<div>Background: Staphylococcus pseudintermedius is a part of the canine skin microflora and an opportunistic pathogen. It plays a central role in canine pyoderma, otitis and surgical wound infections. These conditions correlate with virulence genes distributed in the bacterial genome. These genes determine strain variability on typing, in turn aiding epidemiological surveillance. The aim of this study was to isolate, identify and characterize Staphylococcus pseudintermedius (SP) and Methicillin resistant Staphylococcus pseudintermedius (MRSP) from dogs with skin infections in Chennai, India. 
 
Methods: SP and MRSP positive isolates were identified by multiplex PCR for nuc and mecA genes respectively. Characterization of the isolates for virulence genes responsible for biofilm formation (icaA, icaD), cell wall adherence (SpsO, SpsK, SpsP, SpsQ, SpsF), toxins (ExpA, ExpB, SIET, Sel, Se-int, LukS, LukF) and gene regulation (Agr, SarA) was performed. 
 
Result: Out of 275 samples, 120 SP and 8 MRSP positive isolates were identified. Only one isolate could be typed as SCCmec Type V whereas other MRSP isolates were non typeable. Agr typing of MRSP isolates revealed type II in 7 isolates and type III in one isolate. Our study revealed that there was no significant difference in the detection of virulence genes between MSSP and MRSP.</div>

Characterization of Staphylococcus pseudintermedius Isolates from Skin Infections of Dogs

<div style="text-align: justify;">The attempt was made to study the response of the dog having skin infection with various underlying causes to killed S.pseudintermedius antigen. PBMC were separated and sensitized with killed S. pseudintermedius for 24 hours. cDNA was made from extracted RNA and quantitative PCR were carried out for 8 cytokines with GAPDH as housekeeping gene. IL-6 was the cytokine which was expressed in a statistically significant high level in PBMC of healthy animals to killed antigen than PBMC of infected animals. Except for the IL-6, all other cytokines were expressed at high level in pyoderma animals than healthy control, demodicosis and allergy cases. In the present study, IL-1&acirc;, IL-8 and TNF-&aacute; were the cytokines that were up-regulated and, IL-6 and IFN-&atilde; were down-regulated in demodicosis dogs with pyoderma at apparently significant level than the other tested cytokines. IL-4 was the only cytokine that was expressed in measurable quantity in allergic cases when compared to other cytokines. Thus, it was concluded that dogs with staphylococcal pyoderma infections developed a Th1/Th2 response to fight the infection.&nbsp;</div>

Cytokine response to killed Staphylococcus pseudintermedius antigen in dogs with skin diseases

A bacterium was isolated from benzalkonium chloride solution (5%) stored in an uncovered bottle. The strain was identified as Staphylococcus aureas named SA-RBC through the test of its morphology, biochemical and physiological characterization. The isolated strain SA-RBC has important symbol of pathogenic Staphylococcus by producing coagulase, phosphatase and thermonuclease. Resistance of SA-RBC to disinfectants or antibiotics was tested. The results showed that the strain was highly resistant to benzalkonium chloride, benzalkonium bromide, glutaraldehyde and sodium hypochlorite, but susceptive to phenylic acid. SA-RBC was highly resistant to oxacillin, cefoperazone, cefoxitin, gentamicin, ciprofloxacin, and tetracycline. The tolerable growth concentration of SA-RBC on various surfactants was examined. The results indicated that the strain SA-RBC could grow in a higher concentration of cationic surfactants up to 5%, compared with the control strains (Escherichia coli ATCC 25922, S.epidermidis ATCC12228 and S.aureus ATCC 29213) which were unable to grow in 0.1% cationic surfactants. The strain SA-RBC could not utilize benzalkonium chloride as the sole source of carbon and/or nitrogen. The findings in this study suggested that Staphylococcus aureas might be an original contaminant in the quaternary ammonium disinfectant and has potential risk for public health.

Isolation and identification of a novel Staphylococcus from benzalkonium chloride solution

<div>Background: The present study was done to ascertain prevalence of Staphylococcus aureus from various canine affections in Banaskantha district of Gujarat, India. Along with this, use of classical and molecular techniques were compared in identification and virulence characterization of this pathogen. 
 
Methods: A total of 165 samples were collected and bacterial identification was carried out with bacteriological (phenotypic) techniques and confirmed by genus specific 16S rDNA and Staphylococcus aureus specific sa442 gene based PCR. Isolates were characterized for coagulase production, haemolysis activity and presence of spa gene. &nbsp; 
 
Result: Samples yielded, 88 (53.33%) Staphylococcus spp. via bacteriological and PCR methods. Clinically, 19 (21.59%), 28 (31.82%), 12 (13.64%), 15 (17.04%) and 14 (15.91%) isolates were from abscess/wound, pyoderma, respiratory problems, eye infections and Otitis, respectively. A total of 46/88 (52.27%) isolates were confirmed as Staphylococcus aureus in PCR. In tube coagulase test, 51/88 (57.95%) isolates were found positive. A tota of 42 isolates revealed presence of coa gene, including two tube coagulase negative isolates. Haemolytic activity revealed beta (51.14%) gamma (31.82%), alpha (13.64%) and alpha-beta (3.41%) haemolysis, respectively. X-region of Protein A (spa gene) was detected in 26 /46 (56.52%) isolates in PCR.</div>

<div>Bacteriological and Molecular Identification and Characterization of Staphylococcus aureus from Different Affections of Canines</div>

<div style="text-align: justify;">Present study was undertaken to characterize Staphylococcus spp. associated with respiratory tract infection in small ruminants. A total of 139 bacterial isolates were recovered from clinically sick and apparently healthy small ruminants. The prevalence rate of Staphylococcus spp. in sick and healthy animals was 44.94 and 24% respectively. The sensitivity pattern of selected antibiotics was similar in both clinically sick and apparently healthy animals except streptomycin and penicillin-G. &nbsp;PCR assay was standardized for the detection of pathogenic genes of Staphylococcus spp. viz. clfA and spa. Among 34 random isolates selected, the prevalence rate of clfA and spa gene &nbsp;was 79.41 and 58.82% respectively indicating their pathogenic potential. In conclusion, Staphylococcus spp. was found to be one of the highly prevalent organism in respiratory tract infections.</div>

Molecular characterization of Staphylococcus spp. isolated from respiratory tract of apparently healthy and clinically sick sheep and goat in Nagpur, India

<div>Background: Pyoderma in dog poses complexity owing to involvement of multi drug resistant bacterial infection particularly Staphylococcus species. This may result into failure of antibiotic therapy in pyoderma. Therapeutic use of lytic bacteriophage (phage) may be an alternative to deal antimicrobial resistance in veterinary medicine. There is no report available on isolation and characterization of bacteriophage from clinical cases of pyoderma in animals, hence the study was performed with aim to isolate and characterize lytic phage against methicillin resistant S. aureus (MRSA).&nbsp; 
 
Methods: The phage was isolated from pus sample of dog suffering from severe pyoderma. The phage was isolated using MRSA as a bacterial host and was named as Staphylococcus phage BVC1 (SPBVC1). The morphology of the phage was determined using TME and was characterised by determining the host range and lytic potential of the phage at range of temperature and pH. 
 
Result: The morphology of phage revealed an icosahedral head of diameter 81.31 nm. with sheath and a central tube and a tail of 92.08 nm. It showed strong lytic activity against Methicillin resistant S. aureus and was stable under a range of temperature varying from 4oC to 45oC and pH from 4 to 11. The phage has shown lytic activity against the MRSA however no lytic activity against the MSSA was shown by the phage. The high specificity of the phage for MRSA indicated its potential use as an alternative therapeutic approach against multidrug resistant staphylococcal infections.</div>

Isolation and Characterization of Lytic Bacteriophage against Methicillin Resistant Staphylococcus aureus from Pyoderma in Dog

<div>Background: A study was carried out to screen milk borne Staphylococcus aureus for resistance against Beta lactam antibiotics.&nbsp; 
 
Methods: A total of 45 milk samples were collected over a period of three months from large animal outpatient unit of Madras Veterinary College Hospital, Chennai. Upon collection of samples, ABST followed by its growth in Mannitol Salt Agar was carried out as part of the phenotypic screening. Genotypic screening for Staphylococcus screening was done with the help of PCR by using nuc &nbsp;and mec A primers. MIC for ceftriaxone and cloxacillin was carried out with the samples that were found positive for Staphylococcus aureus. The antibiotic sensitivity pattern is presented: Fluoroquinolones (87.5% sensitive), aminoglycosides (72.5% sensitive), Amoxicillin-Clavulanic acid (Amoxyclave) (72.5% sensitive). The MSA positive samples were subjected to molecular identification with the help of PCR.&nbsp; 
 
Result: The results revealed 10 samples positive for Staphylococcus aureus and 5 among them positive for mecA gene. The MIC results were as follows: MIC50-10.95&micro;g/ml and MIC90- 87.510.95&micro;g/ml for ceftriaxone and MIC50- 43.75 &micro;g/ml and MIC90- 87.5&micro;g/ml for cloxacillin, indicating emergence of resistance. However, further studies are required in a larger sample size that can help us to attain more conclusive results.</div>

<div>Screening of Milk Borne Staphylococcus aureus for Resistance against Beta Lactam Antibiotics</div>

<div style="text-align:justify">Background: India leads the global market in milk production. However, bovine mastitis, which is the mammary gland inflammation in dairy cattle characterized by physical, chemical, bacteriological changes in milk results in commercial losses. Staphylococcus aureus, is the major causative agent. The treatment of mastitis caused by this pathogen is mainly by antibiotics. Emphasizing on the one health concept, phage therapy is an appropriate alternative to antibiotic. The present study was aimed to isolate and characterize bacteriophages against Staphylococcus aureus associated with bovine mastitis. 
 
Methods: Thirty two isolates of S. aureus obtained from the milk of mastitis cows were characterized by phenotypic and genotypic methods. Antibiotic susceptibility of the isolates was carried out by disk diffusion assay. Milk and cow shed wastewater were used for phage isolation. Phages were characterized by host susceptibility and RAPD assay. 
 
Result: Nineteen phages were isolated from the cowshed waste water. All the milk samples showed negative for the presence of phage. The phage 24 (A2) which had the broadest host range, was selected for the CFU drop assay. The phage was able to clear the lawn of S. aureus culture when grown on agar at different time points thus indicating that topical application of this phage would be a potential strategy to control S. aureus infection leading to mastitis. This study provides a basis to continue the exploration of the potential of PSA2 phage as a candidate for the treatment of Staphylococcal mastitis.</div>

<div style="text-align:justify">Isolation of Phages and Study of their In vitro Efficacy on Staphylococcus aureus Isolates Originating from Bovine Subclinical Mastitis</div>

<div>Background: Mitigation process to curb the ever increasing problem of antimicrobial resistance through development of new class of antimicrobials is slow and costly affairs. Research on alternative to conventional antimicrobials using plant based products as good source of numerous phytochemicals have potential to cope up the antimicrobial resistance. The present study was formulated on detection of in vitro antimicrobial and antibiofilm properties of methanol leaf extract of Citrus maxima against clinical isolates of Staphylococcus aureus. 
 
Methods: Leaves of Citrus maxima plants were collected from the campus of College of Veterinary Sciences and Animal Husbandry, Central Agricultural University, Aizawl, Mizoram and processed for preparation of methanol crude extract. The plant extracts were evaluated for their phytochemical and antioxidant properties using DPPH (2, 2-Diphenyl-1-picrylhydrazyl) method. Twenty well characterized biofilm producing and multidrug resistant Staphylococcus aureus strains recovered from milk of mastitic cows from Mizoram were received from the cultural repository of the department. The plant extracts were subjected to determine their antimicrobial and antibiofilm activities against all the bacterial isolates including&nbsp;S. aureus (ATCC 29213) by in vitro agar well diffusion method and 96 well microtiter plate methods, respectively. The MIC value of the plant extracts were determined by microdilution method. 
 
Result: In the methanol leaf extract of C. maxima alkaloids, glycosides, terpenoids, tannin and phenol and flavonoids were detected by qualitative analysis. Saponin, protein, free amino acids, steroids and carbohydrates were not detected. The free radical scavenging potential of the extract was found to be 10.66&plusmn;1.84% to 36.10&plusmn;1.98%, which was comparatively lower than ascorbic acid (83.39&plusmn;0.13% to 89.76&plusmn;0.24%). A total of 8 (40.0%), 5 (25.0%) and 7 (35.0%) strains were recorded as weak, moderate and strong biofilm producer. Maximum antibacterial activity against standard culture was observed with the zone of inhibition of 18 mm at 200 mg/mL concentration and MIC value at 25 mg/mL. Maximum antimicrobial activities against clinical isolates were recorded with 11.8&plusmn;1.13 mm zone of inhibition at 200 mg/mL and MIC value at 25 mg/mL. The clinical isolates exhibited highest (85.94&plusmn;1.00%) biofilm inhibition at 6.25 mg/mL. To the best of our knowledge, this is the first-ever report on antibiofilm and antioxidative activities of C. maxima leaf extracts against any bacteria.</div>

Antimicrobial and Antibiofilm Activities of Methanol Leaf Extract of Citrus maxima against Clinical Isolates of Multidrug Resistant Staphylococcus aureus

<div>Background: Antimicrobial resistance is one of the latest challenges facing the scientific community. Raising the drug resistance is caused mainly by indiscriminate usage of antibiotics in human and animal subjects and the spread of antibiotic resistance between the two has an emerging global threat. Hence, current study aimed to study the antimicrobial resistance pattern and molecular detection of antibiotic resistance genes in Staphylococcus aureus and Escherichia coli isolated from mastitis affected cows.&nbsp; 
 
Methods: Milk samples from mastitis affected cows were subjected to antibiotic sensitivity test and screened for presence of Staphylococcus aureus and Escherichia coli using differential growth media. Molecular characterization of Staphylococcus aureus and Escherichia coli was done with the help of PCR by amplification of &lsquo;nuc&rsquo; and &lsquo;uspA&rsquo; gene respectively. MICs of Penicillin and Tetracycline were determined using microdilution method.&nbsp; 
 
Result: Antibiotic sensitivity pattern for Penicillin G, Ampicillin, Amoxycillin, Cefotaxime, Ceftriaxone, Azithromycin, Ciprofloxacin, Gentamicin, Oxytetracycline, Tetracycline and Vancomycin were 74.19%, 100%, 93.50%, 61.29%, 29%, 35.48%, 9.70%, 9.70%, 70.96% and 70.96% &nbsp;respectively. More than 87.90% of the S. aureus and 50% of the E. coli isolated were resistant to &acirc;-lactam antibiotics while 75% of the E.coli and 65.70% of the S. aureus isolated were resistant to Tetracycline antibiotics. The MICs of Penicillin for S.aureus and E.coli are 26.88 &micro;g/ml and 13.54 &micro;g/ml respectively and the MICs of Tetracycline for S. aureus and E. coli are 243.75 &micro;g/ml and 960.93 &micro;g/ml respectively which is 8-9 folds higher than the standard MICs. From the present study, it can be inferred that bovine mastitis cases are highly resistant to antimicrobial drugs. Results further indicate that Staphylococcus aureus and Escherichia coli are both resistant to Penicillin and Tetracycline with very high MIC. &nbsp;</div>

<div>Molecular Characterization of Antibiotic Resistance Gene Pattern of Staphylococcus aureus and Escherichia coli in Mastitis Affected Dairy Cows</div>