Round-cell mutants of Salmonella typhimurium produced by transposition mutagenesis: Lethality of rodA and mre mutations

Summary Thirty-three insertions of transposon Tn10l6l7 into genes involved in the control of rod cell shape were isolated in Salmonella typhimurium by the characteristic glossy appearance of colonies composed of spherical cells. Genetic tests demonstrated that 25 (76%) were insertions in the rodA gene, 7 (21 %) were mre mutants, and 1 (3%) was a divD mutant. No insertion in the pbpA gene were found. Insertions in cell shape genes only appeared when strains displaying resistance to mecillinam (not caused by -lactamase production) were employed. Neither rodA nor mre insertions could be transduced to wild-type strains but they were normally accepted by mecillinam-resistant derivatives and by cya and crp mutants, which, unlike the corresponding Escherichia coli strains, did not display resistance to mecillinam. On the other hand, the divD insertion could be efficiently transduced to any strain. It is concluded that the rodA, mre, and divD genes are involved in the control of rod cell shape but, in addition, the RodA and Mre products perform some function(s) that is essential for wild-type cells but dispensable for some mecillinam-resistant strains, and for cya and crp mutants.     

Abscisic acid and mefluidide in the regulation of cold hardiness development in Solanum tuberosum L. potato

Plants of Solanum tuberosum L. potato do not cold acclimate when exposed to low temperature such as 5°C, day/night. When ABA (45 M) was added to the culture medium, stem-cultured plantlets of S. tuberosum, cv. Red Pontiac, either grown at 20°C/15°C, day/night, or at 5°C, increased in cold hardiness from –2°C (killing temperature) to –4.5°C. The increase in cold hardiness could be inhibited in both temperature regimes if cycloheximide (70 M) was added to the culture medium at the inception of ABA treatment. Cycloheximide did not inhibit cold hardiness development, however, when it was added to the culture medium 3 days after ABA treatment.When pot-grown plants were foliar sprayed with mefluidide (50 M), ABA content increased from 10 nmol to 30 nmol g^–1 dry weight and plants increased in cold hardiness from –2°C to about –3.5°C. The increases in free ABA and cold hardiness occurred only in plants grown at 20°C/15°C; neither ABA nor cold hardiness increased in plants grown at 5°C.The results suggest that an increase in ABA and a subsequent de novo synthesis of proteins are required for the development of cold hardiness in S. tuberosum regardless of temperature regime, and that the inability to synthesize ABA at low temperature, rather than protein synthesis, appears to be the reason why S. tuberosum does not cold acclimate.     

A scanning electron microscope study of normal and vitrified leaves from Datura insignis plantlets cultured in vitro

The surface anatomy of normal and vitreous leaves of plantlets obtained from Datura insignis Barb Rodr nodal segment cultures was compared using scanning electron microscopy. Normal and vitrified leaves are similar in several ways. They are both amphistomatic, and have similar distributions of glandular and non-glandular trichomes. Stomata have similar length, diameter and distribution in normal and vitreous plants. Immature stomata, which have closed pores, and plugged stomata, which contain an amorphous material between their guard cells, occur in both normal and vitrified leaves. Normal and vitreous leaves differ in the frequency of normal and abnormal stomata. Normal stomata have kidney-shaped guard cells and resemble closely those found in field-grown plants, whereas abnormal stomata have deformed guard cells. Normal stomata represent approximately 80% of the total number of stomata in normal leaves, but only 7% of the total number of stomata in vitreous leaves. Abnormal stomata represent 90% of the total number in vitreous leaves. The deformation of guard cells could possibly be a mechanical impediment to stomatal function.     

Stimulation of Brain Pregnenolone Synthesis by Mitochondrial Diazepam Binding Inhibitor Receptor Ligands In Vivo

Abstract: Evidence that neurosteroids are potent modulators of the action of GABA at GABA_A receptors has prompted the investigation of the mechanism that controls brain neurosteroid synthesis by glial cell mitochondria in vivo. In vitro studies suggest that the interaction of the diazepam binding inhibitor (DBI)—a polypeptide that is abundant in steroidogenic cells—with glial mitochondrial DBI receptors (MDRs) is a crucial step in the physiological regulation of neurosteroid biosynthesis. MDRs bind 4-chlorodiazepam (4′-CD), N,N-di-n-hexyl-2-(4-fluorophenyl)-indol-3-acetamide (FGIN-1–27), and the isoquinoline carboxamide PK 11195 with high affinity, and these ligands have been used to investigate whether the stimulation of glial MDRs increases brain pregnenolone production in vivo. Adrenalectomized and castrated (A-C) male rats (to eliminate peripheral sources of pregnenolone) were pretreated with trilostane (to prevent pregnenolone metabolism to progesterone), and the pregnenolone content in brain regions dissected after fixation with a 0.8-s exposure to microwave irradiation focused to the head was determined by HPLC followed by specific radioimmunoassay. The forebrain and cerebellum of A-C rats contained 4–7 ng of pregnenolone/g of tissue, and the olfactory bulb contained 10–14 ng/g. These concentrations of brain pregnenolone are only 30–40% lower than those of shamoperated rats. In contrast, the plasma pregnenolone content of sham-operated rats was 2–3 ng/ml, but it was only 0.15–0.20 ng/ml in the plasma of A-C rats. In A-C rats, treatment with the MDR ligands 4-CD and FGIN-1–27 increased the pregnenolone content in the brain but failed to change the plasma or peripheral tissue content of this steroid. The effect of 4′-CD on brain pregnenolone content was maximal (70–100% increase) at the dose of 18 μmol/kg, 5–10 min after intravenous injection. The effect of oral administration of FGIN-1–27 on brain pregnenolone content was maximal (80–150% increase) at doses of 400–800 μmollkg and peaked at ～ 1 h. That this effect of FGIN-1–27 was mediated by the MDR was documented by pre-treatment with the MDR partial agonist PK 11195 (100 μmol/kg, i.p.). PK 11195 did not affect basal brain pregnenolone content but prevented the accumulation of brain pregnenolone induced by FGIN-1–27. FGIN-1–27 and 4-CD failed to increase the brain concentration of dehydre epiandrosterone in A-C rats. These data suggest that glial cell MDRs play a role in neurosteroid biosynthesis in vivo.     

Barium permeability of neuronal nicotinic receptor alpha 7 expressed in Xenopus oocytes.

The rat alpha 7 neuronal nicotinic acetylcholine receptor was expressed and studied in Xenopus oocytes. The magnitude and reversal potential of instantaneous whole cell currents were examined in solutions containing varying concentrations of either calcium or barium, and in the presence or absence of the intracellular calcium chelator BAPTA. In external barium, application of nicotine elicits an inwardly rectifying response; in calcium the response is larger and has a linear IV relation. Pretreatment of oocytes with BAPTA-AM could not prevent activation of calcium-dependent chloride channels in external Ringer containing calcium. Using an extended GHK equation, the permeability ratio PBa/PNa of the alpha 7 receptor was determined to be about 17. Our results suggest that alpha 7 nicotinic receptors are highly permeable to divalent cations.     

Survey of bovine mycotic mastitis in dairy herds in the State of São Paulo,Brazil

The purpose of this study was to isolate fungi from the quarter milk of cow udders from several dairy herds and to identify the different genera and species involved in mastitis. A total of 2078 milk samples from normal, clinical and subclinical mastitis quarters from 22 dairy herds of 16 districts in the State of São Paulo, Brazil was utilized in this survey. Two hundred and fifty one (12.07%) fungi were isolated from the samples. Two hundred and eight of these (82.86%) were yeasts and 30 (11.95%) were moulds. The fungi were isolated in pure culture (24.77%) or in cultures mixed with bacteria (72.22%). The yeasts isolated were:Cryptococcus spp. (71 strains),Rhodotorula spp. (40),Candida spp. (68),Trichosporon cutaneum (21),Aureobasidium pullulans (7), andPichia ohmeri (1). Moulds classified in following genera were also isolated:Aspergillus (3),Penicillium (3),Alternaria (3),Phoma (3),Epicoccum (2), andGeotrichum (16).     

Nucleotide substitutions at the -6 position in the promoter region of the factor IX gene result in different severity of hemophilia B Leyden: consequences for genetic counseling

Mutations in the promoter region of the factor IX gene result in hemophilia B Leyden, which is characterized by considerable improvement in the disease after puberty. We have found that distinct nucleotide substitutions at the -6 position in the Leyden-specific (LS) region are associated with a different severity of hemophilia B. The proband (aged 2) from one family is a severe hemophiliac with factor IX activity (F.IXC) and antigen (F.IXAg) levels less than 1.0U/dl. F.IXC and F.IXAg levels in two affected uncles are approximately 30% of normal levels. The LS region was targeted for analysis because the phenotypes suggested the inheritance of a factor IX Leyden gene. An abnormal TaqI digestion pattern was found in amplified DNA from the proband, and sequencing showed a G (-6) to C transversion that was linked to the disease in the family. In another family, two brothers (aged 8 and 9) suffer from mild hemophilia with F.IXC ranging from 7 to 10 U/dl and F.IXAg from 3 to 4 U/dl. They are the only documented members of the family with a bleeding tendency. Denaturing gradient gel electrophoresis on amplified fragments from one of the patient's genomic DNA corresponding to the 8 exons and flanking sequences of the factor IX gene suggested a defect only in a segment from the 5 region. This segment showed an altered TaqI digestion pattern, and sequencing demonstrated a G(-6) to A transition that was traced to the patients's mother and a grandmother. The different phenotypes associated with the G (-6) to A purine nucleotide transition compared with a G(-6) to C transversion provide evidence that this area is directly involved in the regulation of the human factor IX gene expression in vivo by binding of regulatory factors. The ability to predict that the conditions of a hemophilia B patient will improve with age has important implications for genetic counseling of the family. Therefore, the LS region should always be included when scanning the factor IX gene for mutations.     

Cryptophytes: An emerging algal group in the rapidly changing Antarctic Peninsula marine environments

The western Antarctic Peninsula (WAP) is a climatically sensitive region where foundational changes at the basis of the food web have been recorded; cryptophytes are gradually outgrowing diatoms together with a decreased size spectrum of the phytoplankton community. Based on a 11-year (2008–2018) in-situ dataset, we demonstrate a strong coupling between biomass accumulation of cryptophytes, summer upper ocean stability, and the mixed layer depth. Our results shed light on the environmental conditions favoring the cryptophyte success in coastal regions of the WAP, especially during situations of shallower mixed layers associated with lower diatom biomass, which evidences a clear competition or niche segregation between diatoms and cryptophytes. We also unravel the cryptophyte photo-physiological niche by exploring its capacity to thrive under high light stress normally found in confined stratified upper layers. Such conditions are becoming more frequent in the Antarctic coastal waters and will likely have significant future implications at various levels of the marine food web. The competitive advantage of cryptophytes in environments with significant light level fluctuations was supported by laboratory experiments that revealed a high flexibility of cryptophytes to grow in different light conditions driven by a fast photo-regulating response. All tested physiological parameters support the hypothesis that cryptophytes are highly flexible regarding their growing light conditions and extremely efficient in rapidly photo-regulating changes to environmental light levels. This plasticity would give them a competitive advantage in exploiting an ecological niche where light levels fluctuate quickly. These findings provide new insights on niche separation between diatoms and cryptophytes, which is vital for a thorough understanding of the WAP marine ecosystem.     

A surplus no more? Variation in krill availability impacts reproductive rates of Antarctic baleen whales

The krill surplus hypothesis of unlimited prey resources available for Antarctic predators due to commercial whaling in the 20th century has remained largely untested since the 1970s. Rapid warming of the Western Antarctic Peninsula (WAP) over the past 50 years has resulted in decreased seasonal ice cover and a reduction of krill. The latter is being exacerbated by a commercial krill fishery in the region. Despite this, humpback whale populations have increased but may be at a threshold for growth based on these human-induced changes. Understanding how climate-mediated variation in prey availability influences humpback whale population dynamics is critical for focused management and conservation actions. Using an 8-year dataset (2013–2020), we show that inter-annual humpback whale pregnancy rates, as determined from skin-blubber biopsy samples (n = 616), are positively correlated with krill availability and fluctuations in ice cover in the previous year. Pregnancy rates showed significant inter-annual variability, between 29% and 86%. Our results indicate that krill availability is in fact limiting and affecting reproductive rates, in contrast to the krill surplus hypothesis. This suggests that this population of humpback whales may be at a threshold for population growth due to prey limitations. As a result, continued warming and increased fishing along the WAP, which continue to reduce krill stocks, will likely impact this humpback whale population and other krill predators in the region. Humpback whales are sentinel species of ecosystem health, and changes in pregnancy rates can provide quantifiable signals of the impact of environmental change at the population level. Our findings must be considered paramount in developing new and more restrictive conservation and management plans for the Antarctic marine ecosystem and minimizing the negative impacts of human activities in the region.     

Low species turnover of upland Amazonian birds in the absence of physical barriers

Aim

One of the oldest and most powerful ways for ecologists to explain distinct biological communities is to invoke underlying environmental differences. But in hyper-diverse systems, which often display high species richness and low species abundance, these sorts of community comparisons are especially challenging. The classic view for Amazonian birds posits that riverine barriers and habitat specialization determine local and regional community composition. We test the tacit, complementary assumption that similar bird communities should therefore permeate uniform habitat between major rivers, regardless of distance.

Location

Upland (terra firme) rainforests of central Amazonia.

Methods

We conducted intensive whole-community surveys of birds in three pairs of 100-ha plots, separated by 40–60 km. We then used dissimilarity indices, cluster analysis, and ordination to characterize differences among the six avian communities.

Results

In all, we detected 244 forest-dependent birds, with an average of 190 species (78%) per plot. Species turnover was negligible, no unique indicator species were found among plot pairs, and all documented species were already known from a complete inventory at one of the three sites.

Main Conclusions

Our study corroborates the classic biogeographical pattern and suggests that turnover contributes little to regional avian diversity within upland forests. Using a grain size of 100 ha, this implies that upland birds perceive the environment as uniform, at least over distances of ~60 km. Therefore, to maximize both local species richness and population persistence, our findings support the conservation of very large tracts of upland rainforest. Our analyses also revealed that the avifauna at Reserva Ducke, encroached by urban sprawl from the city of Manaus, shows the hallmarks of a disturbed community, with fewer vulnerable insectivores. This defaunation signals that even an enormous preserve (10 × 10 km) in lowland Amazonia is not insulated from anthropogenic degradation within the surrounding landscape.