collected and applied on a nitrocellulose membrane for dot blotting with trypticase soy and cefsulodin‐Irgasan‐novobiocin agars. After 48 hr of incubation, the detection limit was ～10²– 10³ CFU/g by dot blotting.     

Light Scattering Sensor for Direct Identification of Colonies of Escherichia coli Serogroups O26, O45, O103, O111, O121, O145 and O157

Background

Shiga-toxin producing Escherichia coli (STEC) have emerged as important foodborne pathogens, among which seven serogroups (O26, O45, O103, O111, O121, O145, O157) are most frequently implicated in human infection. The aim was to determine if a light scattering sensor can be used to rapidly identify the colonies of STEC serogroups on selective agar plates.

Methodology/Principal Findings

Initially, a total of 37 STEC strains representing seven serovars were grown on four different selective agar media, including sorbitol MacConkey (SMAC), Rainbow Agar O157, BBL CHROMagarO157, and R&F E. coli O157:H7, as well as nonselective Brain Heart Infusion agar. The colonies were scanned by an automated light scattering sensor, known as BARDOT (BActerial Rapid Detection using Optical scattering Technology), to acquire scatter patterns of STEC serogroups, and the scatter patterns were analyzed using an image classifier. Among all of the selective media tested, both SMAC and Rainbow provided the best differentiation results allowing multi-class classification of all serovars with an average accuracy of more than 90% after 10–12 h of growth, even though the colony appearance was indistinguishable at that early stage of growth. SMAC was chosen for exhaustive scatter image library development, and 36 additional strains of O157:H7 and 11 non-O157 serovars were examined, with each serogroup producing unique differential scatter patterns. Colony scatter images were also tested with samples derived from pure and mixed cultures, as well as experimentally inoculated food samples. BARDOT accurately detected O157 and O26 serovars from a mixed culture and also from inoculated lettuce and ground beef (10-h broth enrichment +12-h on-plate incubation) in the presence of natural background microbiota in less than 24 h.

Conclusions

BARDOT could potentially be used as a screening tool during isolation of the most important STEC serovars on selective agar plates from food samples in less than 24 h.     

Composition and variability of airborne fungi in an enclosed rabbit house in China

Sampling was conducted from June 2007 to May 2008 in an enclosed rabbit house to investigate composition and variability of airborne fungi. Samples were collected using an Andersen-6 sampler, with Sabouraud culture medium as sampling medium. The results showed that monthly mean concentration was 2.79–5.46 × 10³ colony forming unit/m³ air (CFU/m³ air), with the maximum level in October, and the minimum level in January. Within a day, the maximum level occurred at 09:00, followed by 17:00 and then 13:00. A total of 6,523 fungal colonies, belonging to 17 genera and 36 species, were obtained. The predominant genera included Cladosporium, Penicillium, Aspergillus and Altemaria, comprising 71.45% of the colony count. The obtained fungi of the year were mainly centralized in the stage D of the sampler (2.0–3.0 μm), accounting for 37.8% of the colonies. The minimum value occurred at stage F (<0.65 μm), accounting for 1.10% of the colonies.     

Behavioural and molecular evidence for selective immigration and group regulation in the social huntsman spider,Delena cancerides

Movement among social groups interacts with the costs and benefits of group‐living in complex ways. Unlike most other social spiders, the social huntsman spider, Delena cancerides, appears to enter foreign colonies, discriminates kin from non‐kin, and has very limited dispersal options because their bark retreats are rare, making this species an interesting model organism with which to examine the role of inter‐colony movement on group‐living. We examined movement among field colonies of D. cancerides in three ways: (1) by tracking the dispersal and immigration of marked spiders into foreign colonies; (2) by recording resident spiders' behaviour toward introduced immigrants; and (3) by inferring intra‐colony relatedness and immigration patterns through allozyme electrophoresis. Of the marked spiders, only young juveniles moved into neighbouring colonies, whereas subadults and adults did not. Introduced juveniles were tolerated in foreign colonies, whereas introduced adult males and subadults were usually attacked by the resident adult female, unless she had similar sized subadult/adult offspring of her own. Allozyme profiles from unmanipulated field colonies showed that 47% of sampled colonies contained at least one immigrant and that average within colony relatedness was below 0.5. These data align with previous research on the costs and benefits of group‐living for D. cancerides, suggesting that spiders actively seek and regulate group membership based on interests of both the immigrant and the colony. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, ?? , ??–??.     

Notes on the almost unknown genus Jeotgalicoccus

Aims: To facilitate isolation and differentiation of the almost entirely unknown Jeotgalicoccus spp. Methods and Results: Jeotgalicoccus spp. have been found in dust samples using SSCP‐PCR analysis. As the cultivation of strains is necessary for further studies on virulence, pathogenicity or metabolism, we developed a method for cultural isolation and further differentiation of Jeotgalicoccus spp. We found that J. halotolerans, J. psychrophilus, J. marinus, as well as the related species Salinicoccus roseus grow on Baird Parker (BP) agar as black colonies without clear zones. J. pinnipedialis and S. jeotgali grow only weakly on BP agar without forming clearly delineated colonies. On BP agar, the colony‐forming Jeotgalicoccus and Salinicoccus spp. are not distinguishable from coagulase‐negative Staphylococcus spp. (CNS). However, unlike CNS, all of the above mentioned species are unreactive in the OF test. SSCP‐PCR was able to differentiate between all investigated Jeotgalicoccus and Salinicoccus spp., as all species had different band positions. Conclusions: Jeotgalicoccus spp. and Salinicoccus spp. may be widely distributed in the environment, but, until now, overlooked or confused with staphylococci. Further epidemiological studies, which are required to prove this hypothesis, are facilitated by the observations of our study. Significance and Impact: This not yet published information enables researchers to carry out epidemiological studies on Jeotgalicoccus spp. in a very cheap and easy way.     

Effects of disturbance on coral communities: bleaching in Moorea,French Polynesia

This study examines patterns of susceptibility and short-term recovery of corals from bleaching. A mass coral bleaching event began in March, 1991 on reefs in Moorea, French Polynesia and affected corals on the shallow barrier reef and to >20 m depth on the outer forereef slope. There were significant differences in the effect of the bleaching among common coral genera, with Acropora, Montastrea, Montipora, and Pocillopora more affected than Porites, Pavona, leptastrea or Millepora. Individual colonies of the common species of Acropora and Pocillopora were marked and their fate assessed on a subsequent survey in August, 1991 to determine rates of recovery and mortality. Ninety-six percent of Acropora spp. showed some degree of bleaching compared to 76% of Pocillopora spp. From March to August mortality of bleached colonies of Pocillopora was 17%, 38% recovered completely, and many suffered some partial mortality of the tissue. In contrast, 63% of the Acropora spp. died, and about 10% recovered completely. Generally, those colonies with less than 50% of the colony area affected by the bleaching recovered at a higher rate than did those with more severe bleaching. Changes in community composition four months after the event began included a significant decrease only in crustose algae and an increase in cover of filamentous algae, much of which occupied plate-like and branching corals that had died in the bleaching event. Total coral cover and cover of susceptible coral genera had declined, but not significantly, after the event.     

Taxonomic classification of the reef coral family Lobophylliidae (Cnidaria: Anthozoa: Scleractinia)

Lobophylliidae is a family‐level clade of corals within the ‘robust’ lineage of Scleractinia. It comprises species traditionally classified as Indo‐Pacific ‘mussids’, ‘faviids’, and ‘pectiniids’. Following detailed revisions of the closely related families Merulinidae, Mussidae, Montastraeidae, and Diploastraeidae, this monograph focuses on the taxonomy of Lobophylliidae. Specifically, we studied 44 of a total of 54 living lobophylliid species from all 11 genera based on an integrative analysis of colony, corallite, and subcorallite morphology with molecular sequence data. By examining coral skeletal features at three distinct levels – macromorphology, micromorphology, and microstructure – we built a morphological matrix comprising 46 characters. Data were analysed via maximum parsimony and transformed onto a robust molecular phylogeny inferred using two nuclear (histone H3 and internal transcribed spacers) and one mitochondrial (cytochrome c oxidase subunit I) DNA loci. The results suggest that micromorphological characters exhibit the lowest level of homoplasy within Lobophylliidae. Molecular and morphological trees show that Symphyllia, Parascolymia, and Australomussa should be considered junior synonyms of Lobophyllia, whereas Lobophyllia pachysepta needs to be transferred to Acanthastrea. Our analyses also lend strong support to recent revisions of Acanthastrea, which has been reorganized into five separate genera (Lobophyllia, Acanthastrea, Homophyllia, Sclerophyllia, and Micromussa), and to the establishment of Australophyllia. Cynarina and the monotypic Moseleya remain unchanged, and there are insufficient data to redefine Oxypora, Echinophyllia, and Echinomorpha. Finally, all lobophylliid genera are diagnosed under the phylogenetic classification system proposed here, which will facilitate the placement of extinct taxa on the scleractinian tree of life.     

Colony expansion model for describing the spatial distribution of populations

Two equations have been used frequently to describe the relation between the sample variance (s ²) and sample mean (m) of the number of individuals per quadrat: Taylor's power law, s ² = am ^b , and Iwao's m ^*−m regression, s ² = cm + dm ², where a, b, c, and d are constants. We can obtain biological information such as colony size and the degree of aggregation of colonies from parameters c and d of Iwao's m ^*−m regression. However, we cannot obtain such biological information from parameters a and b of Taylor's power law because these parameters have not been described by simple functions. To mitigate such in-convenience, I propose a mechanistic model that produces Taylor's power law; this model is called the colony expansion model. This model has the following two assumptions: (1) a population consists of a fixed number of colonies that lie across several quadrats, and (2) the number of individuals per unit occupied area of colony becomes v times larger in an allometric manner when the occupied area of colony becomes h times larger (v≥ 1, h≥ 1). The parameter h indicates the dispersal rate of organisms. We then obtain Taylor's power law with b = {ln[E(h)] + ln[E(v ²)]}/{ln[E(h)] + ln[E(v)]}, where E indicates the expectation. We can use the inverse of the exponent, 1/b, as an index of dispersal of individuals because it increases with increasing E(h). This model also yields a relation, known as the Kono–Sugino relation, between the proportion of occupied quadrats and the mean density per quadrat: −ln(1 −p) = fm ^g , where p is the proportion of occupied quadrats, f is a constant, and g = ln[E(h)]/{ln[E(h)] + ln[E(v)]}. We can use g as an index of dispersal as it increases with increasing E(h). The problem at low densities where Taylor's power law is not applicable is also discussed. Received: January 27, 2000 / Accepted: June 20, 2000     

Mating behavior,spawning, parental care,and embryonic development of some marine pseudocerotid flatworms (Platyhelminthes: Rhabditophora: Polycladida) in Singapore

The reproductive biology of tropical marine cotylean polyclads is presently poorly known. Reproductive aspects of 16 polyclad species from the family Pseudocerotidae in the genera Acanthozoon, Nymphozoon, Phrikoceros, Pseudobiceros, Pseudoceros, Thysanozoon, and Tytthosoceros from Singapore were documented for the first time. Penis fencing was observed to be just a mating ritual and not necessary for insemination, not always aggressive, and could also result in eventual reciprocal insemination. Results showed that all species underwent similar embryonic developmental stages and hatched as Müller's larvae. Only Pseudoceros concinnus and Pseudoceros laingensis, with mean developmental times of >20 days and mean brood sizes of <1,000 eggs, displayed long‐term parental care. Polyclads producing larger brood sizes had shorter developmental times and only covered their egg masses for about one day. Phrikoceros baibaiye and Pseudobiceros spp. produced egg capsules with pointed opercula, whereas all other species possessed smooth, rounded opercula. All genera hatched with brownish‐orange larvae, except for Pseudoceros spp., which hatched with reddish‐purple larvae regardless of the initial embryo color (either grayish‐yellow or reddish‐purple). These could potentially complement current taxonomic characters in distinguishing polyclad genera and species.     

Colony formation in the cyanobacterium Microcystis

Morphological evolution from a unicellular to multicellular state provides greater opportunities for organisms to attain larger and more complex living forms. As the most common freshwater cyanobacterial genus, Microcystis is a unicellular microorganism, with high phenotypic plasticity, which forms colonies and blooms in lakes and reservoirs worldwide. We conducted a systematic review of field studies from the 1990s to 2017 where Microcystis was dominant. Microcystis was detected as the dominant genus in waterbodies from temperate to subtropical and tropical zones. Unicellular Microcystis spp. can be induced to form colonies by adjusting biotic and abiotic factors in laboratory. Colony formation by cell division has been induced by zooplankton filtrate, high Pb²⁺ concentration, the presence of another cyanobacterium (Cylindrospermopsis raciborskii), heterotrophic bacteria, and by low temperature and light intensity. Colony formation by cell adhesion can be induced by zooplankton grazing, high Ca²⁺ concentration, and microcystins. We hypothesise that single cells of all Microcystis morphospecies initially form colonies with a similar morphology to those found in the early spring. These colonies gradually change their morphology to that of M. ichthyoblabe, M. wesenbergii and M. aeruginosa with changing environmental conditions. Colony formation provides Microcystis with many ecological advantages, including adaption to varying light, sustained growth under poor nutrient supply, protection from chemical stressors and protection from grazing. These benefits represent passive tactics responding to environmental stress. Microcystis colonies form at the cost of decreased specific growth rates compared with a unicellular habit. Large colony size allows Microcystis to attain rapid floating velocities (maximum recorded for a single colony, ∼ 10.08 m h⁻¹) that enable them to develop and maintain a large biomass near the surface of eutrophic lakes, where they may shade and inhibit the growth of less‐buoyant species in deeper layers. Over time, accompanying species may fail to maintain viable populations, allowing Microcystis to dominate. Microcystis blooms can be controlled by artificial mixing. Microcystis colonies and non‐buoyant phytoplankton will be exposed to identical light conditions if they are evenly distributed over the water column. In that case, green algae and diatoms, which generally have a higher growth rate than Microcystis, will be more successful. Under such mixing conditions, other phytoplankton taxa could recover and the dominance of Microcystis would be reduced. This review advances our understanding of the factors and mechanisms affecting Microcystis colony formation and size in the field and laboratory through synthesis of current knowledge. The main transition pathways of morphological changes in Microcystis provide an example of the phenotypic plasticity of organisms during morphological evolution from a unicellular to multicellular state. We emphasise that the mechanisms and factors influencing competition among various close morphospecies are sometimes paradoxical because these morphospecies are potentially a single species. Further work is required to clarify the colony‐forming process in different Microcystis morphospecies and the seasonal variation in this process. This will allow researchers to grow laboratory cultures that more closely reflect field morphologies and to optimise artificial mixing to manage blooms more effectively.     

Culturable Rhodobacter and Shewanella species are abundant in estuarine turbidity maxima of the Columbia River

Measurements of dissolved, ascorbate‐reducible and total Mn by ICP‐OES revealed significantly higher concentrations during estuarine turbidity maxima (ETM) events, compared with non‐events in the Columbia River. Most probable number (MPN) counts of Mn‐oxidizing or Mn‐reducing heterotrophs were not statistically different from that of other heterotrophs (10³–10⁴ cells ml^?1) when grown in defined media, but counts of Mn oxidizers were significantly lower in nutrient‐rich medium (13 cells ml^?1). MPN counts of Mn oxidizers were also significantly lower on Mn(III)‐pyrophosphate and glycerol (21 cells ml^?1). Large numbers of Rhodobacter spp. were cultured from dilutions of 10^?2 to 10^?5, and many of these were capable of Mn(III) oxidation. Up to c. 30% of the colonies tested LBB positive, and all 77 of the successfully sequenced LBB positive colonies (of varying morphology) yielded sequences related to Rhodobacter spp. qPCR indicated that a cluster of Rhodobacter isolates and closely related strains (95–99% identity) represented approximately 1–3% of the total Bacteria, consistent with clone library results. Copy numbers of SSU rRNA genes for either Rhodobacter spp. or Bacteria were four to eightfold greater during ETM events compared with non‐events. Strains of a Shewanella sp. were retrieved from the highest dilutions (10^?5) of Mn reducers, and were also capable of Mn oxidation. The SSU rRNA gene sequences from these strains shared a high identity score (98%) with sequences obtained in clone libraries. Our results support previous findings that ETMs are zones with high microbial activity. Results indicated that Shewanella and Rhodobacter species were present in environmentally relevant concentrations, and further demonstrated that a large proportion of culturable bacteria, including Shewanella and Rhodobacter spp., were capable of Mn cycling in vitro.     

Biodiversity of Salix spp. honeydew and nectar honeys determined by RP-HPLC and evaluation of their antioxidant capacity

Rare unifloral willow (Salix spp.) honeys obtained from nectar or honeydew were investigated by direct RP‐HPLC‐DAD method in order to identify and quantify compounds that can be used as possible markers of their origin. Antioxidant and antiradical activities of willow honeys were evaluated using FRAP (=ferric reducing antioxidant assay) and DPPH (=1,1‐diphenyl‐2‐picrylhydrazyl radical) tests, respectively. Also HMF (=5‐(hydroxymethyl)furfural), diastase activity, and CIE L*a*b*C*h* chromatic coordinates were evaluated. Abscisic acids (ABA) are typical of willow nectar honey, with a predominance of (Z,E)‐ABA on (E,E)‐ABA (98.2 and 31.7 mg/kg, resp.). Kinurenic acid and salicylic acid are useful to mark willow honeydew honey. The proposed HPLC‐DAD method proved to be easy and reliable to identify the two different Salix spp. honeys, being not affected from any sample preparation artifact. Total antioxidant activity measured with the FRAP assay ranged from 3.2 to 12.6 mmol Fe²⁺/kg, and the antiradical activity measured with the DPPH assay ranged from 0.6 to 3.0 mmol TEAC (=Trolox equivalent antioxidant capacity)/kg in nectar and honeydew honeys, respectively. Salix spp. nectar and honeydew honeys proved to be two completely different honeys, because, besides color attributes, they show different antioxidant properties and specific compounds.     

The structured diversity of specialized gut symbionts of the New World army ants

Symbiotic bacteria play important roles in the biology of their arthropod hosts. Yet the microbiota of many diverse and influential groups remain understudied, resulting in a paucity of information on the fidelities and histories of these associations. Motivated by prior findings from a smaller scale, 16S rRNA‐based study, we conducted a broad phylogenetic and geographic survey of microbial communities in the ecologically dominant New World army ants (Formicidae: Dorylinae). Amplicon sequencing of the 16S rRNA gene across 28 species spanning the five New World genera showed that the microbial communities of army ants consist of very few common and abundant bacterial species. The two most abundant microbes, referred to as Unclassified Firmicutes and Unclassified Entomoplasmatales, appear to be specialized army ant associates that dominate microbial communities in the gut lumen of three host genera, Eciton, Labidus and Nomamyrmex. Both are present in other army ant genera, including those from the Old World, suggesting that army ant symbioses date back to the Cretaceous. Extensive sequencing of bacterial protein‐coding genes revealed multiple strains of these symbionts coexisting within colonies, but seldom within the same individual ant. Bacterial strains formed multiple host species‐specific lineages on phylogenies, which often grouped strains from distant geographic locations. These patterns deviate from those seen in other social insects and raise intriguing questions about the influence of army ant colony swarm‐founding and within‐colony genetic diversity on strain coexistence, and the effects of hosting a diverse suite of symbiont strains on colony ecology.     

Sexual behavior and male volatile compounds in wild and mass‐reared strains of the Mexican fruit fly Anastrepha ludens (Diptera: Tephritidae) held under different colony management regimes

We compared the calling and mating behavior and volatile release of wild males Anastrepha ludens (Loew) with males from 4 mass‐reared strains: (i) a standard mass‐reared colony (control), (ii) a genetic sexing strain (Tap‐7), (iii) a colony started from males selected on their survival and mating competitiveness abilities (selected), and (iv) a hybrid colony started by crossing wild males with control females. Selected and wild males were more competitive, achieving more matings under field cage conditions. Mass‐reared strains showed higher percentages of pheromone calling males under field conditions except for Tap‐7 males, which showed the highest percentages of pheromone calling males under laboratory cage conditions. For mature males of all strains, field‐cage calling behavior increased during the last hour before sunset, with almost a 2 fold increase exhibited by wild males during the last half hour. The highest peak mating activity of the 4 mass‐reared strains occurred 30 min earlier than for wild males. By means of solid phase microextraction (SPME) plus gas chromatography‐mass spectrometry (GC‐MS), the composition of volatiles released by males was analyzed and quantified. Wild males emitted significantly less amounts of (E,E)‐α‐farnesene but emitted significantly more amounts of (E,E)‐suspensolide as they aged than mass‐reared males. Within the 4 mass‐reared strains, Tap‐7 released significantly more amounts of (E,E)‐α‐farnesene and hybrid more of (E,E)‐suspensolide. Differences in chemical composition could be explained by the intrinsic characteristics of the strains and the colony management regimes. Characterization of calling behavior and age changes of volatile composition between wild and mass‐reared strains could explain the differences in mating competitiveness and may be useful for optimizing the sterile insect technique in A. ludens.