Development of antibodies against recombinant staphylococcal enterotoxin B from food poisoning cases.

Background and Aim: Staphylococcal enterotoxin B (SEB) is the most common serotype involved in food poisoning. The aim of this study was to develop immunoassay detection methods using a recombinant enterotoxin B antigen protein to produce recombinant polyclonal antibodies in vivo. Materials and Methods: Staphylococcus aureus isolated from a food poisoning case (strain JH5800) was analyzed by polymerase chain reaction (PCR) and confirmed to contain a seb gene of 477 bp. A SEB segment was amplified, cloned, sequenced, and aligned. The PCR product corresponding to the predicted mature SEB peptide was inserted into Escherichia coli BL21 (DE-3) expression vector and expressed as a hexahistidine-SEB fusion protein. Antiserum against recombinant SEB protein was produced by immunization of Balb/c mice. Results: In the indirect enzyme-linked immunosorbent assay (ELISA), the polyclonal antibodies produced had a titer of 1:3200. The seb gene of Staphylococcus aureus isolated from a poisoning case (JH5800) had a molecular size of about 477 bp and a band of recombinant SEB toxin was observed at approximately 30 kDa on SDS-PAGE gel. The polyclonal anti-SEB antibody titer, as revealed by indirect ELISA, was 1:3200 at 59 days. Conclusion: SEB recombinant protein could be used to produce polyclonal antibodies. ELISA and Western blotting were used to analyze the specificity and sensitivity of the recombinant polyclonal antibodies. Polyclonal antibodies produced could be used to detect SEB on a large-scale.

Methicillin-resistant Staphylococcus aureus isolates derived from humans and animals in Yogyakarta, Indonesia.

Background and Aim: The emergence of methicillin-resistant Staphylococcus aureus (MRSA) as a highly pathogenic strain in veterinary and human medicine is a growing global problem. This study aimed to evaluate MRSA isolates of human and animal origin against various antibiotics in Yogyakarta, Indonesia. Materials and Methods: The susceptibility test was carried out by the disk diffusion method using Mueller-Hinton agar against nine antibiotic disks. Methicillin-resistant S. aureus strains were genetically confirmed through mecA gene detection encoding for methicillin resistance by polymerase chain reaction. Results: All 240 S. aureus strains isolated from animals and humans were resistant to penicillin G (P) (100% and 99%, respectively), followed by ampicillin (AMP), amoxicillin (AML), oxacillin (OX), erythromycin (E), clindamycin (DA), tetracycline (TE), gentamicin (GEN), and ciprofloxacin (CIP). Eighty-three MRSA strains were resistant to OX (100%), P (100%), AMP (99.27%), AML (95.52%), E (87.77%), TE (71.33%), DA (63.24%), GEN (38.81%), and CIP (26.87%). Conclusion: The antimicrobial resistance pattern of S. aureus human isolates was similar to their animal counterpart, with 77.20% of MRSA strains classified as multidrug-resistant (MDR) bacteria. These findings indicate an increase in MDR S. aureus strains of animal origin in Yogyakarta, thus raising public health concerns about MRSA zoonotic spread.

Cellular immune response of Staphylococcus aureus enterotoxin B in Balb/c mice through intranasal infection.

Background and Aim: Staphylococcus aureus produces various superantigen exotoxins, including staphylococcal enterotoxin B (SEB). It causes fatal anaphylactic reactions and toxic shock. This study aimed to evaluate the reaction of leukocytes and histopathological changes in the respiratory organs of Balb/c mice after intranasal infection with enterotoxigenic S. aureus (SEB). Materials and Methods: The presence of the seb gene in S. aureus was established in this study using polymerase chain reaction-specific primer. Two groups of 8-week-old male Balb-c mice consist of six mice in each group. The treated group was infected with 50 µL and 100 µL of SEB intranasal on days 1 and 14, respectively. NaCl was administered in the second group and was considered as a control group. Blood samples were collected through the retro-orbital plexus on days 1, 4, 7, 14, and 22 after infections. Total cell counts were analyzed with an independent sample t-test and compared using the statistical package for the social sciences (SPSS) version 16.0 (IBM Corp., NY, USA). The infected tissues of the respiratory organ were observed descriptively and compared to the control group. Results: The seb gene with a molecular size of 478 bp, indicating the SEB strain, is present in S. aureus used in this study. Intranasal administration of SEB showed increased leukocytes, lymphocytes, monocytes, and eosinophils on day 22 post-infection. Significant leukocytosis was seen on days 6 and 14; lymphocytosis on days 1, 4, 6, and 16; and eosinophilia on days 6, 14, and 22 compared with the control group (p > 0.05). In contrast, the neutrophil decreased after an increase of immature band cells compared to the control group, indicating a severe acute infection with SEB. The lungs and trachea of the test group had an inflammatory cell accumulation in the respiratory organ. Conclusion: Intranasal route infection of S. aureus containing seb gene significantly induced the cellular immune response and caused pathological changes in the respiratory tissues of the Balb/c mice model. The hematological changes were aligned with marked pathological changes in the respiratory tract. Balb/c mice could be an excellent experimental model to study toxic and anaphylactic shock against SEB to define the future therapeutic agents.

Investigation of chlamydophilosis from naturally infected cats.

BACKGROUND: Chlamydophila felis, formerly known as Chlamydia psittaci var. felis, is frequently associated with ocular, respiratory, and occasionally reproduction tract infections. Even though the infection is sometimes asymptomatic, it potentially results in a latent immunosuppressive infection. OBJECTIVE: This study aimed to identify occurrences of feline chlamydophilosis, rarely reported in cats in Indonesia. METHODS: The observation was conducted in three cats with clinical signs of Cp. felis infection, particularly relapsing conjunctivitis. The cats' histories were recorded based on owners' information. Conjunctival swabs were sampled for cytology examination and molecular assay detection. A phylogenetic tree was generated using MEGA-X software to reveal group clustering. A post-mortem examination was performed on the cat that died during an examination. RESULTS: Cp. felis was detected in both cytological examination and polymerase chain reaction assay. The phylogenetic tree demonstrated that the Cp. felis isolated in this study clustered with several other isolates from the other countries. Cp. felis can be isolated from cats with different clinical manifestations and levels of severity. The chronic fatal infection demonstrated interstitial broncho-pneumonia under histopathological examination. CONCLUSIONS: Molecular assay of Cp. felis is always recommended to obtain a definitive diagnosis of feline chlamydophilosis since the disease can have various clinical manifestations. Even though it may be subclinical and is often not fatal, an infected cat may be a carrier that could spread the pathogen in the surrounding environment. Serious disease management is suggested to avoid high costs associated with regularly relapsing disease.

Streptococcus equi subsp. zooepidemicus finding in confirmed feline infectious peritonitis cat patient.

BACKGROUND: Feline infectious peritonitis (FIP) is a fatal immune-mediated disease in cat, caused by mutated feline coronavirus (FCoV). Due to its difficulties in diagnosis, FIP is sometimes underdiagnosed. Therefore, several laboratory procedures were performed to gain high index suspicion of FIP. However, through several laboratory findings, not only FIP but also SEZ infection was confirmed in this case. CASE DESCRIPTION: A-year-old male, domestic cat was admitted to Veterinary Medicine Clinical Pathology Laboratory, Universitas Gadjah Mada, for further effusion examination due to its high suspicion of feline infectious peritonitis (FIP). Further examination using molecular and post-mortem analysis resulted on confirmed SEZ infection and FIP. This study informed the manifestation and pathological changes in patient with SEZ and FIP in the same time. CONCLUSIONS: This study showed that viral infection followed by bacterial infection could be fatal and untreatable. After these findings, clinicians may consider SEZ infection in cat with respiratory disorder followed by thoracic effusion besides FIP. Companion animal, especially outdoor-kept animal, possibly become infected from its contact to another human or animal in the environment.