Staphylococcus aureus counters organic acid anion-mediated inhibition of peptidoglycan cross-linking through robust alanine racemase activity.

Weak organic acids are commonly found in host niches colonized by bacteria, and they can inhibit bacterial growth as the environment becomes acidic. This inhibition is often attributed to the toxicity resulting from the accumulation of high concentrations of organic anions in the cytosol, which disrupts cellular homeostasis. However, the precise cellular targets that organic anions poison and the mechanisms used to counter organic anion intoxication in bacteria have not been elucidated. Here, we utilize acetic acid, a weak organic acid abundantly found in the gut to investigate its impact on the growth of Staphylococcus aureus. We demonstrate that acetate anions bind to and inhibit d-alanyl-d-alanine ligase (Ddl) activity in S. aureus. Ddl inhibition reduces intracellular d-alanyl-d-alanine (d-Ala-d-Ala) levels, compromising staphylococcal peptidoglycan cross-linking and cell wall integrity. To overcome the effects of acetate-mediated Ddl inhibition, S. aureus maintains a high intracellular d-Ala pool through alanine racemase (Alr1) activity and additionally limits the flux of d-Ala to d-glutamate by controlling d-alanine aminotransferase (Dat) activity. Surprisingly, the modus operandi of acetate intoxication in S. aureus is common to multiple biologically relevant weak organic acids indicating that Ddl is a conserved target of small organic anions. These findings suggest that S. aureus may have evolved to maintain high intracellular d-Ala concentrations, partly to counter organic anion intoxication.

A critical role for staphylococcal nitric oxide synthase in controlling flavohemoglobin toxicity.

Most coagulase-negative staphylococcal species, including the opportunistic pathogen Staphylococcus epidermidis, struggle to maintain redox homeostasis and grow under nitrosative stress. Under these conditions, growth can only resume once nitric oxide (NO) is detoxified by the flavohemoglobin Hmp. Paradoxically, S. epidermidis produces endogenous NO through its genetically encoded nitric oxide synthase (seNOS) and heavily relies on its activity for growth. In this study, we investigate the basis of the growth advantage attributed to seNOS activity. Our findings reveal that seNOS supports growth by countering Hmp toxicity. S. epidermidis relies on Hmp activity for its survival in the host under NO stress. However, in the absence of nitrosative stress, Hmp generates significant amounts of the harmful superoxide radical (O2•-) from its heme prosthetic group which impedes growth. To limit Hmp toxicity, nitrite (NO2-) derived from seNOS promotes CymR-CysK regulatory complex activity, which typically regulates cysteine metabolism, but we now demonstrate to also repress hmp transcription. These findings reveal a critical mechanism through which the bacterial NOS-Hmp axis drives staphylococcal fitness.

Reproducibility of Cytomorphological Diagnosis and Assessment of Risk of Malignancy of Thyroid Nodules Based on the Bethesda System for Reporting Thyroid Cytopathology: A Tertiary Cancer Center Perspective.

Introduction: The Bethesda System for Reporting Thyroid Cytopathology (TBSRTC) was introduced for unifying the terminology and morphologic criteria along with the corresponding risk of malignancy, leading to more consistent management approaches. The aim of this study was to study the utility and reproducibility of TBSRTC in reporting thyroid cytology in a referral cancer center. Methods: The fine-needle aspiration (FNA) of all thyroid nodules were included for a period of 5 years, from January 2016 to December 2021, in this cancer center. They were retrospectively reviewed and recategorized according to TBSRTC by two experienced pathologists. Cytohistopathological correlation was done for the cases which underwent surgical resection. Results: 522 fine-needle aspiration cytology (FNAC) of thyroid swellings were evaluated and categorized according to TBSRTC. There was agreement in the cytological diagnosis of 512 cases, of which 260 (50.78%) were benign lesions, 189 (36.91%) were malignant, 5 (0.97%) were unsatisfactory/nondiagnostic, 41 (8.01%) were follicular neoplasm/suspicious for neoplasm, 13 (2.53%) were suspicious for malignancy, and 4 (0.78%) cases were reported as atypia of undetermined significance. Two cytopathologists were in agreement in 512 cases (98%) of cases. Almost complete concordance was noted in the malignant (99%) and benign categories (98%). Disagreement was seen in 10 cases. Histological follow-up was available in 201 cases with an overall malignancy rate of 62.68% (126/201). Conclusion: TBSRTC proved to be a very simple and effective reporting system for thyroid FNAC, especially in the setting of a cancer center. This enables proper triaging of cases with thyroid masses into those who require surgical intervention and those who can avoid it, thereby preventing unnecessary morbidity.

A Plausible Proposition of CCL20-Related Mechanism in Fusobacterium nucleatum-Associated Oral Carcinogenesis.

OBJECTIVE: The objective of this prospective observational case-control study is to evaluate the prevalence of Fusobacterium nucleatum in the tissues of oral squamous cell carcinoma (OSCC). Reconnoitering the CCL20-related mechanism of carcinogenesis in Fusobacterium nucleatum-positive OSCC is another objective. METHODOLOGY: Tissues from 50 OSCC patients and 30 healthy oral tissues were collected. The prevalence of Fusobacterium nucleatum was evaluated in both tumour and healthy tissue by polymerase chain reaction. The immunohistochemistry of OSCC tissues was conducted to evaluate the difference in the expression of CCL20 between Fusobacterium nucleatum-positive and -negative OSCC tissues. RESULTS: Fusobacterium nucleatum was significantly (p < 0.001) prevalent in OSCC tissues (74%), compared to healthy tissues (26%). No association of Fusobacterium nucleatum or CCL20 immuno-expression with any clinical or histopathological features of OSCC was observed. While the intensity of CCL20 immuno-expression did not differ (p = 0.053), the CCL20-positive cell population was significantly different (p = 0.034) between Fusobacterium nucleatum-positive and -negative OSCC. CONCLUSION: Fusobacterium nucleatum is possibly prevalent in oral cancer tissues in the Indian population. By using immunohistochemistry, this is the first study to propose that the carcinogenesis in Fusobacterium nucleatum-positive OSCC may be CCL20-related. The findings enrich the knowledge of mechanisms involved in Fusobacterium nucleatum-mediated oral carcinogenesis.

Comparison of dosimetric data of bone marrow between standard IMRT and bone marrow sparing IMRT in carcinoma cervix.

BACKGROUND: The aim of the study was to assess the dosimetric comparison of bone marrow between standard IMRT(SD-IMRT) and bone marrow sparing IMRT (BMS-IMRT) among carcinoma cervix patients who underwent radical or adjuvant chemoradiation in a tertiary cancer center. MATERIALS AND METHODS: Forty eligible patients of histo-pathologically proven carcinoma cervix were enrolled in the study that was randomized on a 1:1 basis between SD-IMRT and BMS-IMRT from July 2018 to October 2019. The whole pelvis, bilateral femoral heads, and upper 1/3rd femur were contoured using the whole bone technique as a surrogate marker for the bone marrow. In both arms, V10, V20, and V40, bone marrow was noted along with mean, maximum, minimum dose, and total volume. DVH for the bone marrow in both arms was compared using the unpaired student t-test. RESULTS: We found no significant difference in the mean of various parameters in SD-IMRT arm vs. BMS IMRT arm - for the bone marrow: V10 (89 ± 4.3% vs. 86.7 ± 3.7%), V20 (73.2 ± 5.3% vs. 73.1 ± 4.5%), V40 (23.9 ± 5.4% vs. 26.6 ± 7.4%) and, similarly, for mean dose (28.1 ± 3.5% vs. 28.1 ± 1.8%), maximum dose (53.4 ± 0.58% vs. 53.2 ± 0.58%), minimum dose (0.33 ± 0.18% vs. 0.38 ± 0.38%), total volume (961 ± 110 cc vs. 901 ± 152 cc). CONCLUSION: This study shows no statistically significant difference in dosimetry between the two groups, which suggests that SD-IMRT spares the bone marrow adequately. Therefore, the need for BMS-IMRT using the present contouring technique does not give any added advantage over SD-IMRT. However, large sample size, other novel contouring technique, and multivariate analysis are needed to reach a definite conclusion.

Antibiotic Resistance Profile, Outer Membrane Proteins, Virulence Factors and Genome Sequence Analysis Reveal Clinical Isolates of Enterobacter Are Potential Pathogens Compared to Environmental Isolates.

Outer membrane proteins (OMPs) of gram-negative bacteria play an important role in mediating antibacterial resistance, bacterial virulence and thus affect pathogenic ability of the bacteria. Over the years, prevalence of environmental antibiotic resistant organisms, their transmission to clinics and ability to transfer resistance genes, have been studied extensively. Nevertheless, how successful environmental bacteria can be in establishing as pathogenic bacteria under clinical setting, is less addressed. In the present study, we utilized an integrated approach of investigating the antibiotic resistance profile, presence of outer membrane proteins and virulence factors to understand extent of threat posed due to multidrug resistant environmental Enterobacter isolates. Also, we investigated clinical Enterobacter isolates and compared the results thereof. Results of the study showed that multidrug resistant environmental Enterobacter isolates lacked OmpC, lacked cell invasion abilities and exhibited low reactive oxygen species (ROS) production in neutrophils. In contrast, clinical isolates possessed OmpF, exhibited high invasive and adhesive property and produced higher amounts of ROS in neutrophils. These attributes indicated limited pathogenic potential of environmental Enterobacter isolates. Informations obtained from whole genome sequence of two representative bacterial isolates from environment (DL4.3) and clinical sources (EspIMS6) corroborated well with the observed results. Findings of the present study are significant as it highlights limited fitness of multidrug resistant environmental Enterobacter isolates.

Antibiotic Susceptibility, Virulence Pattern, and Typing of Staphylococcus aureus Strains Isolated From Variety of Infections in India.

Staphylococcus aureus is one of the major causes of nosocomial infections. This organism produces powerful toxins and cause superficial lesions, systemic infections, and several toxemic syndromes. A total of 109 S. aureus strains isolated from a variety of infections like ocular diseases, wound infection, and sputum were included in the study. Minimum inhibitory concentration (MIC) was determined against 8 antimicrobials. PCR determined the presence of 16S rRNA, nuc, mecA, czrC, qacA/B, pvl, and toxin genes in S. aureus isolates. Pulse-field gel electrophoresis (PFGE), multi-locus sequence typing (MLST), SCCmec, spa-, and agr-typing and serotyping determined the diversity among them. All isolates of S. aureus were resistant to two or more than two antibiotics and generated 32 resistance patterns. These isolates were positive for 16S rRNA and S. aureus-specific nuc gene, but showed variable results for mecA, czrC, and qacA/B and pvl genes. Of the 32 methicillin-resistant S. aureus (MRSA), 13 strains carried SCCmec type V, seven type IV, two type III, and nine carried unreported type UT6. Of the 109 strains, 98.2% were positive for hlg, 94.5% for hla, 86.2% for sei, 73.3% for efb, 70.6% for cna, 30.2% for sea, and 12.8% for sec genes. Serotypes VII and VI were prevalent among S. aureus strains. PFGE analysis grouped the 109 strains into 77 clusters. MLST classified the strains into 33 sequence types (ST) and eight clonal complexes (CCs) of which 12 were singletons, and two belong to new allelic profiles. Isolates showed 46 spa-types that included two new spa-types designated as t14911 and t14912. MRSA and methicillin-susceptible S. aureus (MSSA) isolates were diverse in terms of antibiotic resistance pattern, toxin genotypes, SCCmec types, serotypes and PFGE, MLST, and spa-types. However, few isolates from eye infection and wound infection belong to CC239, ST239, and spa-type t037/t657. The study thus suggests that S. aureus strains are multidrug resistant, virulent, and diverse irrespective of sources and place of isolation. These findings necessitate the continuous surveillance of multidrug-resistant and virulent S. aureus and monitoring of the transmission of infection.

Differential expression of cyclin E, p63, and Ki-67 in gestational trophoblastic disease and its role in diagnosis and management: A prospective case-control study.

BACKGROUND: Gestational trophoblastic disease (GTD) constitutes a spectrum of tumors and tumor-like conditions, characterized by proliferation of pregnancy-associated trophoblastic tissue of progressive malignant potential. It is very difficult to differentiate these complex groups of lesions basing on histomorphology alone. Immunohistochemistry (IHC) with cyclin E, P63, and Ki-67 has a definite role in the identification of different trophoblasts and entities of GTD and also in the determination of biological behavior. AIMS: The aim of this study is to find the differential expression of cyclin E, p63, and Ki-67 in normal placenta, hydropic abortus (HA), and various entities of GTD. DESIGN AND SETTINGS: A prospective case-control study conducted in a government medical college. METHODS: Total 96 cases, divided into Group A (48 histologically confirmed cases of GTD) and Group B (controls comprising 8 HA and 40 normal placentas of different trimesters), were studied. The histological samples were subjected to IHC using cyclin E, Ki-67, and p63. STATISTICAL ANALYSIS: Results were analyzed using SPSS statistical method. RESULTS: Among the three immunomarkers used, Cyclin E and Ki-67 show statistically significant difference (P < 0.05) when compared between GTD and control groups, but it was insignificant for p63 (P = 0.369). Strong staining intensity of cyclin E and Ki-67 is seen in complete moles, choriocarcinoma, and placental site trophoblastic tumor. CONCLUSION: This study was done to evaluate the role of cell cycle regulatory proteins such as cyclin E and p63 and proliferation marker Ki-67 in the detection of various trophoblasts and differential diagnosis of the lesions associated with them.

Biofilm Formation by ica-Negative Ocular Isolates of Staphylococcus haemolyticus.

Staphylococcus haemolyticus is the second most frequently isolated CoNS from ocular infections and human blood cultures. In this study, we examined 18 ocular S. haemolyticus isolates for their capacity to form biofilm and conducted detachment assay to determine the composition of the biofilm matrix and involvement of various elements in cell lysis. PCR identified the presence of biofilm-associated genes, and ica operon and CLSM visualized the components of the biofilm matrix. We found that PIA-independent biofilm formation is the characteristic feature of S. haemolyticus isolates, irrespective of the sources of isolation, and protein or DNA or both are the major components of the biofilm matrix. Cell lysis enabling DNA release was an essential step for biofilm attachment during the initial stages of biofilm development. The srtA transcript expression study indicates its role in the early stages of biofilm development. We found the presence of antibiotic resistance genes in the eDNA and gDNA thus suggesting the possible role of biofilm in horizontal gene transfer of antibiotic resistance determinants. The overall study indicates that S. haemolyticus formed the biofilm comprising of protein or DNA or both and srtA play a role in the initial development of biofilm.

Clinicopathological study of malignant melanoma in a regional cancer center.

BACKGROUND: : Malignant melanoma is a tumor of melanocytic origin. Although uncommon in India as compared with the west, its prevalence is increasing. OBJECTIVES: To document the pattern of clinicopathological features of malignant melanoma cases attending in a regional cancer center in eastern India. MATERIAL AND METHODS: The present study was a retrospective study of 182 cases diagnosed histopathologically as malignant melanoma during 2011-2016. RESULTS: Out of the total cases, 170 (93.4%) were cutaneous and 12 (6.6%) were noncutaneous melanoma. The most common age group was sixth decade with a male predominance. Conventional melanotic melanomas were 176 (96.70%), and only 6 cases (3.30%) were amelanotic melanoma. Among noncutaneous melanomas, 6 were in anorectum, 2 in conjunctiva, and 1 case each in nasal cavity, palate, gingivo-buccal sulcus, and vagina. The acrallentigenous type was the most common variety, and the mixed epithelioid and spindle cell type was the most common histopathological pattern. Clark's level III was the most common level of invasion. CONCLUSION: The lower extremity is the most common site for melanoma, whereas extracutaneous melanomas are exceedingly rare and aggressive neoplasms. Melanoma can metastasize to regional lymph nodes, however, visceral metastasis to liver can also occur. In the absence of pigment in amelanotic melanoma, immunohistochemical markers such as HMB 45 can be used for definitive diagnosis.

Extrapulmonary Tuberculosis of Lymph Node Coexist with Diffuse Large B-cell Lymphoma: A Case Report.

Extrapulmonary tuberculosis (TB) coexistent with lymphomas in the same organ are rare. Here, we report a case of a 14-year-old male patient who had cauliflower-like ulcerated mass over cervical lymphadenopathy. He was diagnosed extrapulmonary TB. Unresponsiveness of anti tubercular treatment (anti-tubercular) treatment after 6 months, he was diagnosed as diffuse large B-cell lymphoma.

Serous Tubal Carcinogenesis: The Recent Concept of Origin of Ovarian, Primary Peritoneal and Fallopian Tube High-Grade Serous Carcinoma.

Background: Pelvic (non-uterine) high-grade serous carcinomas (PHGSC) including ovarian, tubal and primary peritoneal serous carcinomas have increased death: incidence ratio due to presentation at advanced stage, rapid progression, poor prognosis and high morbidity. Ambiguity regarding their pathogenesis and lack of a proper screening method is the cause of their late detection and high fatality rate. This study was undertaken to assess the fallopian tube for the presence of precursor lesions in pelvic serous carcinoma. Methods: This was a prospective case-control study carried out in a tertiary care center. Consecutive specimens of 55 cases of pelvic high-grade serous carcinoma and 41 controls inclusive of 21 low-grade serous carcinoma, 10 benign adnexal masses and 10 normal adnexa were included in the study. Both side fallopian tubes in each case were subjected to histopathological examination and p53, Ki67 immunohistochemistry. Results: There were 55 cases of PHGSC comprising of 50 cases of ovarian HGSC, two cases of primary peritoneal carcinoma (PPC) and three cases of tubal carcinoma. Serous tubal intraepithelial carcinoma (STIC) was detected in 14 cases (28%), p53 signature in 13 cases (26%) and tubal intraepithelial lesion in transition in 10 cases (20%) of ovarian HGSC. One case (50%) of PPC and one (33%) case of tubal carcinoma revealed the presence of STIC. None of the controls exhibited any precursor lesion except ovarian low-grade serous carcinoma where p53 was detected in 20% of cases. Conclusion: This revelation concludes that fallopian tubes are the sites of precursors of PHGSC to a large extent. In the absence of a proper screening method of HGSC, prophylactic bilateral salpingectomy at hysterectomy for benign diseases can achieve ultimate goal of reduction in incidence of PHGSC.

Identification of Major Sequence Types among Multidrug-Resistant Staphylococcus epidermidis Strains Isolated from Infected Eyes and Healthy Conjunctiva.

We examined the presence of virulence and antibiotic resistance genes, SCCmec types and determined the genomic diversity among ocular S. epidermidis isolates (patients-23, healthy controls-29). PCR determined the presence of antibiotic resistance genes, virulence genes and SCCmec types among all isolates. MLST and PFGE determined the genomic relatedness among them. All isolates of S. epidermidis showed resistance to at least one class of antibiotics of which 48 isolates were multidrug resistant and carried ARGs. Thirty-five isolates were methicillin resistant and carried mecA gene. Majority of the isolates were resistant to fluoroquinolones and showed mutation in gyrA, parC, and parE genes, however, few isolates showed additional novel mutations in parC gene. Of the MRSE strains, 17 strains carried SCCmec type IV, four type V, two type II, and two UT4. Seven strains carried novel combination of ccr complex and SCCmercury element, not reported earlier. All the S. epidermidis strains harbored icaA and icaD genes, 47 carried ACME operon, and 50 contained IS256. A noteworthy finding was the presence of ST179 among 43% of infected eye isolates an observation rarely reported among S. epidermidis. PFGE and MLST analysis showed genomic diversity among them. Statistical analysis suggests that few healthy conjunctiva isolates had characteristics similar to infected eye isolates. S. epidermidis strains carrying mecA gene are multidrug resistant, virulent and diverse irrespective of sources of isolation. IS256 cannot be used as marker to differentiate isolates of infected eye from healthy conjunctiva.

Role of fine-needle aspiration cytology in diagnosis of disseminated histoplasmosis in an immunocompetent patient: A case report.

Histoplasmosis is a mycotic infection caused by dimorphic fungus, Histoplasma capsulatum. The organisms are usually found within the cells (macrophages). This organism mostly affects lungs in immunocompetent individuals and disseminated forms are seen in immunocompromised cases. Here, we describe a case of disseminated histoplasmosis in an immunocompetent, 35-year-old female with lymphadenopathy diagnosed by fine-needle aspiration cytology and cell block.

Poorly Differentiated Sertoli-Leydig Cell Tumour of Ovary with Heterologous Elements.

Sertoli-Leydig Cell Tumour (SLCT) is included under sex-cord stromal tumour of testis or ovary. Ovarian pathology is an extremely rare entity constituting less than 0.5% of all ovarian neoplasms. Majority of the cases present at younger age group, i.e., 2nd and 3rd decade of life. The clinical presentation depends on either mass effect or excess hormone production. Virillization symptoms are the most common presentation and depend upon the quantity of androgen production. Most of the patients have a unilateral mass without extraovarian spread and present at stage-I. Treatment and prognosis depends on the degree of the differentiation and the stage of the disease. Here, we report a case of SLCT of the ovary in a young female. The patient initially presented with pain abdomen due to mass effect and was treated with conservative surgery followed by adjuvant chemotherapy due to the presence of poorly differentiated pathology with heterologous elements.

Effect of dietary supplementation of fermented fish silage on serum biochemical parameters of broiler Japanese quails (Coturnix coturnix japonica).

AIM: The objective of this study was to evaluate the effect of feeding fermented fish silage (FFS) on serum biochemical parameters of Japanese quails (Coturnix coturnix japonica). MATERIALS AND METHODS: A total of 192, 7-day-old broiler Japanese quail chicks of either sex, were randomly distributed into four dietary treatments with four replicates in each group having 12 chicks in each replicate pen. The dietary treatments were T1 - Control diet, T2 - Diet containing 5% FFS, T3 - Diet containing 10% FFS, and T4 - Diet containing 15% FFS. Group body weight and feed consumption were recorded at weekly intervals. Feed conversion ratio (FCR) was derived by dividing the feed consumed with the weekly body weight gain. At the end of the experiment, 8 birds from each treatment were selected randomly and sacrificed by cervical dislocation to study the carcass traits expressed as % pre-slaughter live weight. At 5 weeks of age, about 2 ml of blood was taken from the jugular vein of each selected bird, and serum samples were separated after centrifugation. Total protein, albumin/globulin (A/G) ratio, calcium, phosphorus, triglyceride, total cholesterol, high density lipoprotein-cholesterol (HDL-C), very low density lipoprotein-cholesterol (VLDL-C), low density lipoprotein-cholesterol (LDL-C), alanine aminotransferase (ALT), and aspartate aminotransferase (AST) were estimated in the serum. RESULT: The cumulative body weight gains from 1 to 5 weeks of age did not vary significantly between control and 5% FFS group. However, a linear decrease in body weight gain was observed by increasing the levels of FFS beyond 5% in the diet. The cumulative feed consumption was significantly higher in case of birds under control group during 1-5 weeks of age as compared to birds fed FFS based diet (5%, 10%, and 15%). No statistical difference in cumulative feed conversion ratio could be noticed during 1-5 weeks of age. The eviscerated yield decreased and giblet weight increased due to the dietary supplementation of FFS at 15% level. The breast meat yield decreased due to dietary supplementation of FFS at 10% and 15% level in the diet of broiler Japanese quails. The effect of FFS was found to be nonsignificant (p>0.05) with respect to serum total protein, globulin and A/G ratio under different treatments whereas significant difference observed in serum albumin concentration. Dietary supplementation of FFS at 10% and 15% level significantly increased the AST concentration in serum. There was no significant difference among the treatments regarding the parameters such as ALT, calcium, phosphorous, triglyceride, total cholesterol, HDL-C, VLDL-C, and LDL-C. CONCLUSION: The serum-biochemical parameters are influenced by the dietary supplementation of FFS in broiler Japanese quails.

Sex-related differences in hemato-biochemical indices of adult Vanaraja chickens during summer and winter seasons.

AIM: The objective of this study was to evaluate the changes in hemato-biochemical indices in male and female Vanaraja chickens under tropical environment during summer and winter season. MATERIALS AND METHODS: A total of 120 day-old sexed Vanaraja chicks were selected as experimental chickens and distributed equally in two groups having 60 female and 60 male chickens in each group, respectively. The experiment was continued for 8 weeks (56 days) and both male and female chickens were slaughtered by cervical dislocation method. All parameters were estimated at the end of the experiment in both seasons. RESULTS: Male had higher blood glucose, Ca and P level. Blood glucose level significantly (p<0.05) reduced in summer. Female had higher total protein, albumin, globulin, and albumin/globulin ratio. Alanine aminotransferase and aspartate aminotransferase enzyme concentration were significantly (p<0.05) higher in summer. Total erythrocyte count, total leukocyte count, hemoglobin (Hb), Hb/lymphocyte ratio, and packed cell volume were significantly (p<0.05) higher in males. Mean corpuscular volume and mean corpuscular Hb were significantly (p<0.05) higher in females. CONCLUSION: Sex of chickens had a significant (p<0.05) effect on different parameters whereas season had nonsignificant (p>0.05) effect in most of the observed parameters. Hence, Vanaraja chickens are adaptable to local tropical climate and can be reared efficiently as backyard poultry.

Identification of Novel Sequence Types among Staphylococcus haemolyticus Isolated from Variety of Infections in India.

The aim of this study was to determine sequence types of 34 S. haemolyticus strains isolated from a variety of infections between 2013 and 2016 in India by MLST. The MEGA5.2 software was used to align and compare the nucleotide sequences. The advanced cluster analysis was performed to define the clonal complexes. MLST analysis showed 24 new sequence types (ST) among S. haemolyticus isolates, irrespective of sources and place of isolation. The finding of this study allowed to set up an MLST database on the PubMLST.org website using BIGSdb software and made available at http://pubmlst.org/shaemolyticus/. The data of this study thus suggest that MLST can be used to study population structure and diversity among S. haemolyticus isolates.

Multidrug-resistant Staphylococcus haemolyticus isolates from infected eyes and healthy conjunctivae in India.

This study aimed to determine the presence of antibiotic resistance genes (ARGs), SCCmec elements and genetic relatedness among Staphylococcus haemolyticus isolated from patients with a variety of eye infections (n=11) and from healthy conjunctiva (n=7). Minimum inhibitory concentrations were determined for 14 antimicrobials according to BSAC guidelines. PCR was used to identify the presence of mecA, mecC, SCCmec type and ARGs. Sequencing was used to determine mutations in gyrA, gyrB, topoisomerase IVA and IVB genes. Genetic relatedness was determined by PFGE. Of the 18 isolates, 17 showed resistance to at least one antibiotic, but none showed resistance to vancomycin or rifampicin. Ten isolates were oxacillin-resistant and carried the mecA gene, eight of which belonged to SCCmec type V. The presence of non-mec SCC elements in two meticillin-susceptible isolates and untypeable SCC elements in meticillin-resistant isolates suggests the involvement of S. haemolyticus in the diversification of SCC elements. Sequence analysis revealed point mutations in gyrA (Ser-84→Leu) and topoisomerase IVA genes (Ser-80→Leu) in 13 isolates, and additional variation in the QRDR (Asp-84→Asn) of two isolates, showing good correlation between mutations in gyrA and topoisomerase IV genes and the level of resistance to fluoroquinolones. PFGE analysis showed distinct pulsotypes forming two major clusters, indicating the existence of diversity among isolates, irrespective of the source of isolation. This study suggests that S. haemolyticus isolates from infected eyes and healthy conjunctivae invariably carried ARGs and SCCmec elements and showed diversity in their genomic content, irrespective of the source of isolation.