Distinct Early Inflammatory Events during Ear Tissue Regeneration in Mice Selected for High Inflammation Bearing S<Emphasis Type="Italic">lc11a1 R</Emphasis> and <Emphasis Type="Italic">S</Emphasis> Alleles

High inflammatory AIRmax mice homozygous for Slc11a1 R and S alleles were produced. AIRmax ^SS mice showed faster ear tissue regeneration than AIRmax ^RR mice, suggesting that the S allele favored tissue restoration. Here, we investigated the gene expression profiles and the inflammatory reactions of AIRmax ^RR and AIRmax ^SS mice during the initial phase of ear tissue regeneration. We observed superior levels of analysis of wound myeloperoxidase and edema in AIRmax ^SS mice, although similar cell influx was verified in both lines. Of the genes, 794 were up- and 674 down-regulated in AIRmax ^RR , while 735 genes were found to be up- and 1616 down-regulated in AIRmax ^SS mice 48 h after punch. Both mouse lines showed significant over-represented genes related to cell proliferation; however AIRmax ^SS displayed up-regulation of inflammatory response genes. Quantitative PCR experiments showed higher expressions of Tgfb1, Dap12 and Trem1 genes in AIRmax ^SS mice. These results indicate that Slc11a1 gene modulated the early inflammatory events of ear tissue regeneration.     

<Emphasis Type="Italic">Ureaplasma parvum</Emphasis> causes hyperammonemia in a pharmacologically immunocompromised murine model

A relationship between hyperammonemia and Ureaplasma infection has been shown in lung transplant recipients. We have demonstrated that Ureaplasma urealyticum causes hyperammonemia in a novel immunocompromised murine model. Herein, we determined whether Ureaplasma parvum can do the same. Male C3H mice were given mycophenolate mofetil, tacrolimus, and prednisone for 7 days, and then challenged with U. parvum intratracheally (IT) and/or intraperitoneally (IP), while continuing immunosuppression over 6 days. Plasma ammonia concentrations were determined and compared using Wilcoxon rank-sum tests. Plasma ammonia concentrations of immunosuppressed mice challenged IT/IP with spent broth (median, 188 μmol/L; range, 102–340 μmol/L) were similar to those of normal (median, 226 μmol/L; range, 154–284 μmol/L, p?>?0.05), uninfected immunosuppressed (median, 231 μmol/L; range, 122–340 μmol/L, p?>?0.05), and U. parvum IT/IP challenged immunocompetent (median, 226 μmol/L; range, 130–330 μmol/L, p?>?0.05) mice. Immunosuppressed mice challenged with U. parvum IT/IP (median 343 μmol/L; range 136–1,000 μmol/L) or IP (median 307 μmol/L; range 132–692 μmol/L) had higher plasma ammonia concentrations than those challenged IT/IP with spent broth (p?<?0.001). U. parvum can cause hyperammonemia in pharmacologically immunocompromised mice.     

Evaluation of the antiplasmodial properties of selected plants in southern Ethiopia

Background

The majority of the Ethiopian population is at risk of malaria largely caused by Plasmodium falciparum. The resistance of the parasite to existing drugs is the main challenge in the control of the disease and thus new therapeutic drugs are required. In Ethiopia, people use different plant species to treat malaria. However, very few of them have so far been evaluated for their safety level and antimalarial activity. Thus, the aim of this study was to evaluate the safety and antimalarial activity of extracts of Ajuga integrifolia, Clerodendrum myricoides, Melia azedarach, Peponium vogelii and Premna schimperi, locally used by the Sidama people of Ethiopia to treat malaria.

Methods

The safety level of 80 % methanol extracts of the plants were evaluated using standard acute toxicity test procedure. The antiplasmodial activity of 80 % methanol extracts of the plants were assessed in vivo using Swiss albino mice against chloroquine sensitive rodent malaria parasite, Plasmodium berghei, using the standard 4-day suppressive test procedure at doses of 200,400 and 800 mg/kg/day. The 80 % methanol extract of Ajuga integrifolia that exhibited better antimalarial activity was fractionated using different solvents and screened for its phytochemical constituents and evaluated in vivo for its antimalarial activity at doses of 100, 200 and 400 mg/kg/day.

Results

All extracts given at the three different doses caused no lethal effect on mice in 24 h and within 10 days of observation. All extracts and fractions exhibited antimalarial activity in a dose dependant manner. The highest inhibition was exhibited by the crude extracts of A. integrifolia (35.17 %) at 800 mg/kg/day (P?<?0.05). Among fractions of A. integrifolia, n-butanol fraction demonstrated the highest inhibition (29.80 %) at 400 mg/kg/day (P?<?0.05). The extracts and fractions prolonged the survival time and prevented weight loss of the mice, but did not prevent PCV reduction. Phytochemical test on Ajuga integrifolia indicated the presence of alkaloids, flavonoids, saponins, terpenoids, anthraquinone, steroids, tannins, phenols and fatty acids.

Conclusions

Findings show that the plants are non-toxic and demonstrate antimalarial activity in a dose dependant manner suporting claims of their traditional therapeutic value for malaria treatment. However, further in-depth investigation is required to assess the potential of the plants towards the development of new antimalarial agent.

     

The importance of <Emphasis Type="Italic">Helicobacter pylori tnpA</Emphasis>, <Emphasis Type="Italic">tnpB</Emphasis>, and <Emphasis Type="Italic">cagA</Emphasis> genes in various gastrointestinal diseases

The cag (cytotoxin-associated gene) pathogenicity island (cagPAI) is one of the major virulence determinants of Helicobacter pylori (H. pylori). The purpose of this study was to investigate the association of the three genes (tnpA, tnpB, and cagA) in H. pylori isolated from Azerbaijani patients with the different gastrointestinal disease. A total of 362 gastric biopsies were collected from hospitals of Tabriz University of Medical Sciences, and were cultured on Brucella agar. The tnpA, tnpB, and cagA genes were detected by PCR. Of the total 264 H. pylori isolates, tnpA, tnpB, and cagA genes were detected in 120 (45.5%), 56 (21.2%) and 172 (65.2%), respectively. A significant association between tnpA and tnpB genes and clinical outcomes were found (P < 0.05). The cagA status was not related to clinical outcomes in our subjects. The predominant genotype among cag-PAI is the cagA. The prevalence of tnpA, tnpB, and cagA genes are high in patients with gastric cancer, and a significant association is revealed between tnpA and tnpB with gastric cancer.     

Characterisation of VIM-2-producing <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> isolates from lower tract respiratory infections in a Spanish hospital

To analyse the antimicrobial phenotype, carbapenem mechanisms, integrons, virulence factors and molecular typing of 164 Pseudomonas aeruginosa isolates recovered from lower tract respiratory samples in a Spanish hospital (1 year) as well as the patients’ clinical data. Susceptibility testing to 12 antipseudomonal agents was determined by microdilution and metallo-beta-lactamase (MBL) phenotype by double disc method. The oprD gene was studied by PCR, sequencing and comparison with P. aeruginosa PAO1 sequence. Detection and characterisation of MBLs, class 1, 2 and 3 integrons, and virulence genes were studied by PCR and sequencing. The prevalence of carbapenem-resistant P. aeruginosa (CRPA) was 26.8%. MBL phenotype was detected in 52.3% CRPA, and all of them were disseminated throughout the intensive care unit. Most of the MBL-carrying patients presented respiratory disease, mechanical ventilation, tracheostomy, bacteraemia, ≥?30 hospitalisation days and previous treatment with carbapenems and/or ≥?3 different antimicrobial families. The bla_VIM-2 gene was the unique MBL encoding gene and was detected inside class 1 integrons. The class 1 integrons detected in 39 strains (23.8%) were associated with aminoglycosides (aadB, aadA1, aadA6, aacA4, aac(3)-I) and carbapenems resistance genes (bla_VIM-2). The aac(3)-I?+?aadA1 and bla_VIM-2 arrangements were the most prevalent ones. Thirty-one different PFGE patterns and 4 STs (ST175, ST235, ST253, ST973) were detected among the 39 intI1-positive isolates, being ST235 the most frequent. CRPA showed a great variety of alterations in oprD gene. The exoU⁺/exoS^? genotype was detected in 82.6% of bla_VIM-2-producing strains (ST235) and the exoU^?/exoS⁺ in the remaining 17.4% (ST973).     

Genomic analysis reveals the presence of a class D beta-lactamase with broad substrate specificity in animal bite associated <Emphasis Type="Italic">Capnocytophaga</Emphasis> species

Capnocytophga canimorsus and Capnocytophga cynodegmi can be transmitted from cats and dogs to humans, and can cause a wide range of infections including wound infections, sepsis, or endocarditis. We and others recently discovered two new Capnocytophaga species, C. canis and C. stomatis, mainly associated with wound infections. The first-line treatment of animal bite related infections is penicillin, and in case of allergy, doxycycline and trimethoprim/sulfamethoxazole. However, there is a lack of antibiotic susceptibility patterns for animal bite associated Capnocytophaga species. Thus, we ?set out to study the antibiotic profiles against animal bite associated Capnocytophaga species isolated from wound and blood cultures after cat and dog bites and coupled the findings to whole genome sequencing data. A total of 24 strains were included in the study. Phenotypic analysis of antibiotic resistance was performed with E-tests. The web-based tool ‘Resfinder’ was used to identify resistance genes in the whole genome dataset. Two strains of C. cynodegmi and two strains of the recently discovered C. stomatis were resistant to penicillin (MIC?> 24 mg?/L) and cephalosporins (MIC?>?24 mg/?L), and three out of these strains also exhibited resistance to imipenem (MIC?=?32 mg/?L). Genomic analysis revealed that these strains carried a class D beta-lactamase gene, which has not previously been found in Capnocytophaga spp. A class D beta lactamase with broad substrate specificity was found in animal bite associated Capnocytophaga species, which could have important implications when treating wound infections after cat and dog bites. It also suggests that pet animal bacteria can harbour resistance genes with relevance for human infections.     

Genetic makeup of Shiga toxin-producing <Emphasis Type="BoldItalic">Escherichia coli</Emphasis> in relation to clinical symptoms and duration of shedding: a microarray analysis of isolates from Swedish children

Shiga toxin (Stx)-producing Escherichia coli (STECs) cause non-bloody diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome, and are the primary cause of acute renal failure in children worldwide. This study investigated the correlation of genetic makeup of STEC strains as revealed by DNA microarray to clinical symptoms and the duration of STEC shedding. All STEC isolated (n?=?96) from patients <10 years of age in Jönköping County, Sweden from 2003 to 2015 were included. Isolates were characterized by DNA microarray, including almost 280 genes. Clinical data were collected through a questionnaire and by reviewing medical records. Of the 96 virulence genes (including stx) in the microarray, 62 genes were present in at least one isolate. Statistically significant differences in prevalence were observed for 21 genes when comparing patients with bloody diarrhea (BD) and with non-bloody stool (18 of 21 associated with BD). Most genes encode toxins (e.g., stx2 alleles, astA, toxB), adhesion factors (i.e. espB_O157, tir, eae), or secretion factors (e.g., espA, espF, espJ, etpD, nleA, nleB, nleC, tccP). Seven genes were associated with prolonged stx shedding; the presence of three genes (lpfA, senB, and stx1) and the absence of four genes (espB_O157, espF, astA, and intI1). We found STEC genes that might predict severe disease outcome already at diagnosis. This can be used to develop diagnostic tools for risk assessment of disease outcome. Furthermore, genes associated with the duration of stx shedding were detected, enabling a possible better prediction of length of STEC carriage after infection.     

Characterization of a novel splicing mutation in <Emphasis Type="Italic">UNC13D</Emphasis> gene through amplicon sequencing: a case report on HLH

Background

Hemophagocytic lymphohistiocytosis (HLH) is a rare but fatal disease caused by uncontrolled proliferation of activated lymphocytes and macrophages. Six genes including SH2D1A, PRF1, UNC13D, STX11, STXBP2 and XIAP were reported as causative genes in most cases.

Case presentation

Here we report a novel splicing mutation in UNC13D gene, which was identified in an 18-year-old female. Patient was diagnosed as HLH base on HLH-2004 guidelines, no history of inherited diseases was revealed in this family, parents were healthy and non-consanguineous. Splenomegaly and hemophagocytosis in bone marrow were observed in clinical examination. Amplicon sequencing for the whole coding region of 6 HLH-related genes was performed on Ion S5XL genetic analyzer. In all, four heterozygous mutations were detected, including 2 nonpathogenic SNPs (PRF1:c.900C > T, STX11:c.*70G > A) and 2 splicing mutations in UNC13D gene (UNC13D:c.1299 + 1G > A and UNC13D:c.2709 + 1G > A), both of which were predicted to be potentially pathogenic by human splicing finder (HSF3) tool. The result was confirmed by two-generation pedigree analysis base on sanger sequencing.

Conclusions

Two compound heterozygous splicing mutations in UNC13D gene were identified and considered to be potential pathogenesis in a female patient of HLH. The mutation UNC13D:c.1299 + 1G > A was reported in HLH for the first time. The inheritance mode and source of the mutation in the proband was examined by family analysis. Our data suggest that further studies of the spectrum of HLH-related mutations in China are warranted.

     

Characterisation of <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> isolates from bloodstream infections,Democratic Republic of the Congo

Staphylococcus aureus is known worldwide as an invasive pathogen, but information on S. aureus from bloodstream infections in Central Africa remains scarce. A collection of S. aureus blood culture isolates recovered from hospitals in four provinces in the Democratic Republic of the Congo (2009–2013) was assessed. A total of 27/108 isolates were methicillin-resistant S. aureus (MRSA), of which >70% were co-resistant to aminoglycosides, tetracyclines, macrolides and lincosamides. For MRSA and methicillin-susceptible S. aureus (MSSA) isolates, resistance to chloramphenicol and trimethoprim–sulphamethoxazole (TMP-SMX) was <10%. However, 66.7% (72/108) of all isolates harboured the trimethoprim resistance gene dfrG. More than three-quarters (84/108, 77.8%) of isolates belonged to CC5, CC8, CC121 or CC152. Genetic diversity was higher among MSSA (31 spa types) compared to MRSA (four spa types). Most MRSA (23/27, 85.2%) belonged to CC8-spa t1476-SCCmec V and 17/23 (73.9%) MRSA ST8 were oxacillin susceptible but cefoxitin resistant. Among MRSA and MSSA combined, 49.1% (53/108) and 19.4% (21/108) contained the genes encoding for Panton–Valentine leucocidin (lukS-lukF PV, PVL) and toxic shock syndrome toxin-1 (tst, TSST-1), respectively. PVL was mainly detected among MSSA (51/53 isolates harbouring PVL were MSSA, 96.2%) and associated with CC121, CC152, CC1 and CC5. TSST-1 was associated with CC8-spa t1476-SCCmec V. The immune evasion cluster (IEC) genes scn, sak and chp were detected in 81.5% of isolates (88/108, equally represented among MSSA and MRSA). The present study confirms the occurrence of MRSA with high levels of multidrug co-resistance and PVL-positive MSSA among invasive S. aureus isolates in Central Africa.     

Matrix metalloproteinase-14 triggers an anti-inflammatory proteolytic cascade in endotoxemia

?

Matrix metalloproteinases can modulate the inflammatory response through processing of cyto- and chemokines. Among them, MMP-14 is a non-dispensable collagenase responsible for the activation of other enzymes, triggering a proteolytic cascade. To identify the role of MMP-14 during the pro-inflammatory response, wildtype and Mmp14 ^?/? mice were challenged with lipopolysaccharide. MMP-14 levels decreased after endotoxemia. Mutant animals showed 100% mortality, compared to 50% in wildtype mice. The increased mortality was related to a more severe lung injury, an impaired lung MMP-2 activation, and increased levels of the alarmin S100A9. There were no differences in the expression of other mediators including Il6, Cxcl2, Tgfb, Il10, or S100a8. A similar result was observed in lung explants of both genotypes cultured in presence of lipopolysaccharide. In this ex vivo model, exogenous activated MMP-2 ameliorated the observed increase in alarmins. Samples from septic patients showed a decrease in serum MMP-14 and activated MMP-2 compared to non-septic critically ill patients. These results demonstrate that the MMP-14-MMP-2 axis is downregulated during sepsis, leading to a proinflammatory response involving S100A9 and a more severe lung injury. This anti-inflammatory role of MMP-14 could have a therapeutic value in sepsis.

Key messages

? MMP-14 levels decrease in lungs from endotoxemic mice and serum from septic patients.? Mmp14 ^?/? mice show increased lung injury and mortality following endotoxemia.? Absence of Mmp14 decreases activated MMP-2 and increases S100A9 levels in lung tissue.? MMP-14 ameliorates inflammation by promoting S100A9 cleavage by activated MMP-2.

     

A novel compound heterozygous mutation in <Emphasis Type="Italic">VARS2</Emphasis> in a newborn with mitochondrial cardiomyopathy: a case report of a Chinese family

Background

Genetic defects in the mitochondrial aminoacyl-tRNA synthetase are important causes of mitochondrial disorders. VARS2 is one of the genes encoding aminoacyl-tRNA synthetases. Recently, an increasing number of pathogenic variants of VARS2 have been reported.

Case presentation

We report the novel compound heterozygous pathogenic VARS2 mutations c.643 C?>?T (p. His215Tyr) and c.1354 A?>?G (p. Met452Val) in a female infant who presented with poor sucking at birth, poor activity, hyporeflexia, hypertonia, persistent pulmonary hypertension of newborn (PPHN), metabolic acidosis, severe lactic acidosis, expansion and hypertrophic cardiomyopathy. These heterozygous mutations were carried individually by the proband’s parents and elder sister; the two mutations segregated in the family and were the cause of the disease in the proband.The c.643 C?>?T (p. His215Tyr) mutation was not described in the ExaC, GNomAD and 1000 Genomes Project databases, and the frequency of c.1354 A?>?G (p. Met452Val) was <?0.001 in these gene databases.The two mutated amino acids were located in a highly conserved region of the VARS2 protein that is important for its interaction with the cognate tRNA. The two missense mutations were predicted by online tools to be damaging and deleterious.

Conclusions

Our report expands the spectrum of known pathogenicVARS2 variants associated with mitochondrial disorders in humans.VARS2 deficiency may cause a severe neonatal presentation with structural cardiac abnormalities.

     

<Emphasis Type="Italic">Mycoplasma hominis</Emphasis> and <Emphasis Type="Italic">Gardnerella vaginalis</Emphasis> display a significant synergistic relationship in bacterial vaginosis

Gardnerella vaginalis plays an important role in bacterial vaginosis (BV,) while the role of genital Mollicutes is less obvious. The diagnosis of BV by use of the current Gram stain Nugent score is also suboptimal for defining the role of Mollicutes that lack a cell wall. Since bacterial load and diversity is an important prerequisite for BV, real-time quantitative polymerase chain reaction (qPCR) assays enable these to be assessed. The purpose of this study was to define the role of genital Mollicutes and potential patterns of synergy with G. vaginalis in women with BV. Vaginal swabs from 130 women categorised by Nugent score as BV (n?=?28), intermediate (n?=?22) and non-BV (n?=?80) were tested against four qPCR TaqMan assays targeting G. vaginalis, Mycoplasma hominis, M. genitalium, Ureaplasma urealyticum and U. parvum. Statistical analyses were used to compare bacterial prevalence and load between the three groups of women. Mycoplasma hominis and G. vaginalis co-infection was significantly more common in BV (60.7 %) compared to intermediate (36.4 %) and non-BV (8.8 %) Nugent scores (p?<?0.001). Significantly higher loads of M. hominis (p?=?0.001) and G. vaginalis (p?<?0.001) were detected in women with BV and the respective loads in M. hominis and G. vaginalis co-infections displayed a significant positive correlation (p?<?0.001; r?=?0.60). No significant associations were seen with the other Mollicutes. The findings strengthen the evidence of a role for M. hominis in BV and a potential synergy with G. vaginalis. This synergy could be an important trigger of the condition and sexual contact the conduit for the transmission of an otherwise commensal bacterium that could initiate it.     

Removal of spermatogonial stem cells (SSCs) from in vitro culture modulates testosterone synthesis in bovine

Niche cells, encompassing spermatogonial stem cells (SSCs) and controlling their development are believed to have a reciprocal communication with SSCs. The present study was conducted to evaluate the impact of the elimination of SSCs on testosterone production and genes contributing to testosterone synthesis. Following isolation, bovine testicular cells were cultured for 12 days on extracellular matrix-coated plates. In the germ cell-removed group, the SSCs were removed from the in vitro culture using differential plating; however, in the control group, no intervention in the culture was performed. Testosterone concentration and the gene expression of luteinizing hormone (LH) receptor (LHR), CYP17A1, and steroidogenic acute regulatory protein (StAR), were assessed on days 6 and 12. The concentration of testosterone was lower in the germ cell-removed than control group (P?<?0.05). Moreover, quantitative real-time PCR revealed lower gene expression of LHR, CYP17A1, and StAR in the germ cell-removed group as compared with the control group (P?<?0.05). In conclusion, the present study showed that elimination of SSCs from in vitro culture could diminish testosterone production via downregulation of factors contributing to testosterone synthesis. Additionally, this study provided evidence for the reciprocal relationship between SSCs and their niche cells.     

Improving effect of <Emphasis Type="Italic">Spirulina platensis</Emphasis> on hematological parameters in <Emphasis Type="Italic">Cyprinus carpio</Emphasis> exposed to sublethal doses of cyanide

The detoxification effects of dietary Spirulina platensis were investigated through semi-static chronic toxicity test developed with potassium cyanide (KCN) in common carp (Cyprinus carpio). The addition of dried Spirulina platensis in diet improves the hematological parameters (mean cell hemoglobin (MCH), hemoglobin (Hb), and mean cell hemoglobin concentrations (MCHC)) in spirulina + cyanide (SC) group in comparison with the fish exposed to cyanide alone (CY). RBC count increased in fish fed with spirulina (SP) in comparison with the other groups. However, this increase was significant compared with SC and CY (P?<?0.05). hematocrit (HCT), MCHC, and Hb level showed no significant difference among groups (P?>?0.05). MCH level improved significantly in the SC group compared with the SP group (P?<?0.05). Mean corpuscular volume (MCV) level had its highest level in the CY group compared with the SP group. WBC count increased significantly in the SC group compared to the CY group (P?<?0.05). To sum up, it seems that, 10 % S. platensis in diet provided some protections against the toxic action of KCN and increased the chance of blood regeneration.     

Hematological and biochemical reference intervals for <Emphasis Type="Italic">Bothrops asper</Emphasis> and <Emphasis Type="Italic">Crotalus simus</Emphasis> (Serpentes: Viperidae), maintained in captivity for venom extraction

This study presents hematological and biochemical reference intervals (RIs) for two species of viper snakes (Bothrops asper and Crotalus simus), which are the first of their type particularly for these two species maintained in captivity and for extraction of venom purposes. A total of 77 snakes were used in developing the study; specifically 39 B. asper and 38 C. simus snakes. Blood samples were obtained by puncturing the caudal vein, and hematological tests were performed manually by two independent technicians. Plasma biochemistry parameters were determined by an automatized analyzer. There are hematological differences between wild-caught and captive B. asper in the percentage of heterophils (P?<?0.001), while in captive and wild-caught C. simus, the differences are found in the percentage of eosinophils, lymphocytes, and monocytes (P?<?0.001). Differences in glucose (P?=?0.012), uric acid (P?<?0.001), and cholinesterase (P?=?0.004) are found in captive and wild-caught B. asper. Differences in albumin (P?=?0.002), calcium (P?<?0.001), CK (P?=?0.04), and LDH (P?=?0.037) are found between captive and wild-caught C. simus. This study constitutes the first set of hematological and biochemical RIs for B. asper and C. simus maintained in captivity with the purpose of producing antivenom in Costa Rica. The establishment of comprehensive RIs for hematology and plasma biochemistry for these two species of snakes is useful for interpretation of blood test results, moreover, to serving as a reference for viper snakes used for research purposes.     

Estimation of flavoniods,antimicrobial, antitumor and anticancer activity of <Emphasis Type="Italic">Carissa opaca</Emphasis> fruits

Background

Carissa opaca Stapf ex Hanes fruits is traditionally used in the treatment of asthma, hepatitis and microbial infections. The present study was arranged to investigate the antimicrobial, cytotoxic and antitumor activity of various fractions of C. opaca extract and its bioactive metabolites responsible for that activity.

Methods

To characterize various fractions of C.opaca antibacterial, antifungal, cytotoxic and antitumor assays are used. Eight strains of bacteria including Bacillus subtilis, Enterobactor aerogenes, Escherichia coli, Klebsiella pneumoniae, Micrococcus luteus, Pseudomonas aeroginosa, Salmonella typhy, and Staphylococcus aureus and four strains of fungal viz: Aspergillus flavus, Aspergillus fumigatus, Aspergillus niger and Fusarium solani are used. Brine shrimps and potato dics are used for anticancer and antitumor potency of extract. High performance liquid chromatography (HPLC) is utilized for determination of bioactive metabolites responsible for the activity.

Results

HPLC chromatogram revealed the presence of orientin, isoquercetin, myricetin and apigenin. Various fractions of C. oapca showed significant antibacterial, antitumor and anticancer activity. In case of C. opaca fruit inhibition growth of Aspergillus niger was ranged between 23.2?±?1.36% to 43.3?±?2.39%, Aspergillus flavus ranged between 27.6?±?1.39% to 65.6?±?3.44%, Aspergillus fumigatus ranged between 13.2?±?1.00% to 52.4?±?1.54% and Fusarium solani ranged between 10.5?±?1.02% to 14.6?±?1.74%.

Conclusion

It can be concluded that, various fractions of C.opaca are accessible source of ethno pharmacy as they are consumed in different areas of Pakistan with ultimate health compensations.

     

Diversity of <Emphasis Type="Italic">Cryptosporidium</Emphasis> species occurring in sheep and goat breeds reared in Poland

The aim of this study was molecular identification of Cryptosporidium species and assessment of their prevalence in different breeds of sheep and goat reared in Poland. In addition, the relationship between animal age, breed type, and the frequency of Cryptosporidium infections was determined. Fecal samples from 234 lambs and 105 goat kids aged up to 9 weeks, representing 24 breeds and their cross-breeds were collected from 71 small ruminant farms across Poland. The identification of Cryptosporidium species was performed at the 18 SSU ribosomal RNA (rRNA) and COWP loci followed by subtyping of C. parvum and C. hominis strains at GP60 gene locus. The presence of Cryptosporidium DNA at the 18 SSU rRNA locus was detected in 45/234 (19.2%) lamb feces samples and in 39/105 (37.1%) taken from goats. The following Cryptosporidium species: C. xiaoi, C. bovis, C. ubiquitum, C. parvum, and C. hominis were detected in small ruminants. Infections caused by C. xiaoi were predominant without favoring any tested animal species. Subsequent GP60 subtyping revealed the presence of C. parvum IIaA17G1R1 subtype in sheep and IIdA23G1 subtype in goats. IIdA23G1 subtype was detected in a goat host for the first time. There were no significant differences found in frequency of infections between the age groups (<3 and 3–9 weeks) of lambs (P = 0.14, α > 0.05) or goat kids (P = 0.06, α > 0.05). In addition, there was no correlation observed between the frequency in occurrence of particular parasite species and breed type in relation to native sheep breeds (F = 0.11; P = 0.990 > 0.05). In the case of goats, more breed-related differences in parasite occurrence were found. The results of this study improve our knowledge on the breed-related occurrence of Cryptosporidium infections in the population of small ruminants reared in Poland.     

Gasterophilosis in horses in Sardinia (Italy): effect of meteorological variables on adult egg-laying activity and presence of larvae in the digestive tract,and update of species

Gasterophilus larvae are common obligate parasites of the digestive tract of the equids. Horses become infected with this parasite by ingesting the larvae hatched from eggs laid by the female flies. In this study carried out monthly, we (i) counted the Gasterophilus eggs deposited by female flies on the coat of 30 grazing horses, (ii) counted and identified the Gasterophilus larvae retrieved from the digestive tract of 128 slaughtered horses, and (iii) compared these results to meteorological data. Eggs were deposited on all monitored horses, and were present from October to January and from May to September, whereas they were absent from February to April. The number of laid eggs was significantly different between the months, body regions, genders, and age classes (p?<?0.05). Larvae were recovered in 112 (87.5 %) horses, and 6 species of Gasterophilus were identified. The prevailing species were Gasterophilus intestinalis (recovered in 110 horses; 85.9 %) and Gasterophilus nasalis (69 horses; 53.9 %), recovered in all months. Gasterophilus inermis (5 horses; 3.9 %), Gasterophilus pecorum (3 horses; 2.3 %), Gasterophilus haemorrhoidalis (3 horses; 2.3 %)¸ and Gasterophilus meridionalis (2 horses; 1.6 %) larvae were also found. Significant differences were found among monthly larval burdens for both Gasterophilus spp. and G. intestinalis (p?<?0.05), but not for G. nasalis (p?>?0.05). Larval burdens and prevalences did not differed significantly between both genders and age classes (p?>?0.05). Monthly eggs and larvae trends were not significantly correlated (p?>?0.05). With regard to the meteorological variables, minimum air temperature was significantly correlated with the eggs trend (rho?=?1.000; p?<?0.001) and maximum air temperature with the Gasterophilus spp. (rho?=?0.972; p?<?0.001) and G. intestinalis (rho?=?0.972; p?<?0.001) larvae trends. In addition, the number of hours with a temperature below +10 °C was significantly correlated with G. intestinalis larvae trend (rho?=?0.602; p?<?0.05). Our findings confirmed that in Sardinia, Gasterophilosis is an important parasitosis in the horses, and it needs more attention and extensive and/or correct treatment to reduce its prevalence.