text:
Local resident and mother of 4 children called North Ridgeville Police department for help her with a feral cat and her new kittens. The humane officer who responded to her dilemma acted caliously and irresponsibley by shooting the kittens within 15 feet of where the children were, telling the children they would be better in "kitty heaven." The North Ridgeville Police Chief Freeman condoned the horrendous act calling it "acceptable" and also said the officer WOULD NOT be fired or repremanded for his actions. THIS BEHAVIOR IS TOTALLY UNACCEPTABLE! Police have a DUTY to be an example to citizens and to have enough intelligence to handle a delicate and difficult situation especially ones that deal with animals and children. The officer who shot the kittens should be fired and punished for animal cruelty. The North Ridgeville Police Dept should be aware of ANY Feral cat groups in their surrounding areas as well as other animal rescue groups. Knowing children and family were present, the officer should have taken the kittens to the local shelter or to a rescue group. Link to news blurb about the incident http://bcove.me/zaj872z9
To: Mayor G David Gilluck, North Ridgeville, Ohio
Resulting from the incident where a Police officer from the North Ridgeville Police Department acted caliously and irresponsibly by shooting 5 innocent kittens to death in front of (and where) children who were present, we believe this Police officer should be fired and punished for animal cruelty.
This action was morally wrong, if not common sense wrong by a public official who was supposed to be humane in the removal or control of the 5 kittens. Police officers and humane officers are supposed to set an example for the community.Is this really the type of example you want your Police force to display?
We also request that the policy of killing cats and kittens by shooting them be reviewed and REVISED and deemed as unacceptable by the North Ridgeville Police department and Chief Freeman. North Ridgeville Police department should work in partnership with local humane animal groups, rescue groups, and animal societys in Ohio to give a positive example to children and familys alike when dealing with delicate and difficult situations such as this incident.
Animals of every kind deserve respect and compassion in every situation. We urge you to fire officer Barry Accorti and charge him with animal cruelty as an example that your city will not tolerate this type of irresponsible and horrendous behavior.

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African Journal of Herpetology
ISSN: 2156-4574 (Print) 2153-3660 (Online) Journal homepage: http://www.tandfonline.com/loi/ther20
Genetic diversity and differentiation of the
Western Leopard Toad (Sclerophrys pantherina)
based on mitochondrial and microsatellite
Jessica M. da Silva , Kevin A. Feldheim, G. John Measey , Stephen Doucette-
Riise, Ryan J. Daniels, Lucas F. Chauke & Krystal A. Tolley
To cite this article: Jessica M. da Silva , Kevin A. Feldheim, G. John Measey , Stephen
Doucette-Riise, Ryan J. Daniels, Lucas F. Chauke & Krystal A. Tolley (2017) Genetic
diversity and differentiation of the Western Leopard Toad (Sclerophrys pantherina) based on
mitochondrial and microsatellite markers, African Journal of Herpetology, 66:1, 25-38, DOI:
To link to this article: http://dx.doi.org/10.1080/21564574.2017.1294115
View supplementary material Published online: 23 Jun 2017.
Submit your article to this journal Article views: 2
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Genetic diversity and differentiation of the Western
Leopard Toad (Sclerophrys pantherina) based on
mitochondrial and microsatellite markers
JESSICA M. DA SILVA
*,KEVIN A. FELDHEIM
G. JOHN MEASEY
,LUCAS F. CHAUKE
&KRYSTAL A. TOLLEY
Kirstenbosch Research Centre, South African National Biodiversity Institute, Private Bag X7, Claremont, Cape
Town, South Africa;
Department of Botany & Zoology, University of Stellenbosch, Private Bag X1, Matieland
7602, Stellenbosch, South Africa;
Pritzker Laboratory for Molecular Systematics and Evolution, The Field
Museum, 1400 S. Lake Shore Drive, Chicago, IL 60605, USA;
Centre for Invasion Biology, Department of
Botany and Zoology, Stellenbosch University, Natural Sciences Building, Matieland, Stellenbosch, South Africa;
Department of Biological Sciences, University of Cape Town, Private Bag X3, Rondebosch 770, Cape Town,
Abstract.—Intraspeciﬁc genetic diversity provides the basis for evolutionary change and is
therefore considered the most fundamental level of biodiversity. Mitochondrial DNA (mtDNA)
and microsatellite loci are the markers most typically used in population-level studies; however,
their patterns of genetic variation are not always congruent. This can result in different
interpretations of the data, which can impact on management decisions, especially for
threatened species. Consequently, in this study, we developed and analysed novel microsatellite
markers for the Endangered Western Leopard Toad (WLT), Sclerophrys pantherina, and
compared the results to previously published mtDNA data to compare the level of genetic
diversity between the two molecular markers. The microsatellite evidence showed signs of a
past bottleneck, yet relatively high levels of genetic diversity and low genetic differentiation
between two sampling sites. In contrast, the mtDNA revealed moderate to low levels of
diversity between sampling sites, and strong genetic differentiation. An explanation for the
conﬂicting patterns may be that the current genetic signature, as depicted by the microsatellite
data, is not yet reﬂected in the mitochondrial dataset; and, as such the data are depicting a
timeline for genetic variation within the WLT. Both markers revealed important information
about the two sampling sites, which can help inform conservation management of the species.
Key words.—Africa; Amietophrynus; Bufonidae; conservation; endangered; genetic
Intraspeciﬁc genetic diversity provides the basis for evolutionary change and is therefore
considered the most fundamental level of biodiversity (May 1994). This diversity is gov-
erned by the loss and gain of alleles, which can be the result of mutations, random genetic
drift, natural and sexual selection or migration. In our human-mediated world, ecological
*Corresponding author. Email: firstname.lastname@example.org
African Journal of Herpetology,
Vol. 66, No. 1, 2017, 25–38
ISSN 2156-4574 print/ISSN 2153-3660 online
© 2017 Herpetological Association of Africa
disturbance is proving to be a key driver of changes to the effective population size (N
a species (for an in-depth review, see Banks et al. 2013); and, over time, these changes can
greatly affect a species’genetic diversity (Alcala & Vuilleumier 2014; Ellegren & Galtier
2016). It is well established that a reduction in genetic diversity compromises the adaptive
potential of a species and, therefore, decreases its long-term survival (Lacy 1997; Markert
et al. 2010). This is especially true for threatened species, which can sometimes be charac-
terised by low levels of genetic variation, often due to inbreeding, population bottlenecks,
or the disproportionate effects of genetic drift on small populations (e.g. England et al.
2003; Spielman et al. 2004; Allendorf 2005; Willoughby et al. 2015).
Molecular markers, such as mitochondrial DNA (mtDNA) and microsatellite loci,
are powerful tools for estimating the diversity among individuals and populations
(e.g. Avise et al. 1987; Schwartz et al. 2007). However, patterns of genetic variation
are not always congruent between the two. At times, greater population differentiation
has been found using microsatellite markers (e.g. Lu et al. 2001; Johnson et al. 2003;
De Oliveira Francisco et al. 2013; Kolleck et al.2013; da Silva et al. 2016); while the
reverse has also been documented (e.g. Castella et al. 2001; Yang & Kenagy 2009).
Some possible explanations for these discrepancies include: (i) differences in the selec-
tion intensities acting on each marker; (ii) the different mutation rates between markers;
and, (iii) the N
for maternally-inherited markers, such as mtDNA, differ from those of
microsatellites which are biparentally inherited markers (Johnson et al. 2003). Genetic
diversity is also inﬂuenced by patterns of mating, sex-biased dispersal and other demo-
graphic parameters (Chesser & Baker 1996; Johnson et al. 2003; Yang & Kenagy 2009;
De Oliveira Francisco et al. 2013; Kolleck et al. 2013), which can further contribute to
the discrepancies in genetic diversity estimated between mtDNA and microsatellite
markers. In general, mtDNA is most often used to analyse phylogeographic events,
while microsatellites typically provide more ﬁne-scale resolution of more recent demo-
graphic events (e.g. Avise et al. 1987). Consequently, researchers must carefully deter-
mine what questions they want to address, so that the appropriate markers can be
utilised (Hoban et al. 2013). This is especially critical for research on threatened
species that have small and/or declining populations, such as the Western Leopard
Toad (WLT: Sclerophrys pantherina; previously Bufo pantherina and Amietophrynus
The WLT is endemic to the winter rainfall region of the southwestern Cape of South
Africa where it falls into two historically disjunct areas: (1) to the west of False Bay,
including the Cape metropolitan area, and (2) to the east of False Bay (De Villiers
2004; Measey & Tolley 2011;Fig. 1). Its Endangered status is based on its small distri-
bution (area of occupancy [AOO] 440 km
) and the ongoing reduction in quantity and
quality of habitat associated with urbanisation and agricultural expansion throughout its
range (de Villiers 2004; SA-FRoG 2016; Measey 2011). Adults are explosive breeders,
moving to breeding sites during the antipodean winter (late July to early September)
where they lay large clutches of spawn in strings (Cherry 1992b; de Villiers 2004). Tad-
poles take around three months to reach metamorphosis, and males are known to breed
after one year and females after two (Cherry 1992a).
In order to investigate the structure and distribution of S. pantherina across its range, a
recent study utilised the mitochondrial marker, ND2 (Measey & Tolley 2011), as it was
thought to have sufﬁcient population-level variation (Cunningham & Cherry 2004).
Two genetically distinct populations were recognised that corresponded to the distribution
of the toads (refer to Fig. 1): population 1 –toads within the Cape Metropolitan area
26 DA SILVA ET AL.—Genetic diversity in the Western Leopard Toad
(CMA) –and population 2 –toads east of False Bay (EFB). Moreover,
haplotype and nucleotide diversity were greater for the CMA population compared to
the EFB population, with no shared haplotypes across the two populations (Measey &
The relatively low genetic diversity found among EFB individuals, coupled with local
extinctions and a small number of remaining breeding sites could be an indication that this
population is more vulnerable than the CMA population (Measey & Tolley 2011). With
ongoing development in the EFB lowlands due to agriculture and tourism, the vulnerability
of this population is likely to worsen. Consequently, it has been recommended that this
population be treated as a separate management unit to safeguard the future of the remain-
ing breeding sites (Measey & Tolley 2011).
Given that this previous study only utilised a single mtDNA marker and the two popu-
lations were estimated to have diverged as recently as 1.2 thousand years ago (Measey &
Tolley 2011), the results may not adequately reﬂect the current genetic diversity within the
species. Consequently, in this study, we developed and analysed novel microsatellite
markers for S. pantherina and compared the results to the previously published ND2
sequence data, speciﬁcally for the EFB population, to compare the level of genetic diver-
sity between the two molecular markers.
Figure 1. Map depicting the distribution of Sclerophrys pantherina (inset) in the southwestern Cape,
South Africa. Shaded blue areas show the disjunct distribution of the species. Circles denote known
sampling sites: black-ﬁlled circles denote the two sampling sites included in this study; while the
white ﬁlled circles are sampling sites within the Cape Metropolitan Area (refer to Measey &
AFRICAN JOURNAL OF HERPETOLOGY 2017 27
MATERIALS AND METHODS
Sampling was conducted during the 2010 breeding season in July and August at night
when the toads are most active. Two sampling sites within the EFB were targeted: Uilenk-
raal (S 34° 34′6″E 19° 27′57″) and Klein Paradijs (S 34° 39′10″E 19° 32′1″:Fig. 1)
which are 11.25 km apart in a straight line. Both sites are artiﬁcial impoundments which
S. pantherina have used for oviposition for many years. A single toe clip (the tip to the
ﬁrst articulation on the inside toe on the left foot) was collected for each toad using ster-
ilised surgical scissors. In addition to providing tissue for DNA analysis, the toe clip
was used as an identiﬁcation of previously captured individuals. Each toe clip was
stored in 99% ethanol and stored at -40 °C until DNA was extracted.
Microsatellite Development and Optimisation
Twelve microsatellite markers were developed for S. pantherina (online supplemental
material, Table S1), using an enrichment protocol (Glenn & Schable 2005). Genomic
DNA (gDNA) from one individual was digested with RsaI and XmnI, and SuperSNX24
linkers were ligated onto the ends of gDNA fragments. Linkers act as priming sites for
polymerase chain reactions (PCR) in subsequent steps. Five tetranucleotide [(AAAT)
] and six trinucleotide [(AAT)
] biotinylated probes were hybridised to gDNA in two
separate reactions. Streptavidin-coated magnetic beads (Dynabeads® M-280 Invitrogen,
Carlsbad, CA) were added to the probe-gDNA complexes, and this mixture was washed
twice with 2× SSC, 0.1% SDS and four times with 2× SSC, 0.1% SDS at 52
Between washes, a magnetic particle collecting unit was used to capture the magnetic
beads which are bound to the biotin-gDNA complex. This allows us to capture gDNA con-
taining repeats while other fragments are washed away. Enriched fragments were removed
from the probes by denaturing at 95
C and precipitated with 3 Msodium acetate and 95%
ethanol. To increase the amount of fragments, a “recovery”PCR was performed in a 25 µl
reaction containing 1× PCR buffer (10 mMTris-HCl, 50 mMKCl, pH 8.3), 1.5 mMMgCl
0.16 mMof each dNTP, 0.52 µMof the SuperSNX24 forward primer, 10× BSA, 1U Ta q
DNA polymerase, and approximately 25 ng enriched gDNA fragments. Thermal
cycling, performed in a BIO-RAD DYAD, was as follows: 95 °C for 2 min followed
by 25 cycles of 95 °C for 20 s, 60 °C for 20 s and 72 °C for 90 s, and a ﬁnal elongation
step of 72 °C for 30 min. PCR fragments were then cloned using the TOPO-TA Cloning®
kit following the manufacturer’s protocol (Invitrogen). Bacterial colonies containing a
vector with gDNA (i.e. white colonies) were used as a template for subsequent PCR.
PCR products were then cleaned using Shrimp Alkaline Phosphatase and Exonuclease I
according to the manufacturer’s protocol (Affymetrix, Santa Clara, CA). DNA sequencing
was performed using the BigDye® Terminator v3.1 Cycle Sequencing Kit (Applied Bio-
systems, Foster City, CA). Sequencing reactions were precipitated with 125 mM EDTA
and 100% ethanol and run on an ABI 3730 DNA Analyzer. Primer3 (http://frodo.wi.
mit.edu/cgi-bin/primer3/primer3_www.cgi) was used to develop microsatellite PCR
Optimisation of the 12 markers was carried out in a 10 µl reaction volume containing
approximately 5–50 ng of DNA template, 80 µMof dNTPs, and 0.2 µMof each primer.
28 DA SILVA ET AL.—Genetic diversity in the Western Leopard Toad
The PCR buffer concentrations, MgCl
concentrations and annealing temperatures (T
varied with the Taq used and the loci targeted (Table S1). Thermal cycling parameters
were as follows: 95 °C for 4 min, followed by 40 cycles at 95 °C for 30 s, T
for 30 s,
72 °C for 45 s and a ﬁnal extension at 72 °C for 5–10 min. Successful products were
then combined in a poolplex format and proﬁled at the Central Analytical Facility at Stel-
lenbosch University using an ABI 3100xl Prism (Applied Biosystems, Foster City, CA),
ROX 500 or LIZ 500 as the internal size standard, and POP-7 as the polymer, as per man-
ufacturer’s recommendation. Alleles were scored and binned using the microsatellite
plugin in Geneious version 9.1.5 (Kearse et al. 2012).
Microsatellite genotypes.—Microsatellites were screened for genotyping errors and null
alleles using Micro-Checker version 2.2.3 (Van Oosterhout et al. 2004) as loci possessing
either of these would not typically be considered useful in population-level studies as they
may affect the estimation of population differentiation, for example, by reducing the
genetic diversity within populations (e.g. Paetkau & Strobeck 1995). As such, some
studies have attempted to correct for null alleles in population genetic studies by statisti-
cally adjusting the visible allele and genotype frequencies (e.g. Roques et al. 1999;
Chapuis & Estoup 2007). However, the estimation of null alleles can be biased upwards
in populations that are either inbred or consist of closely related individuals (Chybicki
& Burczyk 2009; Campagne et al. 2012), and, conversely, that estimations of inbreeding
can be biased upwards when null alleles or genotyping errors occur (e.g. Björklund 2005).
Consequently, automatically correcting for null alleles might be unnecessary and inap-
propriate. To better assess the likely presence of null alleles, we conducted a simultaneous
estimation of the inbreeding coefﬁcient (f), null allele frequencies (n), and random geno-
typing failure (b) using the software INEst v2 (Chybicki et al. 2011). This method uses a
Bayesian approach to simultaneously estimate the three parameters, which should provide
a more accurate estimation of each parameter because it allows for the relative contribution
of each through selection of the best ﬁtting model (i.e. model with the lowest Deviance
Information Criterion [DIC]).
The Markov Chain Monte Carlo (MCMC) was run with 500 000 cycles and 10% burn-in,
saving parameters every 100 cycles. The DICs from each model were then compared, with
the better (lowest) scoring model considered a better ﬁt. Given that the effects of null
alleles are locus-speciﬁc (Dakin & Avise 2004), their presence should be reﬂected in the
best ﬁtting models of each population, or in this case, sampling site. An examination of
the p[j,k] parameter from the INEst results should provide conﬁrmation of the locus/
loci containing null alleles This parameter examines the frequency of the kth allele at
the jth locus, where p[j, 0] denotes the null allele frequency at the jth locus
(Chybicki et al. 2011), For these loci, we manually adjusted allele frequencies using the
Brookﬁeld 1 algorithm implemented in Micro-Checker. This method discounts non-
amplifying individuals when calculating null allele frequencies, as opposed to the Brook-
ﬁeld 2 algorithm, which treats non-ampliﬁcations as data and regards them as null homo-
zygotes when calculating null allele frequencies (Brookﬁeld 1996). By correcting for null
alleles in this way, we were able to maximise sample size and retain loci that may have
sufﬁcient variability to detect patterns. All subsequent analyses were conducted on this cor-
AFRICAN JOURNAL OF HERPETOLOGY 2017 29
Genepop on the Web (Raymond & Rousset 1995; Rousset 2008) was used to test for
linkage disequilibrium (LD) between loci using the log-likelihood ratio test (Cockerham &
Weir 1977). Analyses for LD were run with a MCMC dememorisation of 10 000 for 100
batch runs of 1 000 iterations. For each locus, the number of alleles (allelic richness: A
allelic size range, observed (H
) and expected (H
) heterozygosities (Nei 1987), and devi-
ations from Hardy–Weinberg equilibrium (HWE) were also estimated for the two sampling
sites using ARLEQUIN version 126.96.36.199 (Excofﬁer & Lischer 2010). Deviations from HWE
were tested using 1.0 ×10
Markov chains and 100 000 dememorisation steps. To minimise
the possibility of Type I errors, tests for linkage and Hardy–Weinberg disequilibria were
corrected for multiple comparisons by applying Holm’s sequential Bonferroni correction
(Holm 1979; Rice 1989).
ARLEQUIN was used to test for signatures of genetic bottlenecks in the two sampling
sites using the Garza–Williamson M-ratio (G-W: Garza & Williamson 2001) and to deter-
mine the level of gene ﬂow between sites using R
(Slatkin 1995). The M-ratio is esti-
mated according to the equation M= k/ R+1, where k represents the number of alleles at
a locus and R is the associated allelic range (Garza & Williamson 2001; Excofﬁer &
Lischer 2010). Populations that have experienced a reduction in their effective population
size exhibit a larger reduction in allele numbers than range (Excofﬁer & Lischer 2010).
Accordingly, an M-ratio less than 0.68 (value derived from stable wild populations)
would indicate that the sampling site has been through a bottleneck at the locus under
examination, whereas a value closer to one is indicative of stationary/stable populations
(Garza & Williamson 2001; Peery et al. 2012). Gene ﬂow was measured using R
instead of F
because it relies on a stepwise mutation model which is better suited for
the high mutation rates and memory dependent allele mutations found within microsatellite
loci (Di Rienzo et al. 1994; Slatkin 1995). In contrast, F
relies upon the inﬁnite allele
model, which assumes low mutation rates and a mutation process independent of the
prior allelic state (Weber & Wong 1993; Slatkin 1995). The number of migrants (N
between populations was then calculated from the equation, N
= 0.25*(1- F
(Slatkin & Barton 1989), using R
in place of F
. Genetic differentiation can be cate-
gorised as great if R
> 0.15, moderate if R
= 0.05–0.15, and little if R
(Wright 1978). Similarly, the level of gene ﬂow between sampling sites is categorised as
high if N
>1, intermediate if N
= 0.25 to 0.99, and low if N
< 0.25 (Wright 1978).
mtDNA sequences.—The ND2 sequence data for Uilenkraal and Klein Paradijs data from
Measey & Tolley (2011) was used to estimate genetic diversity within S. pantherina.
Speciﬁcally, the number of haplotypes (h), haplotypic diversity (hd), nucleotide diversity
(π), the number of polymorphic sites (S), and the total number of mutations (Eta) were cal-
culated using DnaSP version 5.10.1 (Librado & Rozas, 2009). Haplotype diversity (or gene
diversity) is deﬁned as the probability that two randomly sampled alleles are different; and,
nucleotide diversity is deﬁned as the average number of nucleotide differences per site in
pairwise comparisons among DNA sequences (Nei 1987). DnaSP was also used to deter-
mine the level of genetic differentiation and gene ﬂow between sampling sites; and, Φ
which is analogous to F
, was calculated in ARLEQUIN. F
is an allelic calculation that
assumes all alleles are equidistant from each other; whereas Φ
(the nucleotide diversity
calculation) allows for different distances between different alleles. Lastly, a median-
joining network was created using NETWORK, version 5.0 (Bandelt et al. 1999) to visu-
alise the relationships among the mtDNA haplotypes between the two sampling sites.
30 DA SILVA ET AL.—Genetic diversity in the Western Leopard Toad
All 12 loci were polymorphic with the number of alleles ranging from 3 to18 (Table 1). There
was no evidence of large allele dropout or scoring errors due to stutter peaks; however, the
likely presence of null alleles due to homozygote excess was detected in both Klein Paradijs
and Uilenkraal sampling sites (Tables 1 and 2). Because null alleles are locus-speciﬁc,
WLT16 and WLT39 likely represent ‘true’null alleles. Although the best-ﬁtting INEst
models also indicated the inﬂuence of inbreeding, with average F
= 0.0247 for Uilenkraal
and 0.0588 for Klein Paradijs sampling sites, the effects are minimal. An examination of the
95% posterior probability intervals for F
included zero for Uilenkraal, indicating that there
is no signiﬁcant inbreeding in this population (Table 2).
No signiﬁcant linkage disequilibrium was detected for any of the loci; however, two
loci were found to deviate from HWE (WLT39 & WLT42: Table 1). Because locus
WLT39 was identiﬁed as possibly having null alleles across populations, this is the
most probable explanation for its deviation from HWE. Locus WLT42, on the other
hand, only showed deviations from HWE for Klein Paradijs, indicating that these devi-
ations are most likely attributed to some biological factor such as the Wahlund effect or
inbreeding (Chakraborty et al. 1992). Given the highly fragmented nature of the WLT
habitat, it is possible that small populations have become ﬁxed for diverse alleles.
Evidence of a genetic bottleneck was found across all loci for both sampling sites (M-ratio:
Table 1), which also show fairly similar levels of microsatellite genetic diversity for all indices
examined (Table 3) and exhibit little differentiation (R
: 0.082, P< 0.001; N
Unlike the microsatellite results, analysis of the mtDNA data revealed differing levels of
diversity between Uilenkraal and Klein Paradijs sampling sites (Table 3). In particular,
Uilenkraal was found to have moderate haplotype diversity (hd = 0.567 ± 0.051), yet
low nucleotide diversity (π= 0.00095 ± 0.00022); while Klein Paradijs has low haplotype
diversity (hd = 0.154 ± 0.126) and very low nucleotide diversity (π= 0.00019 ± 0.0016).
The two populations share one haplotype, possessed by the majority of individuals
sampled (Fig. 2), indicating a previously continuous population or past gene ﬂow
between populations; however, the two populations were found to be strongly differen-
= 0.41991, P=0.002).
Although the mtDNA and microsatellite evidence presented here reveal different patterns
with respect to the genetic diversity within the two sampling sites of S. pantherina, simi-
larities were also present with both markers showing signs of population bottlenecks and
the presence of gene ﬂow between Uilenkraal and Klein Paradijs. A possible explanation
for the conﬂicting patterns of genetic diversity may be that the current genetic signature (as
depicted by the microsatellite data) is not yet reﬂected in the mitochondrial dataset due to
the slower evolution of the mtDNA markers. The data may, therefore, be depicting a time-
line for genetic variation within the WLT, with the mtDNA data revealing the more
AFRICAN JOURNAL OF HERPETOLOGY 2017 31
Table 1. Descriptive statistics of genetic variability for 12 Sclerophrys pantherina microsatellite loci across two sampling sites in the southwestern Cape, South
Uilenkraal (n=39) Klein Paradijs (n=41)
HWE Null MN
HWE Null M
WLT1 13 134–194 0.78 0.89 0.0937 Maybe 0.230 11 138–182 0.70 0.76 0.2801 No 0.244
WLT2 13 119–179 0.81 0.88 0.2772 No 0.230 10 123–167 0.80 0.80 0.5150 No 0.222
WLT16 14 142–358 0.62 0.92 0.0006 Maybe 0.065 15 142–358 0.74 0.91 0.0411 Maybe 0.069
WLT25 11 167–219 0.92 0.90 0.8701 No 0.226 9 159–207 0.69 0.81 0.0443 No 0.204
WLT36 14 216–276 0.90 0.89 0.6666 No 0.230 9 216–280 0.75 0.82 0.1783 No 0.138
WLT38 16 275–387 0.89 0.92 0.1888 No 0.150 18 266–387 0.91 0.93 0.4804 No 0.148
WLT39 5 157–181 0.42 0.71 0.0031 Maybe 0.200 3 165–181 0.32 0.56 0.0087 Maybe 0.176
WLT40 10 152–200 0.61 0.85 0.0054 Maybe 0.204 9 148–200 0.71 0.85 0.0643 No 0.170
WLT42 6 179–211 0.39 0.43 0.1636 No 0.182 5 187–211 0.27 0.60 <0.001* Maybe 0.240
WLT44 15 195–347 0.74 0.87 0.0260 Maybe 0.118 8 195–343 0.75 0.75 0.4391 No 0.054
WLT76 4 174–183 0.57 0.54 0.8948 No 0.400 4 174–183 0.73 0.67 0.5540 No 0.400
WLT88 7 197–245 0.67 0.74 0.7256 No 0.163 6 197–245 0.63 0.78 0.0237 Maybe 0.142
, Number of observed alleles; S, allele size range; H
,observed and H
,expected heterozygosity; HWE, Hardy–Weinberg equilibrium Pvalue; Null, presence of null alleles; M,
Garza–Williamson index. Signiﬁcant deviations from HWE are denoted by *, based on a Bonferroni signiﬁcance value of 0.0042 (P< 0.05/12).
32 DA SILVA ET AL.—Genetic diversity in the Western Leopard Toad
Table 2. Results of Bayesian individual inbreeding model for Sclerophrys pantherina. Models were run using all 12 microsatellite loci. Model parameters include
inbreeding (f), null alleles (n) and random genotyping errors (b).
Klein Paradijs (n=41)
DIC ΔDIC Avg (F
) 95% HPD DIC ΔDIC Avg (F
) 95% HPD
nfb 3294.720 1.274 0.0247 0.0000–0.0619 nfb 3084.212 18.488 0.0588 0.0156-0.1069
nb 3295.994 19.807 NA NA nb 3102.700 3.919 NA NA
fb 3315.801 19.398 0.1053 0.0596–0.1504 fb 3106.619 14.333 0.1254 0.0785–0.1674
b 3335.199 50.288 NA NA nf 3120.952 22.009 0.0805 0.0325–0.1261
nf 3385.487 1.369 0.0372 0.0000–0.0819 n 3142.961 14.834 NA NA
n 3386.856 –NA NA b 3157.795 –NA NA
DIC, Deviance Information Criterion; ΔDIC, difference in DIC from the best model; Avg (F
), average inbreeding coefﬁcient for the population; HPD, highest probability density.
AFRICAN JOURNAL OF HERPETOLOGY 2017 33
ancestral relationships and the microsatellite data the most recent demographic structuring.
Both markers revealed important information about the two sampling sites, which can help
inform conservation management of the species.
The mtDNA data uncovered different levels of diversity between Uilenkraal and Klein
Paradijs, with the former showing moderate haplotype diversity and low nucleotide diversity,
and the latter showing low levels of both diversity measures. The low nucleotide diversity esti-
mates suggest that both populations underwent a bottleneck in the recent past. There may have
been a single WLT population in the EFB region, or possibly even across the entire range of
S. pantherina, during the last glacial maxima when areas of the continental shelf were exposed
by lowered sea levels (Schreiner et al.2013; Mokhatla et al. 2015). Then, during the Holo-
cene, the rising sea levels and a distinct drying period may have caused a signiﬁcant retraction
in the distribution of the species, creating a population bottleneck (Measey & Tolley, 2011),
and through time differentiation between the two sampling sites.
Evidence of a bottleneck is also reﬂected in the microsatellite results; however, the data
also indicate strong gene ﬂow between sites and moderate to high levels of overall genetic
variation as indicated by the allelic richness (the most robust indicator for monitoring
Table 3. Mean microsatellite and mitochondrial DNA genetic diversity indices for Sclerophrys
pantherina from two sampling sites.
Genetic diversity Uilenkraal Klein Paradijs
Sample size (n)3941
Allelic richness (A
) 10.667 8.911
Observed heterozygosity (H
) 0.693 0.667
Expected heterozygosity (H
) 0.795 0.770
Sample size (n)2213
Length (bp) 741 741
No. haplotypes (h)32
Haplotypic diversity (hd) 0.567 ± 0.051 0.154 ± 0.126
Nucleotide diversity (π) 0.00095 ± 0.00022 0.00019 ± 0.00016
No. polymorphic sites (S)3 1
Total no. mutations (Eta)3 1
Figure 2. A median-joining haplotype network based on the ND2 mitochondrial gene for Scler-
ophrys pantherina. Haplotypes are represented by circles and show the proportion of toads from
Uilenkraal (white; n=22) and Klein Paradijs (black; n=13). The area of the circle is proportional
to the number of individuals with that haplotype, and the length of the connecting lines is pro-
portional to the number of base changes between haplotypes (refer to Table 3).
34 DA SILVA ET AL.—Genetic diversity in the Western Leopard Toad
genetic diversity and decline: Hoban et al. 2014)ofeachsamplingsite(A
10.7 and 8.9 for
Uilenkraal and Klein Paradijs, respectively; Tab le 3), compared to other toad species (e.g. Wu
&Hu2010; Roth & Jehle 2016;daSilvaet al. 2016). These microsatellite data tend to depict
the more recent genetic diversity for the species; and, on the surface, it could be interpreted that
the two sampling sites are now healthy and thriving after the bottleneck. However, high levels
of genetic diversity do not always equate to viable populations (Habel & Schmitt 2012).
Levels of genetic diversity differ greatly among species and especially between eco-
logical generalists and specialists. Generalist species tend to have high genetic diversity
within their populations and low genetic differentiation among them due to the absence
of bottlenecks, strong gene ﬂow, and little effects of genetic drift. Specialist species tend
towards low genetic diversity and strong genetic differentiation due to small or ﬂuctuating
populations that experience bottlenecks (e.g. Hughes et al. 1999; Schmitt et al. 2005;
Habel & Schmitt 2009,2012). Based on these distinctions, S. pantherina is probably
not a true habitat specialist nor generalist, but might be an ecologically intermediate
species. Such species must maintain high levels of diversity within their populations,
which is facilitated by gene ﬂow, thereby forcing them to exist in metapopulations
(Habel & Schmitt 2012). Without a network of populations, the species might suffer
greatly from the consequences of population bottlenecks, which result in shifts in allele fre-
quencies, often resulting in the loss of genetic adaptations (Luikart et al. 1998).
Unlike many of the Cape lowland endemics (e.g. Xenopus gilli,Microbatrachella
capensis), the WLT appears to have a high tolerance for disturbance with most, if not
all, of its populations currently breeding in anthropogenically altered habitats (Mokhatla
et al. 2015). This adaptability will be essential to the long-term survival of this species,
which will need to counter predicted fragmentation due to newly created impoundments
in both urban and rural areas (Mokhatla et al. 2015). In addition, this species faces the
threat of an invasive congener currently spreading in the CMA range (Measey et al.
2017). However, there might be a threshold of disturbance beyond which populations
are not able to recover. This might very well explain the loss of other sampling sites in
the EFB region, which were known prior to the 1990s (i.e. Pringle Bay, Betty’s Bay and
Kleinmond: Measey & Tolley 2011;Fig. 1).
Given that the EFB is subject to ongoing development, the two sampling sites exam-
ined here, especially Klein Paradijs, may also be at risk. It is, therefore, imperative that the
current genetic diversity within these two sampling sites be maintained over time, with
conservation efforts focused on preserving connectivity between sites to ensure adequate
gene ﬂow between sampling sites. Future studies using these novel microsatellites could
then be used to monitor the frequency of dispersal between sampling sites and, ultimately,
the extent of genetic erosion/preservation within the EFB.
We would like to thank the South African National Biodiversity Institute and the Field
Museum’s Pritzker Laboratory for Molecular Systematics and Evolution for providing
logistical support and funding for this study. Additional funding was provided by the
SANBI-NORAD Threatened Species Programme. Many thanks also to the landowners,
who gave permission to access their property, Susanna Fuchs and Aletta Groenwald.
Sampling was conducted under permit from the Western Cape provincial conservation
authority, CapeNature (#AAA004-00090-0035) and an ethical clearance certiﬁcate
(# 0001/08 from SANBI Ethics Committee).
AFRICAN JOURNAL OF HERPETOLOGY 2017 35
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Received: 29 September 2016; Final acceptance: 2 February 2017
38 DA SILVA ET AL.—Genetic diversity in the Western Leopard Toad

domain: literature


text:
Pepperoni Pizza Can Be Murder
Kensington Publishing, August 2010
Eleanor Swift will not let the reputation of A Slice of Delight, her pizza parlor, be sullied by customer complaints or by murder. When Wade Hatcher, the brother of her delivery guy Greg, is murdered in her kitchen, she won’t rest until she knows what really happened. After all, the pizza next to the victim was not one of hers! Who would bring a strange pizza to a pizza parlor?
There are obvious reasons to suspect Greg. He and his brother had issues – money, women, and family history. Still, Eleanor and her sister Maddie don’t believe Greg is a killer. She knows Greg is not the person who robbed her a few nights before the murder. The police chief just doesn’t buy it, any of it.
In the meantime, there are the romance issues. Eleanor still isn’t over Joe, her dead husband. Maddie is on the look-out for her next husband, a man who will be one of many. Maddie wants Eleanor to pay more attention to David Quinton, a good friend of Eleanor’s. Eleanor doesn’t want to lead him on, and she’s not sure she wants to take the relationship any farther. Still, he’s a nice guy.
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Reviewed by P.J. Coldren, July 2010.
Note: Chris Cavender is a pseudonym for mystery author Tim Myers.


text:
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text:
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domain: chemistry


We always recommend using an uninterruptible power supply (UPS) as a first step. Please to be sure that anything important to keeping your connection alive is connected to a UPS. This includes your modem, switches, etc!
Next, try using a different outlet or power supply to verify that another piece of hardware isn't causing this issue.
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text:
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I will discuss the fish, filling station, the bight, first death in nova scotia and sestina elizabeth bishop: elizabeth bishop sample essay:. The first occasion in which we “first death in nova scotia” by elizabeth bishop this is clearly demonstrated in her poem “first death in nova. Elizabeth bishop: poems, prose, and letters is kept in print by a gift from deborah and first death in nova scotia filling january first personal essays. Essays and criticism on elizabeth bishop winslow” served as an inspiration for bishop's “first death in nova scotia elizabeth spires (essay date.Download

domain: artistic


In a highly-anticipated culinary showdown at the San Diego Bay Wine & Food Festival, a little duck fat and the gourmet skills of one local chef reigned supreme.
Chef Rich Sweeney, of San Diego’s renowned R-Gang Eatery, was crowned “Chef of the Fest” Saturday at the 2013 San Diego Bay Wine & Food Festival’s Grand Tasting event, the conclusion of the week-long foodie fest.
For the big win, Sweeney effortlessly whipped up some “Duck Fat-Fried Beignets with Burnt Orange Cream and Smoked Duck ‘Dust’ and Crispy-Glazed Duck Skin” – no big deal.
His tasty treats won him the coveted culinary title. When he took the microphone and accepted the honor, the chef had this to say to the cheering crowd:
“[I owe this] to the power of duck fat and whole lot of fat kid creativity,” Sweeney joked.
Each year, the Chef of the Fest competition brings San Diego chefs head-to-head in a fierce battle judged by the San Diego Chapter of the American Culinary Federation, along with celebrity judges.
The judges couldn’t resist Sweeney’s decadent duck-based creation and he walked away with a $3,000 grand prize and feature spread in “Cooking Light” magazine, among other victory swag.
With this being the 10th anniversary of the San Diego Bay Wine & Food Festival, organizer Michelle Metter said Sweeney’s dish, as well as other creations presented by Chef of the Fest participants, did the big event justice.
“I am so impressed with the quality of food and level of innovation in each dish,” said Metter. “With creativity and execution at an all-time high, we commend the winners of this year’s competition on their hard work and passion for culinary artistry.”
In the end, a total of 42 competitors were narrowed down to 10 finalists by judges using a blind-tasting format and graded point system. Participants were ranked based on cumulative scores considering their dish’s presentation, flavor, innovation, quality of product and preparation in the allotted time.
Following Sweeney’s grand prize victory, the other top 10 finalists included, in order:
• Carissa Giacalone of Vela Restaurant (Slow-Braised Pig Jowls and Snouts, Crispy Bearnaise, Chicharones and Pickled Quail Egg)
• Nicholas Bour of AVANT Restaurant (Red Royal Shrimp “Ceviche” with Ponzu Gelee and Crispy Nori)
• Derek Jaeger of The Cravory (Ultimate Ice Cream Sandwich)
• Chad White of Plancha Baja Med (Chicken Hearts, Nopal, Masa, Epazote, Queso Fresco, Cebolla Salsa, Guajillo Mole)
• Jorge Gutierrez of Romesco (Plantain Tostada with Wild Salmon)
• Tommy Fraioli of Sea Rocket Bistro (Pork Belly, Watermelon, Apple, Kimchee)
• Joe Bracero of Katsuya San Diego (Sesame Seed Oil-Marinated Ahi with Roasted Corn, Crisp Watercress, Radishes, Gochujung Walnuts with Lotus Root Chips)
• Eric Bauer of Veladora (House Smoked Lamb Breast with Chestnut and Quinoa Salad, Currants, Mint, and Pomegranate Yogurt)
• Todd Nash of Bub's at the Ballpark (Asian Chopped Chicken Salad with Romaine, Napa Cabbage, Radicchio, Carrots, Mint, Cilantro, Scallions, Grapes, Almonds, Chicken, Crispy Wontons, Lime Hoisin Dressing)
Meanwhile, Francesca Penoncelli of BiCE Ristorante took the win in the vegetarian/raw category and Carissa Giacalone was crowned winner of the meat category. Chad White took the win in the poultry category and Nicholas Bour won the seafood category. Finally, with his beignets, Sweeney also took the dessert category.
Organizers say a combined $50,000 in cash and prizes were awarded to this year’s Chef of the Fest winners.
To learn more about the annual San Diego Bay Wine & Food Festival, click here.


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mcdonalds Essay,full Task,research,mcdonalds In Asia,mcdonalds In India,
NEW DELHI India is increasing a covert uranium enrichment plant that might probably help the development of thermonuclear weapons, a defence research group said on Friday, elevating the stakes in an arms race with China and Pakistan. But native businessmen are betting that as international companies put money into the nation, suppliers will observe market research company in india. A scarcity of fine infrastructure and suppliers are the biggest hurdles to making technology products in the nation, forcing most of India's greater than a hundred completely different telephone corporations to get their products from the mature markets of China and Taiwan.
The frontier from Bhutan eastwards has been clearly outlined by the McMahon line which was fixed by the Simla Convention of 1914.... Our maps present that the McMahon line is our boundary and that's our boundary — map or no map.” Since 1938, however, the Survey of India maps have been surreptitiously altered, displaying the McMahon Line, with the phrase ‘Undemarcated' imprinted on it.
CIA could have given this information to India, however they selected to not. The entire idea was to make India and China bitter enemies, and provoke them to struggle a struggle to sow permanent seeds of mistrust. The border between India and Tibet had remained undefined and undemarcated when the direct rule of India by the British led to 1947. Huge areas within the north-east, mountainous and sparsely populated by tribes, and within the north-west, mountainous, icy and desolate, had been by no means ever beneath the administration of India.
He retired from the military in 1961, nearly 15 months earlier than the Chinese language invasion of India in November 1962. China had lengthy suspected that India was an lively however covert party to this motion. This refined declaration of warfare towards China was the fruits of a coverage that Nehru had been pursuing since as early as April 1947 when India was still a British colony. On 25 April 1947, the exterior affairs division of the federal government of India, of which Nehru was in cost as a member of the viceroy's ‘interim authorities', informed the British secretary of state for India.
On 15 August 1947, the day India grew to become independent , the British mission in Lhasa (Tibet's capital) formally became the Indian mission in a shady manner. In the case of Sikkim, India in 1949 seized the opportunity of a neighborhood rebellion towards the ruler to send in troops and produce the state into closer dependence as a protectorate than it had formally been below the British. In the same yr in 1949, India signed a treaty with Bhutan, through which we took over Britain's right to information Bhutan in foreign affairs. India allowed Chinese language meals material for its troops located close to Yatung to go through Calcutta and Gangtok.

domain: geography


Goodwill Manasota to host job fair Wednesday
BRADENTON- Goodwill Manasota will host its 2018 Job Fair this Wednesday, September 26th.
The non–profit invites the public to come out to their Corporate Campus, located at 2705 51st Avenue East in Bradenton from 10 am–2 pm for a job fair to offer free employment opportunities for all job seekers, including veterans and their families.
More than 35 representative from amazon to the Ritz-Carlton to Manatee County Sheriff’s Office will be in attendance.
For more information call 941-355-2721 Ext. 160.


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First there was the violence and now there is the justice.
After nearly five nights of violence in towns and cities across England, riot suspects are appearing before the courts where magistrates and judges are under pressure from politicians to hand out severe punishments.
In London, the City of Westminster Magistrates Court remained open all night to deal with a backlog of riot suspects held in custody since trouble erupted Aug. 6. Police say some were arrested at the scene of crimes, while others were arrested after having been identified by surveillance cameras or witnesses.
In a speech to the recalled British parliament, Prime Minster David Cameron said there had been more than 1,200 arrests across England, including 922 in London alone.
“It is criminality pure and simple. And there is absolutely no excuse for it," he told members of Parliament. "We have seen houses, offices, and shops raided and torched, police officers assaulted and fire crews attacked as they try to put out fires, people robbing others while they lie injured and bleeding in the street, and even three innocent people being deliberately run over and killed in Birmingham.
“We will not put up with this in our country. We will not allow a culture of fear to exist on our streets," he said. "And we will do whatever it takes to restore law and order and to rebuild our communities.”
[ Video is no longer available. ]
Suspects range from a postman to a millionaire's teen daughter
Despite public outrage few people have turned up at courts to see justice meted out.
At Camberwell Green Magistrates Court, south London, David Pickering from Brixton was the sole person sitting in the public benches waiting to see the court proceedings. He said: “I just wanted to see their faces and who they are. I’m angry about what’s happened and want to know why think they can break windows and steal from shops as if it doesn’t matter.”
In the dock was Piotr Dziedzic, a Polish immigrant who police found in a betting shop in Clapham. Charged with burglary with intent to steal, he appeared indifferent to the charges. He was not required to enter a plea and was remanded in custody.
Others on the court list face similar charges ranging from burglary to possessing an offensive weapon.
At Westminster magistrates court it was revealed that university graduate Natasha Reid had handed herself into police because she was "unable to sleep" after stealing a television from an electrical store in north London.
Despite Ms. Reid acknowledging her remorse and guilty plea, district judge Elizabeth Roscoe said she still faced prison ahead of sentencing because of the serious nature of the offenses.
In other courts the faces of those allegedly taking part in the riots emerged and many were not the impoverished stereotypes assumed to have been behind events. They included a postman, school mentor, students, a lifeguard, a charity worker, and a millionaire’s teenage daughter, Laura Johnson, accused of stealing £5,000 ($8,100) worth of electrical gear from a chain store in southeast London.
Perhaps most shocking was the appearance of an 11-year-old boy at Highbury court in north London and a 12-year-old in Manchester, both charged with looting and stealing.
Peace largely restored
While the courts deal with the list of defendants, peace has largely been restored in London – where 16,000 police will again be patrolling tonight – and other cities such as Birmingham, Wolverhampton, Manchester and Liverpool.
Insurance experts have estimated that claims from riot-hit businesses could run to £200 million ($324 million). Many smaller traders, already struggling in the recession, do not have insurance. In his speech today, Mr Cameron announced a £20 million ($32 million) program to help affected businesses and a £10 million ($16 million) recovery package to help local councils pay for the riot damage.
In a shift away from traditional British policing, Mr. Cameron said police might be allowed to spray rioters with dye to help identify troublemakers, possibly from water cannon.
He also said that police might be given new powers to remove face masks, possible bans on would-be rioters using social-network sites to encourage trouble, and initiatives to stop the spread of gang culture. He cited the successful implementation of the task force set up in Glasgow, Scotland, in 2008 to combat gang membership and the Boston cease-fire project set up in the 1990s.
Cameron pressured to curtail budget cuts on police
Cameron, despite earning plaudits since returning from holiday on Tuesday to take charge of the crisis, is coming under increasing pressure to stop police budget cuts because of the credit crunch, which critics say will lead to as many 16,000 officers lose their jobs.
Opposition leader Ed Miliband of the Labour Party said: “The events of the last few days have been a stark reminder to us all that police on our streets make our communities safer and make the public feel safer.
“Given the absolute priority the public attaches to a visible and active police presence, does the prime minister understand why they would think it is not right that he goes ahead with the cuts to police numbers? Will he now think again on this issue?”
He was joined by London mayor Boris Johnson, a conservative colleague and Oxford University friend of Cameron. He told the BBC: “If you ask me whether I think there is a case for cutting police budgets in the light of these events, then my answer to that would be a 'no.' ”


With over a billion active users, the recent launch of Instagram Reels, and a new monopoly in the aesthetic sector of social media, Instagram can be a goldmine for the right company with the right strategy.
That said, if you’ve tried it, you may have realized that it’s not the simplistic photo-based platform that it used to be. In fact, Instagram is more of a microcosm within the social media universe, with different aspects and characteristics that can take you (and your audience) in several different directions.
Like we said, Instagram is a multi-faceted platform with many options on how to connect with your followers and your overall target audience. There is no one way to post. So if you want to understand how to use the platform, you have to understand how it is designed to be used.
The Instagram feed is the fundamental feature of the platform. While it has evolved over the years, the user experience hasn’t changed much. The feed is where you post images and videos that you want to be visible directly on your profile and on your followers’ feeds.
The best way to use your feed is to create an aesthetic for your brand.
Think in terms of “first impressions”. What do you want your followers to think and feel about your brand as a whole? What do you want to be associated with? Put those visuals in your feed, where they will stay long term while capturing the attention of your audience (and potential new followers).
Stories are probably the most popular feature on Instagram. They appease shorter and shorter attention spans while giving creators enough space to convey a highly engaging message.
Built to compete with Snapchat, Instagram Stories give you 15 seconds to record a video or image that grabs your audience’s attention and expires after 24 hours.
They also come with interactive features like questions and polls, encouraging your followers to interact with you directly (without having to try too hard).
Use stories to start conversations, run short-term promotions, provide news updates to your followers, provide snapshots of a day in the life, and take advantage of the wave of current events.
IGTV is exactly what its name suggests, a television station located within the Instagram platform. This is where you can host longer videos that include interviews, tutorials, long-form creative segments, or whatever your audience cares about.
Instagram Live gives you the ability to connect with your followers on their stories in real time. This is a great feature for connecting with them by asking questions and answers, giving live announcements of product / service changes and launches, and live behind-the-scenes tours with you.
Diehard fans love the opportunity to connect directly with brand leaders to engage with them.
Ultimately, you want to find a good balance between using each type of post listed, so that your users have different ways of interacting and engaging with you.


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Berlin parent representatives: Home office plus childcare is not possible
Berlin parent representatives are critical of the demand for more work in the home office. “Working from home means I can not having a normal working day if I have to look after children at the same time. That doesn’t work, ”said the chairman of the state parents committee for day-care centers, Corinna Balkow, on Friday.
Im Homeoffice To work is one thing in view of the pandemic situation sensible measureto save journeys and limit contacts. Many parents, especially in cramped living conditions, are faced with major problems. “Home office and childcare is not possible with 100 percent work performance.”
That is why politicians and employers must pay more attention to the needs of families, said Balkow. She hit about additional vacation days for parents in front. This would have the advantage that, unlike in the case of the doubling of the childhood illness days per parent from 10 to 20, which has already been decided by the Bundestag accept no loss of earnings would have to.
To discuss whether too many Berlin parents use the daycare centers, although they are currently only set to emergency care, said the chairman of the parents’ committee: “We see a lot of parents who try to look after their children at home and a lot of whom this is not possible and for whom it is completely justified that they bring the child to daycare.”
The situation in the Berlin daycare centers, however, is very different. “ The re are certainly daycare centers with a childcare rate below 30 percent,” reported Balkow. “Individual daycare centers are sure to be full.” But she also observed that Parents be very careful and sometimes do not make full use of care services. Some parents could live with limited childcare hoursespecially if there is to compensate for vacation or, for example, a child benefit bonus give. “
The n fewer children would come to daycare centers every day.”
The federal and state governments want over on Tuesday advise possible further tightening of corona restrictions. According to the government spokesman, this includes expanding work in the home office. (dpa)
[ source link ]
Corona crisis Berlin aboveaverage number employees Berlin fire department infected Berlin

domain: politics


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4 weeks ago
Marvelous picture and coloring 💖💖💖
That is pretty nice praise coming from you, T. Oh, I’m using my voice activated messages so your name is just a letter! Sorry about that. Anyway, thank you so much. Hope you are doing well, and I am loving your gallery as always.@Tee Hoth
Fabulous, I'll have one!!
You are so cute! Love your comments. Thanks @V3RTIGO


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A Twin Cities has been ordered to cease production after children suffered lead poisoning.
According to Bring Me The News, the Minnesota Department of Health and Department of Labor and Industry issued a temporary order for Water Gremlin in White Bear Township to cease manufacturing its lead products after a Saturday inspection.
Commissioners with the MDH and DLI have also asked the Ramsey County District Court to extend the order until Water Gremlin has made moves "to prevent the lead poisoning of workers' children."
Tests showed workers' children had high blood lead levels linked to lead dust unknowingly brought home from Water Gremlin by their parents.
Read more here.

domain: law


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2023 is just around the corner, which means it’s time to start thinking about holiday gifts! If you’re looking for something special for the music lover in your life, look no further. I’ve compiled a list of the best gifts for your piano teacher, music-loving family member, and piano students of all levels.
Whether you’re looking for a new piano book, a set of headphones, or a beautiful piece of jewelry, we have a unique gift idea for everyone on this list. So please sit back and enjoy as I take you through some of my favorite gift ideas for piano lovers this holiday season!
This post may contain affiliate links. As affiliates of the Amazon associate program, Modacity, Dr. Josh Wright ProPractice, Musicnotes, and Piano Marvel, I may receive a commission at no extra cost if you purchase through a link. Please see my full disclosure for further information. I take no credit for the images appearing on this page. All photos are courtesy of Canva.
Jump to a section:
- Practical Gifts
- Fun Gifts
- Studio (or Home) Decor
- Personalized Gifts
- Gifts for Piano Students
- Christmas Ornaments
Sometimes the perfect gift also falls under the category of the most practical gift!
Metal Music Book Clip and Book Holder
Although it may not seem exciting, a music clip can make all the difference between a musical triumph and tragedy. And anyone who has ever had the experience of trying to hold a book open and play piano simultaneously will echo that sentiment!
This particular music clip is functional and beautiful. It’s a thoughtful gift for any pianist in your life!
LED Piano Desk Lamp
A lamp is a gift that keeps giving long after the Christmas lights have been packed away. And this one is convenient because it can either sit with a base or be clipped to the music stand.
This model also features 3 different light settings and has a dim feature to adjust brightness levels. It’s a practical gift that any music teacher will appreciate!
Music Throw Blanket
What’s better than snuggling up with a soft blanket while listening to some of your favorite classical tunes? This holiday season, give the gift of music and comfort with this beautiful throw blanket.
Who says piano gifts have to be all work and no play? Check out these cool gifts that are perfect for any music lovers in your life!
Piano Player Facts Coffee Mug
Start their day off with a cup of coffee (or tea) in this quirky mug! This is a great gift for piano enthusiasts!
Piano Paperweight Crystal Glass Cube
This beautiful piano paperweight is an excellent gift for the pianist or music lover in your life. It’s also a great way to add a touch of elegance to any desk or workspace!
Although not strictly limited to the piano, Skillshare is an online learning platform offering various classes. You can learn multiple skills ranging from painting to photography. There are also several unique music classes that any musician will find interesting.
Studio (or Home) Decor
One of the great things about piano-related gifts is that they make lovely home decor items!
Piano Hanging Wall Art
This particular piece of wall art is simple and elegant. It would make a beautiful addition to any music room or living space!
5-Tier Ladder Shelf
With this ladder shelf, you can give the gift of more space to display all those cute piano knick-knacks and keepsakes. It’s also useful as a bookshelf because I don’t know any pianists with enough shelf space for their music books!
Personalized gifts are always a hit! They show you’ve thought about what would make the recipient feel special.
20 Ounce Personalized Coffee Tumbler
This beautiful 20-ounce tumbler can be personalized with a name. It’s perfect for the piano lover who enjoys a hot cup of coffee or tea on the go!
Personalized Phone Dock and Desk Organizer
This personalized phone dock and desk organizer is perfect for the piano teacher or student in your life. It’s a great way to help piano players keep track of their phones and other tiny yet essential items.
Personalized Name Art
This one could fall under either a personalized gift or decor. One of my colleagues purchased it for me, and I absolutely love it!
This is a unique gift that the recipient will appreciate and remember for years to come.
Gifts for that Special Piano Student
Whether you have a little one who is just starting out or a more experienced player, there are plenty of great gift ideas for piano students!
Piano Note Guide for Beginners
Some students benefit from putting those note stickers on the piano. But they can leave sticky residue behind, which can ruin piano keys.
A simple alternative is these silicon piano stickers that sit at the top of the keys and don’t stick to anything. It can be a great way to jumpstart learning for younger players without ruining your piano!
Adjustable Piano Foot Rest
One of the most frustrating aspects of learning piano as a young child is not reaching the floor. Concentrating is more difficult for younger learners when they can’t put their feet on the ground. And with a strong base, learning proper techniques is more manageable.
Luckily, this piano footrest is the ultimate solution to an age-old piano problem! The footrest is adjustable, so you can move it as your child grows.
It’s a fantastic tool to give your young pianist the best start in their piano journey!
Soundbrenner Pulse Wearable Metronome
Metronomes are a valuable piece of equipment that all pianists will need at some point. In the past, metronomes have only come in traditional or digital formats. But a company called Soundbrenner is revolutionizing this technology.
Soundbrenner has now come up with a wearable metronome. The technology from Soundbrenner is a better way to feel the beat and stay in rhythm.
This incredible technology allows you to use a metronome while performing, which wasn’t possible before. And if you play in a band, you can also sync the metronomes, so everyone plays at the same tempo during practice sessions or performances.
Metronomes are essential for musical development; this model is the ideal gift for students at all levels.
This is one piece of equipment that I can’t wait to try out for myself! Stay tuned for a full review post on the Soundbrenner wearable metronome.
Although paper sheet music will never go out of style, technology has given us a better way to store and carry music books.
With the iPad, your budding piano student can bring their entire music library anywhere. Pianists of all levels have instant access to any piece of music at any time.
Many fantastic game-type learning apps can help accelerate learning. Regardless of your piano student’s current playing level, the iPad is a tremendous music-learning resource!
Donner Wireless Page Turner
This is another piece of equipment that all pianists will find helpful if they play from an iPad.
This device allows you to turn pages hands-free, so you can focus on playing the piano and not worry about flipping pages. Believe me, when I say that manually flipping pages on an iPad is a challenge!
Donner solves that problem with this slick device that helps you stay focused on playing the music instead of worrying about a page turn. It’s also compatible with many digital sheet music apps, including Musicnotes, so that you can use it with your iPad!
This page-turner is also very reasonably priced, which is always a bonus.
If you’ve just started your little one in lessons, now is the perfect time to consider buying a piano. Many parents opt for a digital piano to start out with because they are portable and take up less space than an acoustic piano.
Having an instrument at home can promote a love of music in children. And even if your child doesn’t do formal lessons, there are many YouTube tutorials on how to play covers of virtually every song known to man.
Learning an instrument has so many fantastic benefits for the brain. And with an instrument at home, your child can experiment and fall in love with music!
If you’d like guidance on making a digital piano purchase, check out my guide to the best budget digital pianos.
Adjustable Artist Piano Bench
Give your piano student the gift of comfort with an adjustable artist bench. This bench ranges in height from 18″ up to 21″ to accommodate your growing piano student.
And the leather padding offers comfort for all those hours spent practicing! It’s a great gift idea that’s luxurious and also serves a practical purpose.
Josh Wright ProPractice Membership
If your piano student is serious about playing classical music, a membership to Dr. Josh Wright’s exclusive ProPractice program is also a great gift! Dr. Wright is a gifted classical performer, and in this course, he creates detailed videos on how to play some of the most famous pieces in the repertoire.
Although he is a fantastic performer, he is an equally gifted teacher. The videos are interesting and will leave anyone passionate about classical piano feeling empowered and inspired to try their favorite pieces.
Always an appropriate Christmas gift, ornaments offer the opportunity for a special gift that the piano lover in your life can enjoy year after year! And these ornaments also make great stocking stuffers!
Broadway Gifts Grand Piano Ornament
I love the intricate detail offered by this cute grand piano ornament! From the music on the stand to the strings inside the piano, it’s a delightful little ornament guaranteed to bring a smile to any piano lover’s face year after year!
Broadway Gifts Upright Piano Ornament
This is another charming ornament that any classical music lover would love hanging on their Christmas tree!
Piano ties, t-shirts, and sweatshirts are great gifts for your piano lover! And you can find a wide variety of options to suit any taste.
Piano Pullover Hoodie
I love this cute woman’s hoodie! It’s another fun yet practical gift.
Piano Neck Tie T-Shirt
It’s formal and casual at the same time. And it’s the perfect gift for that piano-loving guy in your life!
Spontaneous Creativity T-Shirt
Celebrate spontaneous creativity with this t-shirt designed to put a smile on any music lover’s face!
Steven Harris Piano Keys Men’s Neck Tie
This is a handsome and stylish piano tie that any man would be proud to wear!
Piano jewelry is a unique and beautiful way to show your love of music!
JOYID Pendant Necklace
This necklace is simple yet stunning! It would make an excellent gift for any woman who loves music.
Stainless Steel Piano Ring
This unisex ring is also a fidget spinner. The ring is a great little piece for anyone who loves to fidget!
Infinity Collection Music Charm Bracelet
This bracelet is a beautiful and unique way to show your love of music! It would make an excellent gift for any woman who loves classical music.
Naimo Vintage Piano Design Jewelry Box
This beautiful jewelry box would make a lovely gift for any woman who loves to keep her jewelry organized and safe. The vintage piano design is gorgeous!
I hope you enjoyed this ultimate piano lover’s gift guide for the 2022 holiday season! I’m sure you’ll find the perfect gift for that special someone on your list. And if you have other great gift ideas, please share them in the comments below!
If you loved this post, check out a few of my other favorites:
- The 30 Most Famous Classical Piano Pieces of All Time
- The Best Piano Bar Songs of All Time
- Can You Really Learn Piano Without a Teacher?
- Your Ultimate Guide to the Perfect Piano Practice Routine
- The Ultimate Guide to Getting Your Kids to Practice Piano
- Why You’ll Never Regret Enrolling Your Kids in Piano Lessons
- How to Find the Right Piano Teacher for You


First making its debut in 1999, the BMW X5 is a mid-sized luxury SUV that featured all-wheel drive and was available with either a manual or an automatic transmission. The car manufacturer branded the BMW X5 as a “Sports Activity Vehicle” (SAV) rather than an SUV, in an attempt to highlight its on-road ability despite its small stature.
The first generation of the X5 was manufactured between 1999 and 2006, with the second generation running from 2006 to 2013, and the third generation being manufactured from 2013 to 2018. The fourth and current generation, known as the BMW G05 X5, was unveiled on June 6, 2018. In 2020, a plug-in hybrid model was introduced to the market.
Anyone who has ever owned one of these luxury cars knows about their perks, as well as their drawbacks. While the BMW X5 vehicle is a splendid automobile, it does come with its own set of unique issues.
Here are all of the BMW X5 problems that you should know about and how to fix them.
Excessive Wind Noise Troubles
If you have ever owned a BMW X5, you probably are no stranger to the sound of the wind gusting by when you are traveling at high speeds. If you’re new to this car, be sure to listen for this excessive wind noise when taking the vehicle out for a test drive. This issue is caused by faulty door aperture seals, which will fail over time and eventually disintegrate.
To fix this problem, you will need to replace the whole seal around the door. This is an inexpensive car component to buy, and will require a simple fitting at your auto body repair shop.
There is a well known issue with the BMW X5’s coolant system, which can develop a leak over time. This important part of your car keeps your engine from overheating and compromising its overall functionality. The way your BMW’s coolant system works is by allowing the engine to maintain its working temperature within safe limits by circulating under pressure a mixture of coolant and water. If there are leaks, the engine will overheat, leading to damage and expensive repairs.
There are many different things that can cause your BM5 X5 to leak, including:
- A blown head gasket
- A hole in your radiator
- Brittle or hard hose attachment points
If you notice puddles of coolant accumulating underneath your car, or if the “fill coolant” light on your dashboard remains illuminated even after you have efficiently topped off the system, you should take your BMW X5 into a mechanic right away.
A coolant leak repair cost can cost you up to $400 if you have a blown head gasket, while replacing a hose is a quick fix and will run you between $35 and $65 for the entire process. Replacing your radiator will cost you around $300 or more.
One of the main culprits behind your coolant issue in your BMW X5 may be the inlet manifolds. They are a known vulnerability in this particular type of BMW model and can cause numerous issues with your engine.
Your BMW’s inlet manifold is the component of its engine that supplies a mixture of fuel and air to the cylinders. The main purpose of this part is to evenly distribute the combustion mixture to each intake port in the cylinder heads.
Some symptoms and signs of bad inlet manifolds include:
- A misfiring engine or decrease in power, acceleration, and fuel economy: This is one of the main symptoms of an intake manifold issue. As your BMW acquires mileage, the intake manifolds can wear out and eventually start to leak.
- Coolant leaks: Another sign of faulty intake manifolds is a coolant leak. This can produce a distinct smell, along with drips or puddles underneath the front of your car.
- An overheating engine: This is yet another symptoms of a possible intake manifold problem. A coolant leak will eventually lead to your engine overheating when the level of the coolant drops too low. However, engine overheating can also happen without any visible leaks taking place.
If you suspect that your BMW X5 vehicle has a bad intake manifold, it’s time to take it into your BMW dealership or local service center right away to prevent any further damage from happening.
Air Mass Sensors
Another issue that can be seen in the BMW X5 car that is widely reported is a problem with the air mass sensors. This is more frequent in older diesel models. This part of your car is a sensor that is used to determine the mass flow rate of air entering into a fuel-injected internal combustion engine. It’s critical for the engine control unit to balance and deliver the proper fuel mass to the engine.
Some signs of faulty air mass flow sensors include:
- Your BMW’s engine is difficult to start or does not turn over right away
- The engine stalls shortly after it is started
- Your car jerks or hesitates when you accelerate
- Excessively rich or lean idling
- Your BMW X5’s engine will “hiccup”
- Your engine drags or hesitates while under idle or load
If you think that your car has bad air mass sensors, take it to a qualified mechanic in order to get a complete computer diagnostic test. If you do need these parts replaced, it will cost you between $300 and $450.
Transmission and Gearbox
If you think your car is suffering from this type of issue, check under it for excessive puddles of oil. This could indicate that your BMW X5’s gearbox plug sleeve seal is defected. This part is prone to failure, causing your car to leak oil. Although this is a fairly cheap component of your car to buy, it’s extremely difficult to install. So seek the assistance of a qualified BMW mechanic.
Are you considering buying a BMW X5 that was made in either 2000 or 2001? Be careful. This is because a large number of these cars made during those years were recalled due to a serious issue found in the steering column. The steering wheel would stop adjusting and would stay in its last position without being able to move, giving different drivers steering troubles.
If your potential purchase was made during this period, be sure to check its service history report to see if the busted part was removed.
The Electric Windows Will Not Shut
This issue will hinder your comfort more than your safety. The electric windows in BMW models, including in BMW X5 cars, frequently break down, causing them to stay open.
We recommend having this issue fixed right away, as BMW cars are among the top stolen vehicles in the world.
Wear and Tear on the X5’s Door Handles
Many BMW X5 luxury SUV owners have reported issues with the vehicle’s door handles. It includes the door handles on both the driver and passenger doors lifting up as normal, but the door will not budge open. This can be more than a minor nuisance if you’re trying to get somewhere in a hurry.
Sadly, this isn’t a quick fix. The entire door opening and locking system will need to be replaced. It requires the use of specialty tools that you can only find in auto shops, so don’t try and do this repair by yourself.
Issues with the Electronics
The problem with windows not closing isn’t the only electronic issue found on BMW X5 vehicles. Numerous BMW owners have reported that the car’s fuse box will sometimes stop working.
More serious issues with BMW cars’ electrical system have been reported, leading them to recall over 300,000 vehicles in the UK during 2018. The cars simply cut out, leading to the death of one BMW driver in Hampshire, England.
Corrosion of Alloy Wheels
The BMW vehicle’s branded alloy wheels are one of the most stunning aspects of this car, and they certainly will make any BMW X5 stand out from the crowd of other SUVs. Sadly, some BMW cars have suffered from corroding wheels over a very short amount of time. This corroding does much more than make your wheels look ugly. It can also affect your tires and lead to safety issues. If you have experienced this issue, opt for a non-branded yet more reliable set of wheels to prevent any future issues.
Along with their electronic issues, BMW X5 luxury SUV drivers may notice that their rides suffer more than their fair share of dead batteries. While the battery is not a permanent part of your BMW and may need a change after a while, batteries should be able to last you between two and five years.
Some signs of a dead or dying battery can include:
- A slow cranking engine: If your BMW X5 is taking longer to start or is sluggish, it could be a sign of a bad battery.
- Dim headlights: If your headlights are operating as dim when your accelerator pedal is at full, your battery is probably on its deathbed. Headlights need a fully charged and properly functioning battery to illuminate properly in the dark, and driving without light can put you, and others, in danger.
- Your “check engine” light is on: There is a number of reasons why this dashboard light is on. A dead battery could be one of them. You’ll need a voltmeter to check your car battery’s voltage.
- Clicking noises: Your BMW’s starter needs the battery’s current to start the engine. When the current is low, the starter does not receive enough power to turn over the engine, so you will probably notice a clicking sound under the hood when this happens.
- Backfiring: A bad batter can cause sporadic sparks in your car’s combustion chamber. But before you replace your battery, ensure that you have ruled out other issues.
- Lack of interior lighting: Sure, BMW cars have ambient interior lighting, but they shouldn’t be dark. If you notice things getting darker inside of your car, it could be a symptom of a bad battery.
If you think you’re having battery troubles, take your BMW X5 into a service station so that they can replace it if necessary.
Faulty Automatic Headlights
Automatic headlights are a new innovation to hit the auto industry, and take the human component out of deciding when it is dark enough to flip your headlights on. The issue with some BMW X5 SUVs is that the automatic headlights will stay on, even when they aren’t needed. This can take a hefty toll on your car’s battery and electrical system.
Brakes are one of the most critical elements in your BMW X5, and should never be taken for granted. If you purchased a BMW X5 with a four-wheel drive system, be sure to have the brakes inspected regularly because this system can be punishing to the brake pads, discs, and calipers.
The Bottom Line on the BMW X5
The BMW X5 is a great luxury SUV that has your comfort and convenience top of mind. But even though it’s an innovative ride, it isn’t without its faults and quirks.
If you notice any of the aforementioned BMW X5 problems, be sure to take your set of wheels into a service center to get diagnosed and repaired right away.